Schizosaccharomyces

J Biol Chem, 1987 Dec 25, 262(36), 17549 - 55
Mutation of a conserved glycine residue modifies the vanadate sensitivity of the plasma membrane H+-ATPase from Schizosaccharomyces pombe; Ghislain M et al.; The structural gene pma+1 for the H+-ATPase from the fission yeast Schizosaccharomyces pombe has been isolated and sequenced . The intron-less gene encodes for a protein of Mr = 99,769 which is 75% homologous to those of Saccharomyces cerevisiae and Neurospora crassa . The S . pombe pma+1 gene complements not only S . pombe pma-1-1 but also S . cerevisiae pma-1-4 mutants selected for in vitro vanadate-resistant ATPase activity . The sequence of the S . pombe mutant pma-1-1 allele reveals that the glycine residue 268, which is perfectly conserved in the transduction domain of all animal and fungal transport ATPases sequenced so far, is modified into an aspartate residue by the mutation . Replacement of glycine 268 by aspartate has been monitored by the appearance of a new PvuI restriction site in the mutant DNA . Mitotic cosegregation has been observed between the PvuI site and vanadate-resistant ATPase activity in a growing population of S . pombe transformants.

Philos Trans R Soc Lond B Biol Sci, 1987 Dec 15, 317(1187), 507 - 16
Control over the onset of DNA synthesis in fission yeast; Simanis V et al.; The fission yeast Schizosaccharomyces pombe has been used to identify gene functions required for the cell to become committed to the mitotic cell cycle and to initiate the processes leading to chromosome replication in S-phase . Two gene functions cdc2 and cdc10 must be executed for the cell to traverse 'start' and proceed from G1 into S-phase . Before the completion of these two functions the cell is in an uncommitted state and can undergo alternative developmental fates such as conjugation . A third gene, suc1, has also been identified whose product may interact directly with that of cdc2 at 'start' . The molecular functions of the genes involved in the completion of 'start' have been investigated . The cdc2 gene has been shown to be a protein kinase, suggesting that phosphorylation may be involved in the control over the transition from G1 into S-phase . The biochemical functions of the cdc10 and suc1 gene products have not yet been elucidated . A control at 'start' has also been shown to exist in the budding yeast Saccharomyces cerevisiae . Traverse of 'start' requires the execution of the CDC28 gene function . The cdc2 and CDC28 gene products (lower-case letters represent genes of Schizosaccharomyces pombe, and capital letters genes of Saccharomyces cerevisiae) are functionally homologous, suggesting that the processes involved in traverse of 'start' are highly conserved . An analogous control may also exist in the G1 period of mammalian cells, suggesting that the 'start' control step, after which cells become committed to the mitotic cell cycle, may have been conserved through evolution.

Eur J Biochem, 1987 Dec 15, 169(3), 527 - 37
The mitochondrial genome of the fission yeast, Schizosaccharomyces pombe . Sequence of the large-subunit ribosomal RNA gene, comparison of potential secondary structure in fungal mitochondrial large-subunit rRNAs and evolutionary considerations; Lang BF et al.; The DNA sequence of the mitochondrial large subunit (LSU) rRNA gene of Schizosaccharomyces pombe has been determined . In the direction of transcription, this gene is located between the gene coding for subunit II of cytochrome oxidase and a cluster of three tRNA genes . Both the 5' and 3' ends of the LSU rRNA have been mapped precisely: whereas the 5' end can be assigned unambiguously to a single nucleotide position, multiple 3' ends occur within a run of eight U residues . Based on these results, the S . pombe LSU rRNA is between 2818 and 2826 nucleotides long . A sequence motif immediately upstream of the 5' end of the gene resembles that of the mitochondrial promoter motif of Saccharomyces cerevisiae; however, the sequence at the 3' end of the gene is not similar to any of the motifs implicated as processing signals in other mitochondrial systems . Unlike its counterparts in S . cerevisiae and Aspergillus nidulans, the mitochondrial LSU rRNA gene of S . pombe does not contain an intron . Comparison of potential secondary structure among the three fungal mitochondrial and Escherichia coli LSU rRNAs has defined a common secondary structure core, held together by long-range hydrogen-bonding interactions . A 5.8S-like structure is present within the 5'-terminal region of all three fungal mitochondrial LSU rRNAs; in contrast, no 4.5S-like structure is evident at the 3' end of these molecules . An evolutionary evaluation of highly conserved regions of a small set of LSU rRNA sequences suggests that S . pombe mitochondria diverged from a mitochondrial proto-fungal branch earlier than either A . nidulans or S . cerevisiae mitochondria . This result, considered in conjunction with the patterns of genome organization and codon usage in fungal mitochondria, points to a slower evolutionary clock speed in the mitochondrial genome of S . pombe.

Nucleic Acids Res, 1987 Dec 10, 15(23), 9727 - 39
Cloning and sequencing of Schizosaccharomyces pombe DNA topoisomerase I gene, and effect of gene disruption; Uemura T et al.; We cloned the structural gene topl+ for Schizosaccharomyces pombe DNA topoisomerase I (topo I) by hybridization . An eight-fold increase of topo I relaxing activity was obtained in S . pombe cells transformed with multicopy plasmid with topl+ insert . Nucleotide sequence determination showed a hypothetical coding frame interrupted by two short introns, encoding a 812 residue polypeptide (M.W . 94,000), 43 residues longer than and 47% homologous to Saccharomyces cerevisiae topo I . We show that the topl (null) strain made by gene disruption is viable, although its generation time is 20% longer than that of wild type . The topl locus is mapped in the long arm of chromosome II, using the Leu+ marker integrated with the cloned topl+ sequence . We constructed a double mutant topl (null) top2 (ts) and found its defective phenotype similar to that of previously obtained topl (heat sensitive) top2 (ts) . The other double mutant topl (null) top2 (cs), however, was lethal . Our results suggest that topl+ gene of S . pombe is dispensable only if topo II activity is abundant.

Mol Gen Genet, 1987 Dec, 210(3), 485 - 9
Strains of Schizosaccharomyces pombe with a disrupted swi1 gene still show some mating-type switching; Schmidt H; The swi1+ gene is necessary for effective mating-type (MT) switching in Schizosaccharomyces pombe . It was cloned on a 4.2 kb genomic DNA fragment . By site-directed integration into the genome and gene disruption experiments it was proved that the swi1+ gene itself and not a suppressor had been isolated . Disruption of the swi1+ gene causes a phenotype identical to that of the original swi1 mutant, i.e . the strain still shows some MT switching . The swi1 gene is unique in the genome and gives rise to a 3 kb mRNA.

Mol Cell Biol, 1987 Dec, 7(12), 4424 - 30
Identification of healed terminal DNA fragments in linear minichromosomes of Schizosaccharomyces pombe; Matsumoto T et al.; The minichromosome Ch16 of the fission yeast Schizosaccharomyces pombe is derived from the centromeric region of chromosome III . We show that Ch16 and a shorter derivative, Ch12, made by gamma-ray cleavage, are linear molecules of 530 and 280 kilobases, respectively . Each minichromosome has two novel telomeres, as shown by genomic Southern hybridization with an S . pombe telomere probe . Comparison by hybridization of the minichromosomes and their chromosomal counterparts showed no signs of gross rearrangement . Cosmid clones covering the ends of the long arms of Ch16 and Ch12 were isolated, and subcloned fragments that contained the breakage sites were identified . They are apparently unique in the genome . By hybridization and Bal 31 digestion, the ends appear to consist of the broken-end sequences directly associated with short stretches (about 300 base pairs) of new DNA that hybridizes to a cloned S . pombe telomere . They do not contain the telomere-adjacent repeated sequences that are present in the normal chromosomes . The sizes of the short telomeric stretches are roughly the same as those of the normal chromosomes . Our results show that broken chromosomal ends in S . pombe can be healed by the de novo addition of the short telomeric repeats . The formation of Ch16 must have required two breakage-healing events, whereas a single cleavage-healing event in the long arm of Ch16 yielded Ch12.

Biochem Biophys Res Commun, 1987 Nov 13, 148(3), 1182 - 8
Revertant of the yeast Schizosaccharomyces pombe with modified alpha subunits of mitochondrial ATPase-ATPsynthase: impaired nucleotide interactions with soluble and membrane-bound enzyme; Falson P et al.; A partial revertant from a mutant with modified alpha subunits of mitochondrial ATPase-ATPsynthase has been obtained for the first time from the yeast Schizosaccharomyces pombe . The purified F1 contains a lower amount of endogenous nucleotides as compared to the wild-strain enzyme . In contrast to the wild-type, the F1 ATPase activity from the revertant does not exhibit bicarbonate-sensitive negative cooperativity . The revertant Michaelis constant for Mg-ATP is very similar to that of normal F1 in the presence of bicarbonate while the Vm is slightly lower . The revertant enzyme is much less sensitive to inhibitions by ADP and by azide . It is proposed that the lack of negative cooperativity of revertant F1 ATPase activity is due to lower affinity for ADP, the release of which is no longer the rate-limiting step.

Nature, 1987 Oct 15-21, 329(6140), 651 - 4
Similarity between cell-cycle genes of budding yeast and fission yeast and the Notch gene of Drosophila; Breeden L et al.; The HO gene of Saccharomyces cerevisiae encodes the endonuclease that initiates mating-type switching . To prevent inopportune switching, HO transcription is restricted to a specific period in the haploid cell cycle, which is just after, and dependent on, the start of the mitotic cell cycle . A repeated promoter element (CACGA4) (refs 7-9) and two trans-acting activators (SWI4 and SWI6) have been identified, which are responsible for the periodic and start-dependent transcription of HO . To understand further the link between start and HO transcription, the SWI6 gene has been cloned and sequenced . The SWI6 protein is similar to the protein in Schizosaccharomyces pombe that is encoded by cdc10 an essential gene specifically required at the start of the cell cycle . The similarity between the SWI6 and cdc10 products, and their common involvement with 'start', suggest that they may share a common mechanism for sensing or executing this critical control step in the cell cycle . The SWI6 and cdc10 proteins also contain two copies of a repeated motif that occurs at least five times in the cytoplasmic domain of the Notch protein of Drosophila melanogaster.

Nucleic Acids Res, 1987 Oct 12, 15(19), 7865 - 76
Resolution of DNA molecules greater than 5 megabases by contour-clamped homogeneous electric fields; Vollrath D et al.; Excellent resolution of chromosomal DNA molecules from Saccharomyces cerevisiae, Candida albicans and Schizosaccharomyces pombe has been obtained using alternating contour-clamped homogeneous electric field (CHEF) gel electrophoresis . The largest of these molecules is greater than 5 Mb in size and is resolved after 130 hours in a 0.6% agarose gel at a field strength of 1.3 V/cm and a switching interval of 1 hour . Separation of concatamers of phage lambda DNA reveals four regions of resolution in alternating CHEF gel electrophoresis . There are two regions of good resolution in which mobility approximates a linear function of molecular weight . These are separated by a region of lower resolution and bounded at high molecular weights by a region of little or no resolution . The four regions are of practical and possibly theoretical importance.

J Mol Biol, 1987 Oct 5, 197(3), 389 - 95
Nucleotide sequences of three tRNA(Ser) from Drosophila melanogaster reading the six serine codons; Cribbs DL et al.; The nucleotide sequences of three serine tRNAs from Drosophila melanogaster, together capable of decoding the six serine codons, were determined . tRNA(Ser)2b has the anticodon GCU, tRNA(Ser)4 has CGA and tRNA(Ser)7 has IGA . tRNA(Ser)2b differs from the last two by about 25% . However, tRNA(Ser)4 and tRNA(Ser)7 are 96% homologous, differing only at the first position of the anticodon and two other sites . This unusual sequence relationship suggests, together with similar pairs in the yeasts Schizosaccharomyces pombe and Saccharomyces cerevisiae, that eukaryotic tRNA(Ser)UCN may be undergoing concerted evolution.

Mol Gen Genet, 1987 Oct, 209(3), 627 - 9
Mapping of the ras1 gene of Schizosaccharomyces pombe; Lund PM et al.; The ras1 gene, an oncogene homologue, is known to be essential for recognition of the mating pheromone and hence for conjugation but not for vegetative growth in Schizosaccharomyces pombe . To facilitate further characterization and genetic manipulation of this gene, we have mapped it by using S . pombe strains which carry the Saccharomyces cerevisiae LEU2 gene inserted next to ras1 on the chromosome . Crosses with tester strains revealed that ras1 is tightly linked to pro2 on chromosome I . Furthermore, we have shown that ras1 is allelic with ste5, one of the sterility genes described by O . Girgsdies . The map position previously reported for ste5 eventually turned out to be false.

J Cell Sci, 1987 Oct, 88 ( Pt 3), 295 - 304
Schizosaccharomyces pombe mutants affected in their division response to starvation; Young PG et al.; Schizosaccharomyces pombe mutants have been selected on the basis of an altered response to nutritional stimulation of cell division (changed division response, cdr) . Two new loci (cdr1 and cdr2) were identified and characterized . When suspended in nitrogen-free medium wild-type cells underwent stimulated rates of division and became reduced to approximately 30% in protein content with a concomitant 3.6-fold increase in cell number after 24 h starvation . cdr cells had significantly smaller increases in cell number . The ratio of starved/unstarved protein content was higher for the cdr strains than for the wild type . cdr cells were also affected in their response to nitrogen-source shifts from proline to glutamate (or vice versa) or when shifted from serine phosphate to inorganic phosphate, showing that the alteration in division response was not restricted to nitrogen metabolism . Upon nitrogen starvation wild-type cells arrested prior to the cdc10 execution point, whereas cdr cells arrested later in the cell cycle . cdc25-22 cdr1 or cdr2 double mutants grew very slowly and were extremely elongated at all temperatures; the restrictive temperature was reduced to 27 degrees C . wee1 was epistatic to cdr mutations with respect to cell length at the cell plate stage . cdr+ genes are postulated to play a role in the nutritional modulation of the mitotic size control.

Nucleic Acids Res, 1987 Sep 25, 15(18), 7369 - 79
A single intronless action gene in the fission yeast Schizosaccharomyces pombe: nucleotide sequence and transcripts formed in homologous and heterologous yeast; Mertins P et al.; The actin gene of the fission yeast Schizosaccharomyces pombe has been isolated by using as a hybridization probe cloned actin DNA from the budding yeast Saccharomyces cerevisiae . In contrast to most actin genes studied from diverse eukaryotic species, the S . pombe gene is not interrupted by introns . The protein sequence deduced from the nucleotide sequence of the gene shows that the S . pombe actin is more closely related to the mammalian gamma-actin than to the actin of S . cerevisiae . Three transcripts of 1240, 1650 and 1850 nucleotides having the same 5' end but differing in the length of their 3' untranslated region are generated in the fission yeast . Only one messenger RNA of 1330 nucleotides is formed from the S . pombe actin gene in S . cerevisiae . Contrary to the observation made with other S . pombe genes transcribed in the budding yeast, the heterologous actin gene transcript is initiated 39 nucleotides upstream of the initiation start site used in the homologous yeast . The mRNA termination (or 3' processing) mechanism in the two ascomycetes also differs as the 3'end of the S . pombe actin gene transcript in S . cerevisiae does not coincide with either of the three 3'ends mapped in the fission yeast.

Cell, 1987 Sep 11, 50(6), 917 - 25
DNA topoisomerase II is required for condensation and separation of mitotic chromosomes in S . pombe; Uemura T et al.; We show that DNA topoisomerase II (topo II) is continuously required for mitotic chromosome changes in Schizosaccharomyces pombe . We constructed cold-sensitive (cs) or temperature-sensitive (ts) strains mutated in the genes coding for topo II (top2) and beta-tubulin (nda3) . The ATP-dependent activity of the top2cs gene product is cs in vitro . The cloned top2cs gene sequence predicts an amino acid substitution . A cs top2-cs nda3 double mutant at 20 degrees C shows long, entangled chromosomes, which condense and separate upon the shift to permissive temperatures . If spindle formation is prevented at permissive temperatures, the chromosomes condense but do not separate . Thus topo II is required for final chromosome condensation; moreover, pulse-shift experiments show that topo II is required for chromatid disjuction . Experiments with ts top2-cs nda3 cells show that topo II is also required for chromosome separation in anaphase: inactivation of topo II and activation of beta-tubulin allow normal spindle formation but result in "streaked" chromosomes.

FEBS Lett, 1987 Aug 10, 220(1), 27 - 30
Heterogeneous glycosylation of the EXG1 gene product accounts for the two extracellular exo-beta-glucanases of Saccharomyces cerevisiae; Nebreda AR et al.; Two exo-beta-glucanases of glycoprotein nature can be detected in culture supernatants of Saccharomyces cerevisiae cells . These exo-beta-glucanases show different Mr values and kinetic properties, although they are immunologically related . Their carbohydrate content and the electrophoretic mobility of both endoglycosidase H-treated exo-beta-glucanases suggest that they share the same protein fraction . Studies at genetic level relate the production of both extracellular exo-beta-glucanases with the expression of a single-copy gene in S . cerevisiae . Expression of this gene in another yeast, Schizosaccharomyces pombe, demonstrates that it codes for a protein with exo-beta-glucanase activity whose heterogeneous N-glycosylation accounts for both extracellular exo-beta-glucanases of S . cerevisiae.

Cell, 1987 Jul 31, 50(3), 391 - 403
A fission yeast chromosome can replicate autonomously in mouse cells; Allshire RC et al.; To test the functional capacity of a fission yeast chromosome in mouse cells, a strain of the fission yeast Schizosaccharomyces pombe, ED628 Int5, was constructed . A plasmid bearing the SV2NEO gene, which can confer G418 resistance to mouse cells, was integrated at the ura4 locus on S . pombe chromosome III . S . pombe Int5 chromosomes were introduced into mouse C127 cells by PEG-facilitated protoplast fusion . Here we describe two independent G418-resistant cell lines with distinct growth characteristics, F1.1 and F7.1, and examine the structure of material derived from S . pombe Int5 chromosome III in these lines . F1.1 is shown to contain a single rearranged block of chromatin from S . pombe chromosome III integrated into a mouse chromosome, maintained in the absence of selection . In contrast, the data for F7.1 are consistent with the presence of linear, unintegrated copies of S . pombe chromosome III, which are apparently intact and maintained in an unstable but autonomous state . The unstable maintenance of this chromosome may be due to defective centromere function leading to missegregation at mitosis or to over- or underreplication.

J Mol Biol, 1987 Jul 20, 196(2), 355 - 61
High level of complexity of small nuclear RNAs in fungi and plants; Tollervey D; The complexity of the trimethylguanosine-capped, small nuclear RNA (snRNA) populations in a number of organisms has been examined using immunoprecipitation and two-dimensional gels . From the fungi Aspergillus nidulans and Schizosaccharomyces pombe, over 30 major snRNAs can be resolved . The most abundant of these correspond to the putative analogues of vertebrate U1, U2, U4 and U5, which have been reported to be precipitated by anti-Sm antibodies, but other snRNAs are little less abundant than the major Sm-precipitable species . A similarly high level of complexity of snRNAs is detected in pea plants . In Candida albicans, the snRNAs are somewhat less numerous (about 22 major species) and are substantially less abundant than those of the above fungi, features shared with another budding yeast, Saccharomyces cerevisiae . Ten species of human snRNA have been reported; on two-dimensional gels, a number of additional snRNAs can be resolved from human cells . Each fungus, as well as pea plants, contains snRNAs substantially larger than any reported from vertebrates or detected in the human RNA used here . It appears that many eukaryotes contain substantially more species of snRNA than was previously believed.

Cell, 1987 Jul 17, 50(2), 319 - 25
Identification of p34 and p13, human homologs of the cell cycle regulators of fission yeast encoded by cdc2+ and suc1+; Draetta G et al.; cdc2+ and CDC28 play central roles in the cell division cycles of the widely divergent yeasts Schizosaccharomyces pombe and Saccharomyces cerevisiae, respectively . The genes encode protein kinases that show 62% protein sequence identity and are capable of cross-complementation . Monoclonal antibodies were raised against p34cdc2, and a subset recognize p36cdc28 . The cross-reacting antibodies detected a 34 kd homolog of the p34cdc2/p36CDC28, protein in HeLa cells . Human p34 was also recognized by an affinity-purified polyclonal anti-p34cdc2 serum . Peptide mapping of p34cdc2, p36CDC28, and human p34 revealed complete conservation of four tryptophan residues in the three proteins . p34 thus appears to be closely related to the two yeast proteins . In addition, a p34 immune complex showed protein kinase activity in vitro, and HeLa cell p34 interacts with p13, the human homolog of the suc1+ gene product of S . pombe.

Nucleic Acids Res, 1987 Jun 25, 15(12), 4705 - 15
A novel sequence common to the centromere regions of Schizosaccharomyces pombe chromosomes; Nakaseko Y et al.; An approximately 4 kb long sequence (designated dh) is located in the centromere regions of all three chromosomes of S . pombe . There is one copy each of dh per centromere in chromosomes I and II and multiples in the centromere of chromosome III . Nucleotide sequence determination shows that dhI and dhII are highly homologous . A part of the sequence (ca . 300-400 bp) contains short direct repeats, otherwise dh is in general internally non-repetitious . Although there are three segmental deletions (total 821 bp) and two insertions (27 bp) in dhII (an 80% overall homology to dhI), there are only nine substitutions between dhI and dhII in the remaining 3980 bp, giving a 99.77% homology . The substitutions are restricted to the non-repetitious domains and are only of the pyrimidine-pyrimidine or purine-purine types . A possible conformational role of dh is discussed.

Nucleic Acids Res, 1987 Jun 11, 15(11), 4481 - 9
An electrophoretic karyotype for Schizosaccharomyces pombe by pulsed field gel electrophoresis; Smith CL et al.; The three chromosomal DNAs of S . pombe have been fractionated by pulsed field gel electrophoresis . The resulting molecular karyotype will greatly speed gene mapping in this organism, and it indicates that the separation range of the technique extends to DNA molecules as large as 9,000,000 base pairs.

EMBO J, 1987 Jun, 6(6), 1757 - 63
Nuclear pre-mRNA splicing in the fission yeast Schizosaccharomyces pombe strictly requires an intron-contained, conserved sequence element; Mertins P et al.; It has recently been argued that pre-mRNA splicing in the fission yeast Schizosaccharomyces pombe may be more similar to splicing in metazoan species than in the budding yeast Saccharomyces cerevisiae . In this report we show that, contrary to this assumption, the conserved sequence element 5'-CTPu APy-3' found in all S . pombe introns 6-18 nucleotides upstream of the 3' splice site is, like the TACTAAC box in S . cerevisiae, indispensable for efficient splicing . The conserved adenine residue of this sequence is used for branch formation and point mutations introduced into the CTPuAPy sequence abolish splicing and seem not to result in the recruitment of cryptic branch sites . We also show that an S . cerevisiae intron is correctly excised in S . pombe whereby the TACTAAC box is used in branch formation.

Proc Natl Acad Sci U S A, 1987 Jun, 84(11), 3580 - 4
Analysis and in vivo disruption of the gene coding for calmodulin in Schizosaccharomyces pombe; Takeda T et al.; Calmodulin is a low molecular weight calcium-binding protein that modulates many enzyme systems in eukaryotes . We have cloned the gene encoding calmodulin from the fission yeast, Schizosaccharomyces pombe, by using synthetic oligonucleotide probes that correspond to three distinct regions of Tetrahymena calmodulin . A 1.6-kilobase (kb) DNA fragment that hybridized to all of them contains a gene whose deduced product possesses 74% amino acid homology with bovine calmodulin . This gene, which is unique in the S . pombe genome and is named cam1, encodes 149 amino acids excluding the first methionine and is transcribed into mRNA of 1.2-kb length . It has an intron that apparently starts immediately after the initiation codon and is 126 bp long . S . pombe calmodulin exhibits more homology to vertebrate calmodulin than to that of the budding yeast, Saccharomyces cerevisiae . Gene disruption experiments revealed that cam1 gene function is essential for vegetative growth of S . pombe . Spores bearing disrupted cam1 halt growth soon after germination and rarely carry out the first cell division, indicating that calmodulin does not exist in excess in those cells.

Proc Natl Acad Sci U S A, 1987 May, 84(9), 2585 - 9
Cloning and heterologous expression of glycosidase genes from Saccharomyces cerevisiae; Kuranda MJ et al.; Genomic clones were isolated that code for three glycosidases proposed to be involved in the catabolism of cell wall components in Saccharomyces cerevisiae . alpha-Mannosidase (AMS1), exoglucanase (BGL1), and endochitinase (CTS1) genes were isolated with the aid of filter assays based on the hydrolysis of 4-methylumbelliferyl glycosides, which permitted the in situ monitoring of these glycosidase activities in yeast colonies . Uracil prototrophs resulting from transformation with a multicopy YEp24 yeast genomic library were screened, leading to the identification of transformants possessing high levels of glycosidase activity . Restriction maps of plasmids from multiple isolates were used to localize glycosidase-overproduction genes, which were subcloned into a Schizosaccharomyces pombe/S . cerevisiae shuttle vector . Transformation of Sch . pombe with BGL1 and CTS1 subclones resulted in the appearance of these activities in this organism, and an AMS1 plasmid caused a 2-fold increase in endogenous alpha-mannosidase levels . Insertion of the marker gene LEU2 into putative AMS1 sequences disrupted plasmid-encoded alpha-mannosidase overproduction . S . cerevisiae strains that incorporated a restriction fragment containing ams1::LEU2 into their chromosomal DNA by homologous recombination expressed no detectable alpha-mannosidase activity in either the haploid or homozygous recessive diploid states, whereas heterozygous and wild-type cells exhibited levels proportional to AMS1 gene dosage . No readily apparent phenotype was associated with the alpha-mannosidase deficiency; however, labeling experiments utilizing {2-3H}mannose suggest that alpha-mannosidase may function in mannan turnover.

Mol Gen Genet, 1987 Apr, 207(1), 161 - 4
Characterisation of an autonomously replicating sequence from the fission yeast Schizosaccharomyces pombe; Johnston LH et al.; A DNA sequence has been isolated from Schizosaccharomyces pombe which promotes high frequency transformation of plasmids in the same organism . It is closely linked to the DNA ligase gene CDC17 and has therefore been named ARS17 although in structure it differs substantially from ARS elements in Saccharomyces cerevisiae . ARS17 spans some 1.8 kb of DNA and deletion of any part of this region affects activity . Moreover, there does not appear to be any short sequence which is, by itself, sufficient for high frequency transformation . ARS17 lies between and partly overlaps two divergently transcribed genes and it is extremely AT rich . It lacks the consensus sequence found in S . cerevisiae ARSs and it has no ARS activity in S . cerevisiae.

Microbiol Sci, 1987 Apr, 4(4), 115 - 8
Yeast tubulin genes; Yanagida M; There are two alpha-tubulin genes and one beta-tubulin gene in Schizosaccharomyces pombe and Saccharomyces cerevisiae . Detailed analyses employing tubulin mutants and cloned genes have revealed different cellular roles for the tubulin genes in these organisms.

J Biol Chem, 1987 Mar 15, 262(8), 3754 - 61
Kinetic characterization of yeast alcohol dehydrogenases . Amino acid residue 294 and substrate specificity; Ganzhorn AJ et al.; A three-dimensional model of yeast alcohol dehydrogenase, based on the homologous horse liver enzyme, was used to compare the substrate binding pockets of the three isozymes (I, II, and III) from Saccharomyces cerevisiae and the enzyme from Schizosaccharomyces pombe . Isozyme I and the S . pombe enzyme have methionine at position 294 (numbered as in the liver enzyme, corresponding to 270 in yeast), whereas isozymes II and III have leucine . Otherwise the active sites of the S . cerevisiae enzymes are the same . All four wild-type enzymes were produced from the cloned genes . In addition, oligonucleotide-directed mutagenesis was used to change Met-294 in alcohol dehydrogenase I to leucine . The mechanisms for all five enzymes were predominantly ordered with ethanol (but partially random with butanol) at pH 7.3 and 30 degrees C . The wild-type alcohol dehydrogenases and the leucine mutant had similar kinetic constants, except that isozyme II had 10-20-fold smaller Michaelis and inhibition constants for ethanol . Thus, residue 294 is not responsible for this difference . Apparently, substitutions outside of the substrate binding pocket indirectly affect the interactions of the alcohol dehydrogenases with ethanol . Nevertheless, the substitution of methionine with leucine in the substrate binding site of alcohol dehydrogenase I produced a 7-10-fold increase in reactivity (V/Km) with butanol, pentanol, and hexanol . The higher activity is due to tighter binding of the longer chain alcohols and to more rapid hydrogen transfer.

J Biol Chem, 1987 Mar 5, 262(7), 3146 - 53
Study of the nucleotide binding site of the yeast Schizosaccharomyces pombe plasma membrane H+-ATPase using formycin triphosphate-terbium complex; Ronjat M et al.; The plasma membrane of yeasts contains an H+-ATPase similar to the other cation transport ATPases of eukaryotic organisms . This enzyme has been purified and shows H+ transport in reconstituted vesicles . In the presence of Mg2+, formycin triphosphate (FTP) is hydrolyzed by the H+-ATPase and supports H+ transport . When combined with terbium ion, FTP (Tb-FTP) and ATP (Tb-ATP) are no longer hydrolyzed . Competition between Mg-ATP and Tb-FTP for ATP hydrolysis indicates that terbium-associated nucleotides bind to the catalytic site of the H+-ATPase . The fluorescent properties of the Tb-FTP complex were used to study the active site of the H+-ATPase . Fluorescence of Tb-FTP is greatly enhanced upon binding into the nucleotide site of H+-ATPase with a dissociation constant of 1 microM . Tb-ATP, Tb-ADP, and Tb-ITP are competitive inhibitors of Tb-FTP binding with Ki = 4.5, 5.0, and 6.0 microM, respectively . Binding of Tb-FTP is observed only in the presence of an excess of Tb3+ with an activation constant Ka = 25 microM for Tb3+ . Analysis of the data reveals that the sites for Tb-FTP and Tb3+ binding are independent entities . In standard conditions these sites would be occupied by Mg-ATP and Mg2+, respectively . These findings suggest an important regulatory role of divalent cations on the activity of H+-ATPase . Replacement of H2O by D2O in the medium suggests the existence of two types of nucleotide binding sites differing by the hydration state of the Tb3+ ion in the bound Tb-FTP complex.

J Cell Sci, 1987 Mar, 87 ( Pt 2), 323 - 5
Periodic cell cycle changes in the rate of CO2 production in the fission yeast Schizosaccharomyces pombe persist after a block to protein synthesis; Novak B et al.; CO2 production has been followed by manometry in synchronous and asynchronous control cultures of Schizosaccharomyces pombe prepared by elutriation from the same initial culture . Earlier results showed a periodic change in the rate of production, which took place once per cell cycle . These changes were most clearly shown as oscillations in the difference between values of the second differential (acceleration) for the synchronous and asynchronous cultures . This paper shows that the oscillations continue for at least three cycles in the presence of cycloheximide (with and without chloramphenicol) . Protein synthesis is virtually absent and there is no cell division . The control of this metabolic oscillation is therefore not directly dependent on translation . The period of the oscillation under these conditions is about 60% of the normal cycle time.

Nature, 1987 Mar 26-Apr 1, 326(6111), 414 - 6
Need for DNA topoisomerase activity as a swivel for DNA replication for transcription of ribosomal RNA; Brill SJ et al.; Yeast strains with mutations in the genes for DNA topoisomerases I and II have been identified previously in both Saccharomyces cerevisiae and Schizosaccharomyces pombe . The topoisomerase II mutants (top2) are conditional-lethal temperature-sensitive (ts) mutants . They are defective in the termination of DNA replication and the segregation of daughter chromosomes, but otherwise appear to replicate and transcribe DNA normally . Topoisomerase I mutants (top1), including strains with null mutations are viable and exhibit no obvious growth defects, demonstrating that DNA topoisomerase I is not essential for viability in yeast . In contrast to the single mutants, top1 top2 ts double mutants from both Schizosaccharomyces pombe and Saccharomyces cerevisiae grow poorly at the permissive temperature and stop growth rapidly at the non-permissive temperature . Here we report that DNA and ribosomal RNA synthesis are drastically inhibited in an S . cerevisiae top1 top2 ts double mutant at the restrictive temperature, but that the rate of poly(A)+ RNA synthesis is reduced only about threefold and transfer DNA synthesis remains relatively normal . The results suggest that DNA replication and at least ribosomal RNA synthesis require an active topoisomerase, presumably to act as a swivel to relieve torsional stress, and that either topoisomerase can perform the required function (except in termination of DNA replication where topoisomerase II is required).

Nucleic Acids Res, 1987 Feb 25, 15(4), 1477 - 92
Sequence and regulatory responses of a ribosomal protein gene from the fission yeast Schizosaccharomyces pombe; Nischt R et al.; We have determined the nucleotide sequence and mapped the 5' and 3' termini of a ribosomal protein gene . The gene is transcribed into a RNA molecule of about 770 nt and appears to initiate at multiple sites, as judged by SI nuclease analysis . Gene dosage experiments with a plasmid born gene leads to a proportional increase of the messenger RNA, but not to an overproduction of the protein, suggesting a posttranscriptional control mechanism . However, the heat shock response of this gene indicates that there is also a potential for transcriptional control . Comparison of the 5' flanking region of this gene with the ribosomal protein gene S 6 from Schizosaccharomyces pombe and with ribosomal protein genes from Saccharomyces cerevisiae revealed homologous sequences, which may be involved in the regulation of ribosomal protein genes.

FEBS Lett, 1987 Feb 23, 212(2), 323 - 7
Pi in equilibrium ATP exchange in the absence of proton gradient by the H+-ATPase from yeast plasma membranes; de Meis L et al.; Purified soluble H+-ATPase from Schizosaccharomyces pombe catalyzes a Pi in equilibrium ATP exchange in the absence of a H+ gradient . When the pH of the assay medium is raised from 5.5 to 8.0 there is a decrease of the ATPase activity and an increase of the rate of Pi in equilibrium ATP exchange . At pH 7.0 the addition of the organic solvent dimethyl sulfoxide (20%, v/v) promotes a decrease of ATPase activity and an increase of the Pi in equilibrium ATP exchange reaction . The effect of the organic solvent on the Pi in equilibrium ATP exchange is related to a decrease of the apparent Km for Pi.

Eur J Biochem, 1987 Feb 2, 162(3), 659 - 67
Molecular characterisation of the DNA ligase gene, CDC17, from the fission yeast Schizosaccharomyces pombe; Barker DG et al.; We have sequenced a 4200-base-pair fragment of Schizosaccharomyces pombe DNA which encompasses the entire DNA ligase gene, CDC17 . S1 mapping has enabled us to identify two small introns (40 and 62 nucleotides) at the 5' end of the coding region of the gene and their 3' internal conserved sequences match the CTRAY consensus found in other S . pombe introns . The major transcription initiation and 3' polyadenylation sites have been mapped and are preceded by higher eukaryotic-like TATA and AATAAA sequences respectively . Furthermore, the CDC17 mRNA carries a poly(A) tail whose length (approximately 250 nucleotides) is typical of that found in higher eukaryotic mRNAs, and is in contrast to the much shorter polyadenylated sequences found for the mRNAs of the budding yeast, Saccharomyces cerevisiae . The deduced amino acid sequence of the S . pombe DNA ligase predicts a protein of 86182 daltons, and an overall 53% homology with the same enzyme from S . cerevisiae . In particular, a stretch of 24 amino acids with 100% sequence homology spans the putative ATP-binding region which is also conserved in T4 and T7 bacteriophage DNA ligases.

EMBO J, 1987 Feb, 6(2), 469 - 76
Fungal small nuclear ribonucleoproteins share properties with plant and vertebrate U-snRNPs; Tollervey D et al.; snRNAs with properties closely related to those of the major vertebrate U-snRNAs are present in the fungi Aspergillus nidulans, Neurospora crassa and Schizosaccharomyces pombe . These RNAs possess a tri-methyl guanosine cap structure and a subset cross-hybridizes with human U1 and U2 clones . In the form of snRNPs, snRNAs from these fungi as well as from Saccharomyces cerevisiae and pea plants are immunoprecipitated by human and anti-Sm or anti-(U1)RNP autoimmune antibodies . On micro-injection into the cytoplasm of Xenopus oocytes, the snRNAs are packaged into ribonucleoprotein particles and migrate into the nucleus . The results demonstrate a hitherto unsuspected degree of evolutionary conservation in snRNA structure, snRNP protein structure, and sites of RNA-protein interaction within snRNPs.

J Biol Chem, 1987 Jan 5, 262(1), 223 - 8
A single mutation confers vanadate resistance to the plasma membrane H+-ATPase from the yeast Schizosaccharomyces pombe; Ulaszewski S et al.; A single-gene nuclear mutant has been selected from the yeast Schizosaccharomyces pombe for growth resistance to Dio-9, a plasma membrane H+-ATPase inhibitor . From this mutant, called pma1, an ATPase activity has been purified . It contains a Mr = 100,000 major polypeptide which is phosphorylated by {gamma-32P} ATP . Proton pumping is not impaired since the isolated mutant ATPase is able, in reconstituted proteoliposomes, to quench the fluorescence of the delta pH probe 9-amino-6-chloro-2-methoxy acridine . The isolated mutant ATPase is sensitive to Dio-9 as well as to seven other plasma membrane H+-ATPase inhibitors . The mutant H+-ATPase activity tested in vitro is, however, insensitive to vanadate . Its Km for MgATP is modified and its ATPase specific activity is decreased . The pma1 mutation decreases the rate of extracellular acidification induced by glucose when cells are incubated at pH 4.5 under nongrowing conditions . During growth, the intracellular mutant pH is more acid than the wild type one . The derepression by ammonia starvation of methionine transport is decreased in the mutant . The growth rate of pma1 mutants is reduced in minimal medium compared to rich medium, especially when combined to an auxotrophic mutation . It is concluded that the H+-ATPase activity from yeast plasma membranes controls the intracellular pH as well as the derepression of amino acid, purine, and pyrimidine uptakes . The pma1 mutation modifies several transport properties of the cells including those responsible for the uptake of Dio-9 and other inhibitors (Ulaszewski, S., Coddington, A., and Goffeau, A . (1986) Curr . Genet . 10, 359-364).

Mol Cell Biol, 1987 Jan, 7(1), 76 - 84
Substrate recognition and identification of splice sites by the tRNA-splicing endonuclease and ligase from Saccharomyces cerevisiae; Greer CL et al.; We have examined the substrate requirements for efficient and accurate splicing of tRNA precursors in Saccharomyces cerevisiae . The effects of Schizosaccharomyces pombe tRNASer gene mutations on the two steps in splicing, intron excision and joining of tRNA halves, were determined independently by using partially purified splicing endonuclease and tRNA ligase from S . cerevisiae . Two mutations (G14 and A46) reduced the efficiency of excision and joining in parallel, whereas two others (U47:7 and C33) produced differential effects on these two steps; U47:7 affected primarily the excision reaction, and C33 had a greater impact on ligation . These data indicate that endonuclease and ligase recognize both common and unique features of their substrates . Another two mutations (Ai26 and A37:13) induced miscutting, although with converse effects on the two splice sites . Thus, the two cutting events appear to be independent . Finally, we suggest that splice sites may be determined largely through their position relative to sites within the tRNA-like domain of the precursors . Several of these important sites were identified, and others are proposed based on the data described here.

Curr Genet, 1987, 12(8), 591 - 7
Sequence of the bifunctional ade1 gene in the purine biosynthetic pathway of the fission yeast Schizosaccharomyces pombe; McKenzie R et al.; The ade1 gene of the fission yeast Schizosaccharomyces pombe encodes a bifunctional polypeptide with glycinamide ribotide synthetase (GARSase) and aminoimidazole ribotide synthetase (AIRSase) enzyme activities . These enzyme activities carry out the 2nd and 5th steps, respectively, of the purine synthetic pathway . We report the cloning of the ade1 gene on a 4.4 kb Sau3A insert in the yeast shuttle vector pWH5 . Integration of this genomic insert at or near the ade1 locus and its ability to complement, by transformation, three different types of ade1 mutants proved that it contains the ade1 chromosomal gene . Analysis of the nucleotide sequence of this insert revealed the presence of an uninterrupted open reading frame of 2,367 pb . This sequence, and the predicted 789 amino acid sequence encoded, both show a high degree of homology with the functionally equivalent ade5,7 gene sequence of Saccharomyces cerevisiae (approx . 60% overall in both cases) and Gart gene sequences of Drosophila melanogaster . The size of the ade1 RNA transcript is about 2.7 kb.

Curr Genet, 1987, 12(5), 329 - 36
Distribution of mitochondrial introns in the species Schizosaccharomyces pombe and the origin of the group II intron in the gene encoding apocytochrome b; Zimmer M et al.; The mitochondrial genome size of 26 different Schizosaccharomyces pombe strains varies between 17.6 and 24.6 kilobase pairs due to the presence or absence of introns . One of these is the group II intron in the gene encoding apocytochrome b (cob: intron cobI1) . Partial DNA sequences of continuous cob genes from six strains (including strain EF1: Trinkl et al . 1985) revealed identical nucleotide sequence in the region where the group II intron is inserted in the mosaic form of the gene . In contrast, analysis of the mosaic cob gene in strain UCD-FstI revealed several base pair changes in the exon regions flanking the splice point, compared with the continuous genes and with the mosaic cob gene in strain 50 (Lang et al . 1985) . The base pair differences between the exons of the two mosaic cob genes and the identity of exons in all continuous cob genes argue in favour of the two cob introns in strains 50 and UCD-FstI as independent later acquisitions of the genes, rather than loss of the intron from a common mosaic ancestor of all strains . Other introns present in some but not all strain include two group I introns without open reading frame in the gene encoding subunit 1 of cytochrome c oxidase (cox1: introns cox1I2a and cox1I3), and two group I introns with open reading frames in the same gene (introns cox1I1 and cox1I2b).

Gene, 1987, 60(2-3), 157 - 61
The RNA components of Schizosaccharomyces pombe RNase P are essential for cell viability; Cherayil B et al.; The fission yeast Schizosaccharomyces pombe contains in the haploid genome one copy of the gene (designated rrkl) for the RNA components of RNase P . Gene disruption in diploid cells of one copy of rrkl resulted in a moderate reduction of the level of cellular RNase P activity . Haploidization by meiosis demonstrated that rrkl is required for cell growth . Thus, the RNA components of S . pombe RNase P are essential in vivo . This is similar to the situation in Escherichia coli.

Curr Genet, 1987, 11(8), 575 - 89
Genetic nomenclature and gene list of the fission yeast Schizosaccharomyces pombe; Kohli J; The nomenclature rules for the genetics of the fission yeast Schizosaccharomyces pombe have been fixed for the first time, after discussion among scientists working with this organism . Conventions are proposed for the naming of genes and alleles that are obtained by classical means or by reverse genetics . In addition a list has been compiled of 460 known genes of S . pombe . It includes genes defined both by classical mutation analysis and by molecular cloning . 270 genes have been assigned either to one of the three nuclear chromosomes or the mitochondrial genome.

Gene, 1987, 58(1), 59 - 66
The unique histone H2A gene of Aspergillus nidulans contains three introns; May GS et al.; The histone H2A gene of the filamentous fungus Aspergillus nidulans has been cloned and sequenced . There is a single H2A gene in the genome of A . nidulans, and it contains three introns . The introns are 51 nucleotides (nt), 56 nt and 50 nt in length and split codons for amino acids (aa) 18, 48 and 116 of the predicted protein . The transcriptional start and termination points have been determined using an S1 nuclease protection assay . The predicted protein is 132 aa residues in length and surprisingly has a threonine after the initiator methionine instead of the usual serine . The sequence of the predicted histone H2A protein is compared to histone H2A proteins from Schizosaccharomyces pombe, Saccharomyces cerevisiae and calf thymus . Comparison of the amino acid sequence to these other H2A proteins shows that the divergence of amino acid sequences between H2A proteins is found in two clustered sites.

Mol Cell Biol, 1987 Jan, 7(1), 504 - 11
Sucl+ encodes a predicted 13-kilodalton protein that is essential for cell viability and is directly involved in the division cycle of Schizosaccharomyces pombe; Hindley J et al.; Sucl+ was originally identified as a DNA sequence that, at high copy number, rescued Schizosaccharomyces pombe strains carrying certain temperature-sensitive alleles of the cdc2 cell cycle control gene . We determined the nucleotide sequence of a 1,083-base-pair Sucl+ DNA fragment and S1 mapped its 866-nucleotide RNA transcript . The protein-coding sequence of the gene is interrupted by two intervening sequences of 115 and 51 base pairs . The predicted translational product of the gene is a protein of 13 kilodaltons . A chromosomal gene disruption of Sucl+ was constructed in a diploid S . pombe strain . Germinating spores carrying a null allele of the gene were capable of very limited cell division, following which many cells became highly elongated . The Sucl+ gene was also strongly overexpressed under the control of a heterologous S . pombe promoter . Overexpression of Sucl+ is not lethal but causes a division delay such that cells are approximately twice the normal length at division . These data suggest that Sucl+ encodes a protein which plays a direct role in the cell division cycle of S . pombe.

J Bacteriol, 1987 Jan, 169(1), 93 - 6
Cloning and analysis of transcription of the mei2 gene responsible for initiation of meiosis in the fission yeast Schizosaccharomyces pombe; Shimoda C et al.; We have isolated a hybrid plasmid, pDB(mei2)2, containing a 7.4-kilobases (kb) DNA fragment from a Schizosaccharomyces pombe genomic library which is able to complement the mei2 mutation of S . pombe . Integration of the cloned DNA sequence at the mei2 site on chromosome I demonstrated that it contained the mei2 gene . This gene was localized on a 4.7-kb HindIII-PvuII fragment in the subclone pFMV402 . Transcriptional regulation was studied by Northern blot analysis in which polyadenylated RNA was prepared from a heterozygous (h+N/h-S) diploid strain cultured either in nitrogen-rich growth medium or in nitrogen-free sporulation medium . The size of the major mei2 mRNA, which always gave a broad band, was estimated to be 4.2 +/- 0.2 kb, and a few minor bands (e.g., 3.2 and 1.8 kb) appeared as well . These transcripts appeared more abundantly in sporulating cells than in growing cells . Neither the mating type genes (mat) nor the mei3 gene was essential for transcription of the mei2 gene, since ample mei2 mRNA was detected in sporulation-deficient cells transferred to sporulation medium, such as h+N/h+N and h-S/h-S homozygotes, as well as mei1 and mei3 mutants.

Proc Natl Acad Sci U S A, 1987 Jan, 84(2), 388 - 92
Homology probing: identification of cDNA clones encoding members of the protein-serine kinase family; Hanks SK; Mixed oligonucleotide probes were used to screen a HeLa cDNA library for clones encoding amino acid contiguities whose conservation is characteristic of the protein-serine kinase family . Eighty thousand clones were screened, from which 19 were identified as showing strong hybridization to two distinct probes . Four clones were chosen for characterization by partial DNA sequence analysis and 3 of these were found to encode amino acid sequences typical of protein-serine kinases . One deduced amino acid sequence shares 72% identity with rabbit skeletal muscle phosphorylase kinase gamma-subunit, while another is closely related to the yeast protein-serine kinases CDC2 in Schizosaccharomyces pombe and CDC28 in Saccharomyces cerevisiae . This screening approach should have applications in the identification of clones encoding previously unknown or poorly characterized members of other protein families.

Curr Genet, 1987, 12(7), 527 - 34
Cloning and expression of the OMP decarboxylase gene URA4 from Schizosaccharomyces pombe; Bach ML; URA4, the gene coding for orotidine monophosphate decarboxylase (OMPdecase), has been cloned from the fission yeast by homologous complementation and restricted in an Escherichia coli-Schizosaccharomyces pombe (E . coli-S . pombe) replicative plasmid to a 1.76 kb HindIII fragment . This plasmid is maintained at a high copy number in S . pombe and allows OMPdecase expression in Saccharomyces cerevisiae (S . cerevisiae) as well as in E . coli . After characterisation by restriction mapping and Southern hybridisation, the cloned gene was used as a probe to measure URA4 transcription and to examine its regulation . Messenger RNA levels were measured by DNA/RNA filter-hybridisation with pulse labelled RNAs during 6-azauridine (6-AUR) inhibited growth in wild type and 6-AUR sensitive strains . We found that in S . pombe the OMP analogue 6-AUR does not regulate the level of OMPdecase formation as it does in S . cerevisiae but rather modifies the ratio of total polyA+ to polyA- RNAs in the cell . Based on these results and on corresponding enzyme activities this study demonstrates divergent pyrimidine pathway regulation in the two yeasts S . cerevisiae and S . pombe . Finally, we propose the use of the URA4 gene as a convenient selective marker for genetic engineering in S . pombe.

NCI Monogr, 1987, (4), 7 - 10
Molecular genetic analysis of topoisomerase II gene from Drosophila melanogaster; Hsieh TS et al.; The gene encoding the Drosophila type II DNA topoisomerase has been isolated and characterized . The enzyme is coded for by a messenger RNA of about 5000 nucleotides, the expected size for the enzyme with MWr 170,000, and the gene is interrupted by four introns . The cytogenetic location of this gene has been mapped to the left arm of chromosome 2 at 37D2-6 by both in situ hybridization to polytene chromosomes and genomic blot-hybridization . The complete nucleotide sequence of the coding region and the flanking sequence has been determined . The deduced amino acid sequence shows interesting homology with topoisomerases II from bacteria, bacteriophage T4, and yeasts . The sequence homology between the Drosophila enzyme and those from Saccharomyces and Schizosaccharomyces is very extensive . This, in combination with the similar biochemical properties among all the eucaryotic DNA topoisomerases II, indicates that these enzymes are conserved during the course of evolution.

EMBO J, 1986 Dec 20, 5(13), 3665 - 71
Homology between the ran1+ gene of fission yeast and protein kinases; McLeod M et al.; The ran1+ gene of the fission yeast Schizosaccharomyces pombe is a negative regulator of both sexual conjugation and meiosis . The nucleotide sequence of the gene has been determined and contains a region of open reading frame (ORF) capable of encoding a protein of 52,000 daltons . S1 nuclease analysis of ran1+-encoded RNA showed that the ORF was spanned by an uninterrupted transcript . A fragment of DNA containing the entire ran1+ gene was expressed in a bacterial expression vector and found to encode the expected product of 52,000 daltons . The putative ran1+ gene product shares significant sequence homology with known protein kinases . The level of the ran1+ transcript was similar in vegetative and meiotic cells suggesting that the ran1+ protein product rather than its transcript is regulated during sexual differentiation.

J Biol Chem, 1986 Dec 15, 261(35), 16351 - 5
Antisuppressor mutations and sulfur-carrying nucleosides in transfer RNAs of Schizosaccharomyces pombe; Grossenbacher AM et al.; Antisuppressor mutations reduce the efficiency of nonsense suppressors . A mutation in the gene sin4 of Schizosaccharomyces pombe leads to loss of 5-(methoxycarbonylmethyl) thiouridine (mcm5s2U) from the first anticodon position of tRNAs . This resembles the phenotype of sin3 (Heyer, W . D., Thuriaux, P., Kohli, J., Ebert, P., Kersten, H., Gehrke, C., Kuo, K . C., and Agris, P . F . (1984) J . Biol . Chem . 259, 2856-2862), but the mutations reside in different genes . In vivo 35S-labeled tRNA from the parental suppressor strain sup3, the antisuppressor strains sin3 and sin4, and the double mutant sin3 sin4 has been digested to nucleosides and analyzed with high performance liquid chromatography methods . The major sulfur-carrying nucleoside in wild-type S . pombe tRNA is mcm5s2U . It is reduced in the mutant strains . Two other thiolated nucleosides are also present: 2-thiouridine and a nucleoside of unknown structure . Neither was affected by the antisuppressor mutations . Thiocytidine has not been found . Independent from their effect on suppressors, the two mutations sin3 and sin4 reduce the growth rate of cells, and sin3 also increases cell length . In vivo decoding of the serine codon UCG by the UCA reading serine tRNA is not promoted by the two antisuppressor mutations.

J Biol Chem, 1986 Dec 5, 261(34), 15877 - 82
Identification and characterization of thiamin repressible acid phosphatase in yeast; Schweingruber ME et al.; We have identified a genetic locus, pho4, in Schizosaccharomyces pombe which encodes a minor expressed cell surface acid phosphatase that is repressed by low concentrations (0.5 microM) of thiamin . The enzyme was purified from a strain that overproduces the enzyme . It is an Asn-linked glycoprotein . Removal of the carbohydrates by endoglycosidase H does not abolish enzymatic activity . The molecular mass of deglycosylated and unglycosylated enzyme that accumulates in membranes when cells are grown in the presence of tunicamycin is 56 kDa as determined by sodium dodecyl sulfate-gel electrophoresis . Thiamin regulation, at least in part, operates by reducing the level of pho4-mRNA . Pho4 is not genetically linked to the phosphate repressible acid phosphatase gene pho1 . Phosphate and thiamin repressible acid phosphatase differ in their substrate specificity . Their protein moieties are immunologically related . Pho4 and pho1 are the only genes in S . pombe that express cell surface acid phosphatases being enzymatically active with nitrophenyl phosphate as substrate . S . pombe is not unique in having a thiamin repressible acid phosphatase . In Saccharomyces cerevisiae this enzyme is encoded by PHO3.

J Bacteriol, 1986 Dec, 168(3), 1439 - 43
Malate transport in Schizosaccharomyces pombe; Osothsilp C et al.; The transport of malate was studied in a Schizosaccharomyces pombe wild-type strain and in mutant strains unable to utilize malic acid . Two groups of such mutants, i.e., malic enzyme-deficient and malate transport-defective mutants, were differentiated by a 14C-labeled L-malate transport assay and by starch gel electrophoresis followed by activity staining for malic enzyme (malate dehydrogenase {oxaloacetate decarboxylating} {NAD+}; 1.1.1.38) and malate dehydrogenase (1.1.1.37) . Transport of malate in S . pombe was constitutive and strongly inhibited by inhibitors of oxidative phosphorylation and of the formulation of proton gradients . Transport was a saturable function of the malate concentration . The apparent Km and Vmax values for transport by the parent were 3.7 mM and 40 nmol/min per mg of protein, respectively, while those of the malic enzyme-deficient mutant were 5.7 mM and 33 nmol/min per mg of protein, respectively . Malate transport was pH and temperature dependent . The specificity of transport was studied with various substrates, including mono- and dicarboxylic acids, and the possibility of a common transport system for dicarboxylic acids is discussed.

J Cell Sci, 1986 Dec, 86, 191 - 206
Change in the rate of CO2 production in synchronous cultures of the fission yeast Schizosaccharomyces pombe: a periodic cell cycle event that persists after the DNA-division cycle has been blocked; Novak B et al.; CO2 production has been followed by manometry in synchronous and asynchronous cultures of Schizosaccharomyces pombe prepared by elutriation from the same initial culture . The rate of production follows a linear pattern in synchronous cultures with a rate change once per cycle at the time of cell division . This pattern is most clearly shown in oscillations of the difference between values of the second differential (acceleration) for the synchronous and asynchronous cultures . The association between the rate change and the time of division is maintained during growth speeded up in rich medium and slowed down in poor medium and at lower temperature . It is also maintained after a shift-up in temperature . Results with wee mutants suggest that the association is with the S period rather than division itself . The rate and acceleration of CO2 production are approximately proportional to cell size (protein content) in asynchronous cultures . When synchronous cultures of the temperature-sensitive mutants cdc2.33 and cdc2.33 wee1.6 are shifted up to the restrictive temperature, the DNA-division cycle is blocked . The oscillatory pattern of CO2 production, however, continues for one to two cycles until the acceleration reaches a constant value, after which the oscillations are undetectable . This point is reached later in the double mutant and there is a phase difference in the oscillations compared to those in the single mutant . With both blocked mutants the 'free-running' oscillations are about 15% shorter than the normal cycle time . There are well-known examples of such oscillations in eggs but they are rare in growing systems.

J Cell Sci, 1986 Dec, 86, 207 - 15
Nucleoside diphosphokinase, an enzyme with step changes in activity during the cell cycle of the fission yeast Schizosaccharomyces pombe . I . Persistence of steps after a block to the DNA-division cycle; Creanor J et al.; In confirmation of earlier results, nucleoside diphosphokinase is shown to be a 'step' enzyme in Schizosaccharomyces pombe with a sharp doubling in activity at the beginning of the cell cycle . These doubling steps occur at the same time in the cycle in the smaller cells of the mutant wee1.6 . An important result is that the activity steps persist with normal cell cycle timing after a block to the DNA-division cycle imposed by the cycle mutants cdc2.33 and cdc2.33wee1.6 . This is clear proof that oscillatory controls of some cell cycle events can persist after the main periodic events of the DNA-division cycle have been abolished.

Proc Natl Acad Sci U S A, 1986 Nov, 83(21), 8253 - 7
Analysis of centromeric DNA in the fission yeast Schizosaccharomyces pombe; Clarke L et al.; The Schizosaccharomyces pombe centromere-linked genes, LYS1 and CYH1 on chromosome I and TPS13 and RAN1 on chromosome II, have been isolated . The genetic order of these markers with respect to their centromeres was determined to establish relative directionality on the genetic and physical maps . Chromosome walking toward the centromeres reveals a group of repetitive sequences that occur only in the centromere regions of chromosomes I and II and at one other specific location in the S . pombe genome, presumably the centromere of chromosome III . The major class of large repeated sequence elements is 6.4 kilobases (kb) long (repeat K), portions of which occur at least twice on chromosome II and in several tandemly arranged intact copies at another centromeric location . Repeat K in turn contains groups of smaller repeats . Genetic recombination is strongly suppressed in the centromere II region, which contains at least 30 kb of repeated sequences . Centromeric DNA organization is much more complex in fission yeast than has been described in budding yeast (Saccharomyces cerevisiae), possibly because of the larger more condensed nature of the S . pombe chromosomes.

Cell, 1986 Oct 10, 47(1), 49 - 59
U2 RNA from yeast is unexpectedly large and contains homology to vertebrate U4, U5, and U6 small nuclear RNAs; Ares M Jr; I have determined the structure of the gene from Saccharomyces cerevisiae coding for the yeast homolog of vertebrate U2 snRNA . Surprisingly, the RNA is 1175 nucleotides long, six times larger than U2 RNAs from other organisms, including Schizosaccharomyces pombe . Nearly 100 nucleotides of the large RNA share sequence homology and potential secondary structure with metazoan U2 . The large RNA also contains homology to vertebrate U4, U5, and U6 snRNAs, implying a "poly-snRNP" structure for the RNP containing the large RNA . The gene LSR1, encoding the large RNA, is essential for growth, suggesting that the yeast spliceosome can be dissected using genetic approaches . The different organization of spliceosomal RNA may underlie differences in splicing between yeast and metazoans.

Mol Cell Biol, 1986 Oct, 6(10), 3523 - 30
Site-specific mutagenesis of cdc2+, a cell cycle control gene of the fission yeast Schizosaccharomyces pombe; Booher R et al.; The cdc2+ gene of Schizosaccharomyces pombe is homologous to the CDC28 gene of Saccharomyces cerevisiae . Both genes share limited homology with vertebrate protein kinases and have protein kinase activity . cdc2+ has been subjected to mutagenesis in vitro . A null allele of the gene, constructed by insertion of the S . cerevisiae LEU2 gene into a site within the gene, has a phenotype similar to that of many temperature-sensitive alleles of cdc2 . Mutations within the predicted ATP-binding site and in a region which may be a site of phosphorylation result in loss of cdc2+ activity . A single substitution of Gly-146 to Asp-146 has been identified in cdc2-1w, a dominant activated allele of the gene . The four introns within the cdc2+ gene have been deleted . The resulting gene not only functions in fission yeast but also rescues cdc28(Ts) strains of S . cerevisiae, a property which is not shared by the genomic cdc2+ gene.

Proc Natl Acad Sci U S A, 1986 Oct, 83(20), 7860 - 4
Functional complementation between mutations in a yeast suppressor tRNA gene reveals potential for evolution of tRNA sequences; Willis I et al.; Successive rounds of mutagenesis of a Schizosaccharomyces pombe strain bearing the UGA-reading sup3 tRNASer suppressor have been carried out for two cycles of inactivation and reactivation of the suppressor . The suppressor phenotype at each stage was found to involve different combinations of three mutations, A30, A53, and A67, in the sup3-UGA gene . Single mutations A30 and A53 inactivate the suppressor as does the presence of all three mutations . A67 by itself is phenotypically neutral, but in combination with either A30 or A53 suppressor function is restored . The frequency with which these and other complementation events occur in S . pombe demonstrates a significant potential for nucleotide sequence evolution in tRNA . Differential expression of the S . pombe genes in Saccharomyces cerevisiae suggests that the two yeasts have diverged at the transcriptional and RNA processing level . Processing of the mutant tRNA precursors in S . cerevisiae reveals a hierarchy of structural domains within the tRNA that vary in their importance for RNase P cleavage.

FEBS Lett, 1986 Sep 29, 206(1), 135 - 41
Immunological cross-reactivity of fungal and yeast plasma membrane H+-ATPase; Vai M et al.; The plasma membrane H+-ATPases from fungi and yeasts have similar catalytic and molecular properties . A structural comparison has been performed using immunoblot analysis with polyclonal antibodies directed toward the 102 kDa polypeptide of the plasma membrane H+-ATPase from Neurospora crassa . A strong cross-reactivity is observed between the fungal H+-ATPase and the enzyme from the yeasts Saccharomyces cerevisiae and Schizosaccharomyces pombe . Structural homologies are indicated also by the analysis of the cross-reactive peptides originated by proteolytic digestion of Neurospora and S . cerevisiae purified enzymes . Neither enzyme from these two sources appears to be glycosylated by a highly sensitive concanavalin A affinity assay on blotted proteins . A glycoprotein of Mr 115000 and pI 4.8-5, which comigrates with a cell cycle-modulated protein on 2D gel, is present in partially purified preparations of plasma membrane H+-ATPase of S . cerevisiae and it is shown to be structurally unrelated to H+-ATPase.

EMBO J, 1986 Sep, 5(9), 2355 - 61
The nucleotide sequence of the fission yeast DNA topoisomerase II gene: structural and functional relationships to other DNA topoisomerases; Uemura T et al.; We have determined the complete nucleotide sequence of a 5.3-kb long genomic DNA fragment of the fission yeast Schizosaccharomyces pombe that encodes DNA topoisomerase II . It contains a 4293 bp long single open reading frame . The predicted polypeptide has 1431 residues (mol . wt 162,000) and shows three characteristic domains; the large C-terminal region, which consists of alternating acidic-basic stretches and might be a chromatin-binding domain, the NH2 half domain homologous to the ATP-binding gyrB subunit of bacterial gyrase and the central-to-latter part which is homologous to the NH2 domain of the catalytic gyrA subunit, suggesting a possible evolutionary consequence of the gene fusion of the bacterial gyrase subunits into the eucaryotic DNA topoisomerase II gene . We have found that the cloned fission yeast TOP2 gene can complement the budding yeast top2 mutation, although the fission yeast TOP2 protein sequence is only 50% homologous to the recently determined sequence of budding yeast (J.C . Wang, personal communication) . Conversely, the budding yeast TOP2 gene can complement the fission yeast top2 mutations, indicating that their DNA topoisomerase II genes are functionally exchangeable.

Cell, 1986 Aug 29, 46(5), 725 - 31
Initiation of meiotic recombination by double-strand DNA breaks in S . pombe; Klar AJ et al.; Mitotic gene conversion and reciprocal recombination have recently been shown to be efficiently initiated by double-strand DNA breaks (DSBs) in both Saccharomyces cerevisiae and Schizosaccharomyces pombe . We tested whether DSBs could also initiate meiotic recombination at the mat1 locus in S . pombe . The mat1 switching-mechanism-generated DSB found in mitotically growing cells can be repaired without mat1 switching, since strains deleted for both donor loci (mat2-P and mat3-M) have the break but do not produce inviable cells . A (mat1-P X mat1-M) cross produced a high frequency (20%) of 3:1 gene conversions of mat1 in meiotic tetrads . Gene conversion events were associated with the recombination of flanking markers . Strains lacking the DSB failed to convert . Thus, the DSB at mat1 promotes efficient meiotic recombination in fission yeast.

Microbiol Sci, 1986 Aug, 3(8), 234 - 7
Regulation of meiosis in Schizosaccharomyces pombe; Yamamoto M; Analysis of a new class of meiotic mutants isolated in the fission yeast Schizosaccharomyces pombe strongly indicates that the gene in which they are deficient codes for a factor whose physiological role is inhibition of initiation of meiosis . A negative control mechanism for meiosis is discussed.

EMBO J, 1986 Jul, 5(7), 1697 - 703
Two RNA species co-purify with RNase P from the fission yeast Schizosaccharomyces pombe; Krupp G et al.; RNase P activity from Schizosaccharomyces pombe co-purifies with two RNA species . These RNAs are associated with enzyme activity as judged by titrated micrococcal nuclease inactivation experiments . The two RNAs, K1- and K2-RNA, are 285 and 270 nucleotides long, respectively . Both RNAs are transcribed from one gene, present in a single copy in the haploid genome . The primary and a secondary structure of K RNAs have been determined and compared with M1 RNA, their counterpart from Escherichia coli . Very limited sequence homology was observed, and this agrees with the finding that no cross-hybridization with M1 RNA can be detected in a Southern analysis with yeast genomic DNA . However, the secondary structures of K RNA and M1 RNA show the same basic organization and one conserved local motif, the sequence GUG--AGGPu in an exposed hairpin loop.

Eur J Biochem, 1986 Jul 1, 158(1), 133 - 40
Glycosylation and secretion of acid phosphatase in Schizosaccharomyces pombe; Schweingruber AM et al.; We have purified secreted acid phosphatase of Schizosaccharomyces pombe . The enzyme is N-glycosylated, the associated carbohydrate accounts for 90% of the total molecular mass and the protein moiety has a molecular mass of 54 kDa . The deglycosylated enzyme still exhibits enzymatic activity . Using antibodies recognizing the protein moiety of the enzyme we have identified two intracellular precursors of acid phosphatase: an unglycosylated membrane-bound 54-kDa form that accumulates in the presence of tunicamycin and a partially glycosylated 72-kDa form that accumulates mostly in membranes of cells grown in rich medium . We further showed that the conversion of the 54-kDa and 72-kDa forms to partially glycosylated and fully glycosylated acid phosphatase is a regulated process . Growth conditions determine how much of translated 54-kDa acid phosphatase is glycosylated to the 72-kDa form and how much remains unglycosylated in membranes . When cells are grown in a rich medium, 5% of the total acid phosphatase protein remains as unglycosylated enzyme and 8% as partially glycosylated 72-kDa form . In cells grown in the minimal medium, however, all of the 54-kDa and 72-kDa forms of acid phosphatase are rapidly processed to fully glycosylated enzyme . The 72-kDa form and the unglycosylated form of acid phosphatase are not secreted or transported to the plasma membrane.

Exp Cell Res, 1986 Jul, 165(1), 243 - 54
A second growth state for Schizosaccharomyces pombe; Kubitschek HE et al.; The kinetics of volume increase in individual cells of Schizosaccharomyces pombe were determined by phase microscopy at osmolalities lower than those reported in the literature . At the highest osmolality, 550 mmol/kg, all cells followed a biphasic pattern of growth, in which cell volumes increased to their maximum values approximately four-fifths of the way through the growth cycle . At lower osmolalities (400-420 mmol/kg), many or most of the cells followed a different growth pattern, with a linear increase in cell volume throughout the cycle . The following evidence indicates that a different regulatory mechanism is responsible for the linear growth pattern: (1) Regulation of cell length and diameter differed for the two cases . During biphasic growth, cell length also increased biphasically and cell diameters remained essentially constant during the cycle, whereas during linear growth, both cell length and diameter increased linearly until formation of the cell plate very late in cycle . (2) The two different growth states were observed for cells growing on two very different kinds of medium . (3) Frequency distributions of the two growth patterns showed that there were two distinct groups of growing cells, with and without a cell volume plateau; these results rule out a single growth state in which plateaus are graded from large to infinitesimally small . (4) Linear regressions fitted to the data for linear growth did not differ significantly from the theoretical model for linear growth without a terminal plateau . These results reveal the operation of a second regulatory system for cell growth in S . pombe at osmolalities closer to those in liquid medium . The occurrence of transitions between the two growth states in successive generations and the agreement between several growth parameters for the two modes suggest that the growth states are closely related.

EMBO J, 1986 Jul, 5(7), 1705 - 9
Periodic transcription as a means of regulating gene expression during the cell cycle: contrasting modes of expression of DNA ligase genes in budding and fission yeast; White JH et al.; Using cultures synchronised by three independent procedures, we have shown that the CDC9 gene, coding for DNA ligase, is periodically expressed in the Saccharomyces cerevisiae cell cycle . The level of CDC9 transcript increases many fold in late G1 reaching a peak at about the G1/S phase boundary and preceding the peak in histone message by some 20 min . The level of DNA ligase itself also fluctuates, showing the expected pattern for a stable enzyme synthesised periodically . In contrast, the transcript from the DNA ligase gene (CDC17) of Schizosaccharomyces pombe is present at a constant level throughout the cell cycle, and no fluctuation in amount was detected, although the histone H2A showed the expected periodic synthesis . Furthermore, DNA ligase activity remains at a constant level during the S . pombe cell cycle showing that there is unlikely to be any form of translational control . These contrasting modes of expression of the DNA ligase genes in the two organisms suggests that when periodic transcription is observed from an essential cell cycle gene, it may have no particular significance for regulating progress through the cell cycle . Also, regulatory circuits may be less well conserved between organisms than the processes they control and thus different organisms may utilise quite different modes of control to achieve the same ends.

J Biol Chem, 1986 Jun 5, 261(16), 7151 - 9
Chemical modification of thiol groups of mitochondrial F1-ATPase from the yeast Schizosaccharomyces pombe . Involvement of alpha- and gamma-subunits in the enzyme activity; Falson P et al.; Mitochondrial F1-ATPase from the yeast Schizosaccharomyces pombe has been prepared under a stable form and in relatively high amounts by an improved purification procedure . Specific chemical modification of the enzyme by the thiol reagent N-ethylmaleimide (NEM) at pH 6.8 leads to complete inactivation characterized by complex kinetics and pH dependence, indicating that several thiols are related to the enzyme activity . A complete protection against NEM effect is afforded by low concentrations of nucleotides in the presence of Mg2+, with ADP and ATP being more efficient than GTP . A total binding of 5 mol of {14C}NEM/mol of F1-ATPase is obtained when the enzyme is 85% inactivated: 3 mol of the label are located on the alpha-subunits and 2 on the gamma-subunit . Two out of the 3 mol on the alpha-subunits bind very rapidly before any inactivation occurs, indicating that the two thiols modified are unrelated to the inactivation process . Complete protection by ATP against inactivation by NEM prevents the modification of three essential thiols out of the group of five thiols labeled in the absence of ATP: one is located on a alpha-subunit and two on the gamma-subunit . These two essential thiols of the gamma-subunit can be differentiated by modification with 6,6'-dithiodinicotinic acid (CPDS), another specific thiol reagent . A maximal binding of 4 mol of {14C}CPDS/mol of enzyme is obtained, concomitant to a 25% inhibition . Sequential modification of the enzyme by CPDS and {14C}NEM leads to the same final deep inactivation as that obtained with {14C}NEM alone . One out of the two thiols of the gamma-subunit is no longer accessible to {14C}NEM after CPDS treatment . When incubated at pH 6.8 with {3H}ATP in the presence of Mg2+, F1-ATPase is able to bind 3, largely exchangeable, mol of nucleotide/mol of enzyme . Modification of the three essential thiols by NEM dramatically decreases the binding of 3H-nucleotide down to about 1 mol/mol of enzyme . Partial modification modifies the cooperative properties, the enzyme being no longer sensitive to anion activation.

Mol Cell Biol, 1986 Jun, 6(6), 2168 - 78
Differential expressions of essential and nonessential alpha-tubulin genes in Schizosaccharomyces pombe; Adachi Y et al.; The fission yeast Schizosaccharomyces pombe has two alpha-tubulin genes and one beta-tubulin gene . Gene disruption experiments showed that the alpha 1-tubulin gene (NDA2) is essential whereas the alpha 2 gene is dispensable . The alpha 2-disrupted cells missing alpha 2 transcript and alpha 2 polypeptide could grow and sporulate normally . The alpha 2 gene, however, was expressed in the wild type and the alpha 1 mutant . Alpha 2-Tubulin was distinguished as an electrophoretic band and was assembled into microtubules . The alpha 2-disrupted cells had an increased sensitivity to an antimicrotubule drug thiabendazole, and the alpha 1(cold-sensitive {cs}) alpha 2 (disrupted) cells became not only cs but also temperature sensitive . Northern blot experiments indicated that alpha 2 transcription was minor and constitutive whereas alpha 1 transcription was major and modulated, depending on the gene copy number of the alpha 2 gene . The amounts of alpha 1 and alpha 2 polypeptides estimated by beta-galactosidase activities of the lacZ-fused genes integrated on the chromosome and by intensities of the electrophoretic bands in crude tubulin fractions, however, were comparable, indicating that alpha 2 tubulin is not a minor subtype . We assume that the cells of Schizosaccharomyces pombe have no excess tubulin pool . alpha 1 mutants would then be blocked in the cell cycle because only half the amount of functional alpha-tubulin required for growth can be produced by the alpha 2 gene . On the other hand, the alpha 2-disrupted cells became viable because the synthesis of alpha 1 tubulin was increased by transcriptional or translational modulation or both . The real cause for essential alpha 1 and dispensable alpha 2 genes seems to be in their regulatory sequences instead of the coding sequences.

J Bacteriol, 1986 Jun, 166(3), 779 - 86
Metabolism of the phospholipid precursor inositol and its relationship to growth and viability in the natural auxotroph Schizosaccharomyces pombe; Fernandez S et al.; Phospholipid metabolism in the fission yeast Schizosaccharomyces pombe was examined . Three enzymes of phospholipid biosynthesis, cytidine diphosphate diacylglycerol synthase (CDP-DG), phosphatidylinositol (PI) synthase, and phosphatidylserine (PS) synthase, were characterized in extracts of S . pombe cells . Contrary to an earlier report, we were able to demonstrate that CDP-DG served as a precursor for PI and PS biosynthesis in S . pombe . S . pombe is naturally auxotrophic for the phospholipid precursor inositol . We found that S . pombe was much more resistant to loss of viability during inositol starvation than artificially generated inositol auxotrophs of Saccharomyces cerevisiae . The phospholipid composition of S . pombe cells grown in inositol-rich medium (50 microM) was similar to that of S . cerevisiae cells grown under similar conditions . However, growth of S . pombe at low inositol concentrations (below 30 microM) affected the ratio of the anionic phospholipids PI and PS, while the relative proportions of other glycerophospholipids remained unchanged . During inositol starvation, the rate of PI synthesis decreased rapidly, and there was a concomitant increase in the rate of PS synthesis . Phosphatidic acid and CDP-DG, which are precursors to these phospholipids, also increased when PI synthesis was blocked by lack of exogenous inositol . The major product of turnover of inositol-containing phospholipids in S . pombe was found to be free inositol, which accumulated in the medium and could be reused by the cell.

J Biol Chem, 1986 May 5, 261(13), 5878 - 85
A single base change in the intron of a serine tRNA affects the rate of RNase P cleavage in vitro and suppressor activity in vivo in Saccharomyces cerevisiae; Willis I et al.; Differences in the processing of dimeric tRNASer-tRNAMet precursors derived from the Schizosaccharomyces pombe sup9 wild-type and opal suppressor genes can be attributed to conformational alterations in the tRNASer anticodon/intron domain . A comparison of the patterns obtained upon transcription of the sup9+ (wild-type) and sup9-e (opal suppressor) genes in a coupled transcription/processing extract from Saccharomyces cerevisiae reveals that the latter exhibits a greatly reduced efficiency of 5'-end maturation and is susceptible to specific endonucleolytic cleavage(s) within the intron . Free energy calculations indicate that these effects coincide with a destabilization of the wild-type anticodon/intron stem and suggest that the predominant sup9-e conformer lacks secondary structure in this region . Evidence in support of this hypothesis was obtained by analyzing the processing of sup9+ and sup9-e precursors carrying the intron base substitution, G37:10, which destroys and restores, respectively, the base-pairing potential of the proposed secondary structure and comparing the strength and temperature sensitivity of sup9-e and sup9-e G37:10 suppression in vivo in S . cerevisiae . The data indicate that the anticodon/intron structure of tRNA precursors can influence the rate of RNase P cleavage in vitro and affect tRNA expression in vivo.

J Bacteriol, 1986 May, 166(2), 484 - 90
Cloning and expression of a Saccharomyces diastaticus glucoamylase gene in Saccharomyces cerevisiae and Schizosaccharomyces pombe; Erratt JA et al.; A recombinant plasmid pool of the Saccharomyces diastaticus genome was constructed in plasmid YEp13 and used to transform a strain of Saccharomyces cerevisiae . Six transformants were obtained which expressed amylolytic activity . The plasmids each contained a 3.9-kilobase (kb) BamHI fragment, and all of these fragments were cloned in the same orientations and had identical restriction maps, which differed from the map of the STA1 gene (I . Yamashita and S . Fukui, Agric . Biol . Chem . 47:2689-2692, 1983) . The glucoamylase activity exhibited by all S . cerevisiae transformants was approximately 100 times less than that of the donor strain . An even lower level of activity was obtained when the recombinant plasmid was introduced into Schizosaccharomyces pombe . No expression was observed in Escherichia coli . The 3.9-kb BamHI fragment hybridized to two sequences (4.4 and 3.9 kb) in BamHI-digested S . diastaticus DNA, regardless of which DEX (STA) gene S . diastaticus contained, and one sequence (3.9 kb) in BamHI-digested S . cerevisiae DNA . Tetrad analysis of crosses involving untransformed S . cerevisiae and S . diastaticus indicated that the 4.4-kb homologous sequence cosegregated with the glucoamylase activity, whereas the 3.9-kb fragment was present in each of the meiotic products . Poly(A)+ RNA fractions from vegetative and sporulating diploid cultures of S . cerevisiae and S . diastaticus were probed with the 3.9-kb BamHI fragment . Two RNA species, measuring 2.1 and 1.5 kb, were found in both the vegetative and sporulating cultures of S . diastaticus, whereas one 1.5-kb species was present only in the RNA from sporulating cultures of S . cerevisiae.

J Mol Biol, 1986 Apr 5, 188(3), 343 - 53
Inactivation of nonsense suppressor transfer RNA genes in Schizosaccharomyces pombe . Intergenic conversion and hot spots of mutation; Heyer WD et al.; Intergenic conversion is a mechanism for the concerted evolution of repeated DNA sequences . A new approach for the isolation of intergenic convertants of serine tRNA genes in the yeast Schizosaccharomyces pombe is described . Contrary to a previous scheme, the intergenic conversion events studied in this case need not result in functional tRNA genes . The procedure utilizes crosses of strains that are homozygous for an active UGA suppressor tRNA gene, and the resulting progeny spores are screened for loss of suppressor activity . In this way, intergenic convertants of a tRNA gene are identified that inherit varying stretches of DNA sequence from either of two other tRNA genes . The information transferred between genes includes anticodon and intron sequences . Two of the three tRNA genes involved in these information transfers are located on different chromosomes . The results indicate that intergenic conversion is a conservative process . No infidelity is observed in the nucleotide sequence transfers . This provides further evidence for the hypothesis that intergenic conversion and allelic conversion are the result of the same molecular mechanism . The screening procedure for intergenic revertants also yields spontaneous mutations that inactivate the suppressor tRNA gene . Point mutations and insertions of A occur at various sites at low frequency . In contrast, A insertions at one specific site occur with high frequency in each of the three tRNA genes . This new type of mutation hot spot is found also in vegetative cells.

Environ Health Perspect, 1986 Mar, 65, 13 - 9
Unique properties of Cd-binding peptides induced in fission yeast, Schizosaccharomyces pombe; Hayashi Y et al.; Metallothioneins, a class of low molecular weight cysteine-rich proteins that bind heavy metal ions, have been found in various eucaryotic organisms . When fission yeasts are grown in the presence of high concentration of CdCl2, large amounts of Cd-binding peptides (Cd-BP1 and Cd-BP2) are synthesized . Cd-BP1 (MW 4000) contains 4 mole of small unit peptide (cadystin, MW 771), 6 mole of Cd2+, and 1 mole of the labile sulfide; on the other hand, Cd-BP2 (MW 1800) contains 2 mole of cadystin and 2 mole of Cd2+ . While Cd-BP2 shows similarities to mammalian Cd-thioneins in UV and CD spectra, Cd-BP1 has a characteristic shoulder at 265 nm in the UV absorption spectrum and shows two marked Cotton bands at 257 nm (negative) and 275 nm (positive) . These characteristics of Cd-BP1 are not found in the other Cd-thioneins . When Cd-BP1 is acidified (pH 2.0) and successively neutralized, a shoulder of 265 nm in the UV spectrum and a Cotton band at 275 nm disappear, and the molecular weight changes from 4000 to 1800, with simultaneous loss of the labile sulfide . While the reconstituted complex without labile sulfide showed the characteristics of Cd-BP2, the reconstituted complex in the presence of labile sulfide indicated partial reconstitution of Cd-BP1 . The UV and CD spectra differences between reconstituted and native Cd-BP1 suggest the requirement for some additional molecular architecture including another peptide-Cd2+ interaction . Induction of cadystin synthesis is almost exclusive for Cd, but an exception is a small amount of cadystin also induced by the higher concentration of CuCl2 (2.5 mM).(ABSTRACT TRUNCATED AT 250 WORDS)

Plasmid, 1986 Mar, 15(2), 156 - 8
Vectors for the construction of gene banks and the integration of cloned genes in Schizosaccharomyces pombe and Saccharomyces cerevisiae; Wright A et al.; We have constructed a variety of vectors for use in both budding yeast (Saccharomyces cerevisiae) and fission yeast (Schizosaccharomyces pombe) . Four of these, pDB262, pWH4, pWH5, and pMAK262, have positive selection for the insertion of cloned DNA, making them convenient for the construction of gene banks . pDB262, pWH4 and pWH5 contain the 2 mu ARS and the LEU2 gene from S . cerevisiae and can be used for gene isolation . They can also be converted into integration vectors for use in the genetic mapping of cloned sequences . pMAK262 contains only the LEU2 gene from budding yeast and can be used to screen for ARS elements or for gene integration . We also describe two other integration vectors, pDAM3 and pDAM6, which have a variety of restriction sites suitable for subcloning.

J Biol Chem, 1986 Feb 25, 261(6), 2936 - 41
Isolation and characterization of the structural gene for secreted acid phosphatase from Schizosaccharomyces pombe; Elliott S et al.; The Schizosaccharomyces pombe acid phosphatase structural gene (PHO 1) was isolated by complementation of an S . pombe acid phosphatase mutant with a wild type S . pombe DNA recombinant plasmid library . Northern analysis indicates that acid phosphatase is encoded by a 1.4-kilobase mRNA of which approximately 100 bases are 3'-poly(A) . The gene contains no introns and the 3' and 5' untranslated regions are short . According to DNA and amino acid sequence data, the S . pombe acid phosphatase has a molecular weight of 50,600 . An 18-amino acid sequence at the N terminus was found that is similar to previously identified signal peptides in other eukaryotic secretory proteins . This signal peptide is apparently removed during secretion, since it is absent in the mature secreted acid phosphatase . The gene can be induced 2--3-fold by starvation for phosphate . The signals required for this induction are contained on the isolated DNA clone . Although the gene can be expressed in Saccharomyces cerevisiae, secretion is abnormal.

J Theor Biol, 1986 Feb 21, 118(4), 405 - 26
Sloppy size control of the cell division cycle; Tyson JJ et al.; In an asynchronous, exponentially proliferating cell culture there is a great deal of variability among individual cells in size at birth, size at division and generation time (= age at division) . To account for this variability we assume that individual cells grow according to some given growth law and that, after reaching a minimum size, they divide with a certain probability (per unit time) which increases with increasing cell size . This model is called sloppy size control because cell division is assumed to be a random process with size-dependent probability . We derive general equations for the distribution of cell size at division, the distribution of generation time, and the correlations between generation times of closely related cells . Our theoretical results are compared in detail with experimental results (obtained by Miyata and coworkers) for cell division in fission yeast, Schizosaccharomyces pombe . The agreement between theory and experiment is superior to that found for any other simple models of the coordination of cell growth and division.

J Cell Sci, 1986 Feb, 80, 253 - 68
Mitosis in the fission yeast Schizosaccharomyces pombe as revealed by freeze-substitution electron microscopy; Tanaka K et al.; Nuclear division in Schizosaccharomyces pombe has been studied in transmission electron micrographs of sections of cells fixed by a method of freeze-substitution . We have found cytoplasmic microtubules in the vicinity of the spindle pole bodies and two kinds of microtubules, short discontinuous ones and long, parallel ones in the intranuclear mitotic spindle . For most of the time taken by nuclear division the spindle pole bodies face each other squarely across the nuclear space but early in mitosis they briefly appear twisted out of alignment with each other, thereby imparting a sigmoidal shape to the bundle of spindle microtubules extending between them . This configuration is interpreted as indicating active participation of the spindle in the initial elongation of the dividing nucleus . It is proposed that mitosis is accompanied by the shortening of chromosomal microtubules simultaneously with the elongation of the central pole-to-pole bundle of microtubules of the intranuclear spindle . Daughter nuclei are separated by the sliding apart of interdigitating microtubules of the spindle at telophase . Some of the latter bear dense knobs at their ends.

Mol Gen Genet, 1986 Feb, 202(2), 291 - 3
The fission yeast cell cycle control gene cdc2: isolation of a sequence suc1 that suppresses cdc2 mutant function; Hayles J et al.; A DNA fragment called suc1 has been found to rescue cells mutated in the cell cycle control gene cdc2 of the fission yeast Schizosaccharomyces pombe . The suppressing activity of suc1 is observed when it is present on a multicopy number plasmid . The gene does not hybridize to cdc2 and maps elsewhere in the genome . Its effect is cdc2 allele specific suggesting that it interacts directly with the cdc2 gene function.

Cell, 1986 Jan 31, 44(2), 329 - 36
Role of a ras homolog in the life cycle of Schizosaccharomyces pombe; Fukui Y et al.; We have analyzed the function of the only ras homolog in S . pombe detectable by Southern blotting, ras1, which is homologous to mammalian ras genes and has been cloned . We have disrupted the ras1 gene and have replaced it with ras1Val17, which corresponds to a transforming variant of mammalian ras . Loss of ras1 activity by disruption results in the complete inability to mate . The cell body of a ras1- strain is extensively deformed, and a ras1-/ras1- diploid sporulates very poorly . Unlike RAS1 and RAS2 of S . cerevisiae, ras1 of S . pombe appears to have no effect on adenylate cyclase activity . This suggests that the target enzymes presumably modulated by ras proteins in signal transduction are not the same for all organisms.

Antonie Van Leeuwenhoek, 1986, 52(1), 45 - 51
Long-chain fatty acid composition of selected genera of yeasts belonging to the Endomycetales; Viljoen BC et al.; The cellular long-chain fatty acids of 36 strains representing 18 genera of the Saccharomycetace, Endomycetaceae, Metchnikowiaceae, Saccharomycodaceae, Schizosaccharomycetaceae and Dipodascaceae were extracted and analyzed as methyl esters by gas chromatography . On the basis of their fatty acid content the set of strains was divided into 6 groups, coinciding with the above families . The members of the Saccharomycetaceae (group I) had a high percentage of oleic acid while the strains classified under the Endomycetaceae (group II) and Metchnikowiaceae (group III) were characterized by oleic acid and linoleic acid as major fatty acids . The Saccharomycodaceae (group IV) had the highest percentage of palmitoleic acid . The Schizosaccharomycetaceae (group V) had the highest percentage of oleic acid, while the Dipodascacea (group VI) were characterized by a high percentage of linoleic acid.

CRC Crit Rev Biochem, 1986, 21(2), 153 - 223
Control of cell growth and division in Saccharomyces cerevisiae; Hanes SD et al.; Considerable advances have been made in recent years in our understanding of the biochemistry of protein and nucleic acid synthesis and, particularly, the molecular biology of gene expression in eukaryotes . The yeast Saccharomyces cerevisiae, and to a lesser extent Schizosaccharomyces pombe, has had a preeminent role as a focus for these studies, principally because of the facility with which these organisms can be experimentally manipulated biochemically and genetically . This review will be designed to critically examine and integrate recent advances in several vital areas of regulatory control of enzyme synthesis in yeast: structure and organization of DNA, transcriptional regulation, post-transcriptional modification, control of translation, post-translational modification and secretion, and cell-cycle modulation . It will attempt to emphasize and illustrate, where detailed information is available, principal underlying molecular mechanisms, and it will attempt to make relevant comparisons of this material to inferred and demonstrated facets of regulatory control of enzyme and protein synthesis in higher eukaryotes.

J Cell Sci Suppl, 1986, 5, 229 - 41
Growth polarity and cytokinesis in fission yeast: the role of the cytoskeleton; Marks J et al.; The distribution of F-actin in the fission yeast Schizosaccharomyces pombe was investigated by fluorescence microscopy using rhodamine-conjugated phalloidin . Fluorescence was seen either at the ends of the cell or at the cell equator . End staining was predominantly in the form of dots whilst equatorial actin was resolved as a filamentous band . The different staining patterns showed a close correlation with the known pattern of cell wall deposition through the cell cycle . In small, newly divided cells actin was localized at the single growing cell end whilst initiation of bipolar cell growth was coincident with the appearance of actin at both ends of the cell . As cells ceased to grow and entered cell division, a ring of actin was seen to anticipate the deposition of the septum at cytokinesis . The relationship between actin and cell wall deposition was further confirmed in three temperature-sensitive cell division cycle (cdc) mutants; cdc10, cdc11 and cdc13 . Immunofluorescence microscopy of S . pombe with an anti-tubulin antibody revealed a system of cytoplasmic microtubules extending between the cell ends . The function of these was investigated in the cold-sensitive, benomyl-resistant mutant ben4 . In cold-grown cells actin was seen to form conspicuous filamentous rings around the nucleus . The origin of these and the possible role of microtubules in the cell-cycle-dependent rearrangements of F-actin are discussed.

Curr Genet, 1986, 11(2), 113 - 7
Acid phosphatase deficient mutants of Schizosaccharomyces pombe are defective in tyrosine uptake; Coddington A et al.; The uptake of tyrosine and arginine into wild type and acid phosphatase deficient mutants (pho 1) of Schizosaccharomyces pombe was investigated . All 11 pho 1-alleles tested exhibited a reduced tyrosine uptake and impaired uptake cosegregated with the lack of acid phosphatase activity . Kinetic analyses using wild type cells grown in high phosphate medium (acid phosphatase repressed) and low phosphate medium (acid phosphatase derepressed) showed staturation kinetics for tyrosine with a KM of about 2 x 10(-4) M for both media and a V of about 5 nmol min-1 mg-1 and 2 nmol min-1 mg-1 for derepressed and repressed cells respectively . The pho 1-118 strain completely lacked this saturable uptake system for tyrosine . Preliminary evidence suggests that tyrosine uptake may be via a general amino acid permease system and we conclude that mutations in the structural gene of acid phosphatase which abolish enzyme activity lead to a loss of this uptake system . In contrast to tyrosine, arginine uptake seems not to be significantly affected either by different acid phosphatase levels in wild type cells or by the pho 1-118 mutation.

Curr Genet, 1986, 10(7), 509 - 14
Isolation of a novel type of mutation in the mitotic control of Schizosaccharomyces pombe whose phenotypic expression is dependent on the genetic background and nutritional environment; Ogden JE et al.; The major cell cycle control in the fission yeast Schizosaccharomyces pombe acts at entry to mitosis, and involves three previously identified genes cdc2, cdc25 and wee1 . The presence of a wee1 mutation phenotypically suppresses cdc25 mutations . This paper describes the isolation and subsequent analysis of a strain in which the suppression is reversed by the presence of a new mutation, designated win1.1 . The mutation causes a slight increase in cell size at division in most genetic backgrounds . However, when combined with a wee1 mutation and cdc25.22, the win1.1 mutation interacts strongly to generate a novel phenotype: cells are phenotypically cdc during growth on minimal medium but cdc+ when cultured on complex medium . The win1 locus is unlinked to previously identified genes involved in mitosis.

Curr Genet, 1986, 10(7), 503 - 8
Transformation of Schizosaccharomyces pombe by non-homologous, unstable integration of plasmids in the genome; Wright AP et al.; In the fission yeast, Schizosaccharomyces pombe, transformation with recombinant plasmids always results in a high proportion of mitotically unstable transformants . This suggested that specialised (ARS) sequences might not be required for autonomous replication of plasmids in S . pombe, contrary to the situation in Saccharomyces cerevisiae . We have shown that specialised ARS sequences, analogous to those in S . cerevisiae, do exist in S . pombe, supporting the view that ARS elements are a general feature of eukaryotes . In addition, there is a further mechanism of plasmid maintenance which involves homologous and non-homologous integration into, and excision from the genome.

Curr Genet, 1986, 10(6), 443 - 7
Genetic mapping of eleven spo genes essential for ascospore formation in the fission yeast Schizosaccharomyces pombe; Kishida M et al.; Sporulation-deficient mutants of the fission yeast Schizosaccharomyces pombe were isolated from a homothallic strain mutagenized with ethyl methanesulfonate . Complementation tests defined two new genetic loci (spo19 and spo20) essential for ascospore formation, in addition to the 18 known spo loci (Bresch et al . 1968) . A novel mapping procedure using random spore analysis prior to tetrad analysis allowed us to map 11 spo genes . Four genes (spo3, spo15, spo19 and spo20) were mapped on chromosome I, 6 genes (spo2, spo4, spo5, spo6, spo14 and spo18) on chromosome III and 1 gene (spo13) on chromosome III . Although there was no noticeable clustering of spo genes on the chromosomes, three pairs of linked genes (spo15-spo20, spo3-spo19 and spo2-spo18) were found.

Curr Genet, 1986, 10(5), 365 - 70
Molecular cloning of a ribosomal protein gene from the fission yeast Schizosaccharomyces pombe; Nischt R et al.; Using the structural gene for the ribosomal protein L3 from Saccharomyces cerevisiae as a probe, we isolated a homologous fragment from genomic DNA of Schizosaccharomyces pombe . Analysis of the plasmid carrying this fragment by hybridization selection and 2D-electrophoresis revealed a 31 kDa ribosomal protein . Transformation of the vector pDB248x containing this fragment into Schizosaccharomyces pombe leads to an increased level of mRNA suggesting that we have cloned the entire and actively transcribed gene.

Curr Genet, 1986, 11(2), 107 - 12
Induction of yeast DNA ligase genes in exponential and stationary phase cultures in response to DNA damaging agents; Johnson AL et al.; UV-irradiation of stationary phase cells of Saccharomyces cerevisiae and Schizosaccharomyces pombe leads to a 9-fold and 90-fold increase in transcript levels from the respective DNA ligase genes CDC9 and CDC17, whereas exponential cells show only 3-fold and 2-fold increases . Induction of CDC9 after MMS treatment and gamma-irradiation was also observed by using a CDC9-lacZ translational fusion and assaying for beta-galactosidase . Surprisingly, irradiation of S . cerevisiae induces only a 50% increase in DNA ligase itself, probably reflecting the extremely high in vivo stability of the enzyme . The UV-induction of ligase may be part of a "fail-safe" mechanism which, together with the enzyme stability, ensures adequate supplies of this essential enzyme.

J Mol Evol, 1986, 23(4), 337 - 42
Examination of protein sequence homologies: III . Ribosomal protein YS25 from Saccharomyces cerevisiae and its counterparts from Schizosaccharomyces pombe, rat liver, and Escherichia coli; Otaka E et al.; The sequences of the ribosomal proteins YS25, SP-S28, RL-S21, and Ec-S6, from Saccharomyces cerevisiae, Schizosaccharomyces pombe, rat liver, and Escherichia coli, respectively, have been examined using a computer program that searches for homologous tertiary structures . Matrices of comparisons among the eukaryotic sequences show that they match each other sequentially without any internal gaps . The average values of the correlation coefficients obtained from the comparison matrices are higher for the first halves of the sequences than for the latter halves . This result suggests that the first halves of the sequences may represent a more important domain than the latter halves . The comparison matrices between the eukaryotic and bacterial sequences of ribosomal proteins, however, do not show sequentially arranged homology, though there are six well-matching segments arranged in different orders in the two types of sequences . This implies that the eukaryotic sequences of the ribosomal protein were reconstituted by two internal transpositions and six deletions of 4-12 residues each from the ancestral sequence during the divergence between bacterial and eukaryotic genes . These findings may give insight into structural and quantitative studies of evolutionary divergence between eukaryotes and prokaryotes.

Gene, 1986, 47(2-3), 245 - 59
Cloning of genes related to exo-beta-glucanase production in Saccharomyces cerevisiae: characterization of an exo-beta-glucanase structural gene; Nebreda AR et al.; The EXG1 gene of Saccharomyces cerevisiae was cloned and identified by complementation of a mutant strain (exg1-2) with highly reduced extracellular exo-beta-1,3-glucanase (EXG) activity . Two recombinant plasmids containing an overlapping region of 5.2 kb were isolated from a genomic DNA library and characterized by restriction mapping . The coding region was located by subcloning the original DNA inserts in a 2.7-kb HindIII-XhoI fragment . Exg+ strains and Exg- mutants transformed with yeast multicopy plasmids containing this DNA fragment showed an EXG activity 5- to 20-fold higher than for the untransformed Exg+ wild-type (wt) strains . The overproduced EXG had the same enzymic activity on different substrates, and showed the same electrophoretic behaviour on polyacrylamide gels and identical properties upon filtration through Sephacryl S-200 as those of the main EXG from Exg+ wt strains . The EXG1 gene transformed Schizosaccharomyces pombe, yielding extracellular EXG activity which showed cross-reactivity with anti-S . cervisiae EXG antibodies . A fragment including only a part of the EXG1 region was subcloned into the integrating vector YIp5, and the resulting plasmid was used to transform an Exg+ strain . Genetic and Southern analysis of several stable Exg- transformants showed that the fragment integrated by homology with the EXG1 locus . The chromosomal DNA fragment into which the plasmid integrated has a restriction pattern identical to that of the fragment on which we had previously identified the putative EXG1 gene . Only one copy of the EXG1 gene per genome was found in several strains tested by Southern analysis . Furthermore, two additional recombinant plasmids sharing a yeast DNA fragment of about 4.1 kb, which partially complements the exg1-2 mutation but which shows no homology with the 2.7-kb fragment containing the EXG1 gene, were also identified in this study . This 4.1-kb DNA fragment does not appear to contain an extragenic suppressor and could be related in some way to EXG production in S . cerevisiae.

J Mol Evol, 1986, 23(1), 41 - 51
The cloning and characterization of a RAS gene from Schizosaccharomyces pombe; Nadin-Davis SA et al.; We have cloned and determined the complete nucleotide sequence of a RAS gene from the yeast Schizosaccharomyces pombe (SP-RAS) . The putative RAS protein of 214 amino acids is encoded by two noncontiguous reading frames separated by an intron of 86 bp . The SP-RAS gene product shares extensive homology with the proteins of the Saccharomyces cerevisiae (SC), Dictyostelium, Drosophila, and human RAS genes in its N-terminal region but not in its C-terminal region . The extended C-terminal regions found in the SC-RAS genes have no counterpart in the SP-RAS gene . Thus the RAS genes of these two yeasts are structurally quite distinct . The SP-RAS sequence was expressed in vivo.

Gene, 1986, 45(3), 289 - 97
The mosaic cox1 gene in the mitochondrial genome of Schizosaccharomyces pombe: minimal structural requirements and evolution of group I introns; Trinkl H et al.; The gene encoding subunit 1 of cytochrome oxidase (cox1) in the fission yeast Schizosaccharomyces pombe is polymorphic . In strain 50 it contains two group I introns with open reading frames (ORFs) in phase with the upstream exons (Lang, 1984) . In strain EF1 two additional very short group I introns which do not possess ORFs were detected by DNA sequencing . These two introns (AI2a and AI3) share distinct characteristics concerning their nucleotide sequence and secondary structure and are located at identical positions as the introns AI4 and AI5 beta, respectively, in the cox1 gene of Saccharomyces cerevisiae . The sequence homology of the cob and cox1 genes around the splice points of introns AI2a, AI4, and BI4 (cob intron 4) might reflect horizontal gene transfer between the distantly related species S . pombe and S . cerevisiae.

Mol Cell Biol, 1986 Jan, 6(1), 80 - 9
Replicating plasmids in Schizosaccharomyces pombe: improvement of symmetric segregation by a new genetic element; Heyer WD et al.; We characterized a number of widely used yeast-Escherichia coli shuttle vectors in the fission yeast Schizosaccharomyces pombe . The 2 micron vectors pDB248 and YEp13 showed high frequency of transformation, intermediate mitotic and low meiotic stability, and a low copy number in S . pombe, analogous to their behavior in {cir0} strains of Saccharomyces cerevisiae . The S . cerevisiae integration vectors pLEU2 and pURA3 transformed S . pombe at very low frequencies but, surprisingly, in a nonintegrative fashion . Instead, they replicated autonomously, and they showed very high copy numbers (up to 150 copies per plasmid-containing cell) . This could reflect a lack of sequence specificity for replication of plasmid DNA in S . pombe . pFL20, an S . pombe ars vector, and a series of plasmids derived from it were studied to analyze the unusually high stability of this plasmid . Mitotic stability and partitioning of the plasmids was measured by pedigree analysis of transformed S . pombe cells . An S . pombe DNA fragment (stb) was identified that stabilizes pFL20 by improvement of plasmid partitioning in mitosis and meiosis.

Gene, 1986, 41(2-3), 321 - 5
Cloning and characterization of the Schizosaccharomyces pombe DNA ligase gene CDC17; Johnston LH et al.; The Schizosaccharomyces pombe CDC17 gene has been cloned by complementation of the cdc17 mutant coding for temperature-sensitive DNA ligase . An allele-specific suppressor active only in the presence of a high osmotic pressure was also isolated . The cloned CDC17 gene failed to complement the analogous DNA ligase mutation, cdc9, in Saccharomyces cerevisiae, although the reverse complementation was successful {Barker and Johnston, Eur . J . Biochem . 134 (1983) 315-319} . The CDC17 gene specifies a 2.8-kb transcript.

Curr Genet, 1986, 10(5), 359 - 64
A new mutation for multiple drug resistance and modified plasma membrane ATPase activity in Schizosaccharomyces pombe; Ulaszewski S et al.; The mutant JV66 was selected from the wild type strain of S . pombe 972h- ade7-413 by its ability to grow on solid rich medium containing 200 micrograms Dio-9/ml . The single nuclear mutation, designated pma1 gives resistance towards diguanidines and several other positively charged compounds . The pma1 mutation also decreases plasma membrane ATPase activity and confers resistance of ATPase to vanadate . The pma1 locus is localized on chromosome I at 5.3 map units from cyh1-C7 and at about 20.7 map units from the centromere . This new mutation is genetically and phenotypically different from the mutation cyh3 and cyh4 previously described (Johnston and Coddington 1983).

Nucleic Acids Res, 1985 Dec 20, 13(24), 8739 - 47
Dimeric tRNA gene arrangement in Schizosaccharomyces pombe allows increased expression of the downstream gene; Hottinger-Werlen A et al.; Three Schizosaccharomyces pombe dimeric tRNA genes, consisting of a tRNASer gene encoding a minor species with an intervening sequence followed by a tRNAMeti gene, have been described {Mao et al . (1980) Cell 21, 509-516; Hottinger et al . (1982) Mol . Gen . Genet . 188, 219-224; Willis et al . (1984) EMBO J . 3, 1573-1580} . We have examined the reason for the dimeric structure by comparing the transcriptional efficiencies and competitive abilities of the genes subcloned from the dimeric arrangement . Both of the subcloned genes are active in vivo in Saccharomyces cerevisiae, but only the tRNASer gene is efficiently transcribed in vitro . The tRNASer gene competes efficiently for transcription factors, while the tRNAMeti gene does so only weakly . Thus, it appears that the dimeric arrangement is required to support expression of the tRNAMeti gene . S . pombe genes encoding major species of tRNASer are transcribed considerably less efficiently than are the minor genes from the dimers, so coupling of the tRNAMeti gene to the minor species genes should lead to efficient production of tRNAMeti.

Biochim Biophys Acta, 1985 Dec 18, 826(4), 180 - 5
DNA ligase-AMP adducts: identification of yeast DNA ligase polypeptides; Banks GR et al.; Yeast DNA ligase is radioactively labelled in vitro by incubating a crude cell extract with {alpha-32P}ATP . The product of this reaction is the stable covalent ligase-AMP adduct, which can be characterized by its reactivity with either pyrophosphate or nicked DNA and visualized by gel electrophoresis and autoradiography . The Saccharomyces cerevisiae DNA ligase was identified as an 89 kDa polypeptide by exploiting the fact that transformants with multiple copies of the plasmid-encoded DNA ligase (CDC9) gene overproduce the enzyme by two orders of magnitude . A similar strategy has been used to identify the Schizosaccharomyces pombe DNA ligase as an 87 kDa polypeptide . Both values agree well with the coding capacities of the respective cloned gene sequences . When the S . cerevisiae ligase is greatly overproduced with respect to wild-type levels, a second polypeptide of 78.5 kDa is also labelled and has the same properties as the 89 kDa adduct . We suggest that this polypeptide is generated by proteolysis.

EMBO J, 1985 Dec 16, 4(13A), 3531 - 8
Histone gene organization of fission yeast: a common upstream sequence; Matsumoto S et al.; Histone genes of the fission yeast Schizosaccharomyces pombe were cloned from Charon 4A and cosmid gene libraries by hybridization, and their nucleotide sequences were determined . The genome of S . pombe has a single, isolated H2A, a pair of H2A-H2B and three pairs of H3-H4 (one H2B, two H2A and three each of H3 and H4) . This non-assorted histone gene organization is distinct from that of the budding yeast which has two pairs of H2A-H2B and H3-H4 . The predicted amino acid sequences of S . pombe histone H2As, H3s and H4s were identical except for three residue changes in H2As . Compared with those os S . cerevisiae and human, variable residues were clustered near the NH2- and COOH-terminal regions of H2A and H2B . Sequence homologies to the two organisms were roughly the same in H2A (79-83%), H3 (92-93%) and H4 (91%), but differed in H2B (82% to S . cerevisiae and 68% to human) . The coding sequences in pairs of S . pombe histone genes were divergently directed . A 17-bp long highly homologous sequence (AACCCT box) that had internal 6-bp direct repeats was present in the intergene spacer sequences or in the 5' upstream region of all the cloned histone genes . A possible regulatory role of the common upstream sequence for histone gene expression is discussed.

EMBO J, 1985 Dec 16, 4(13A), 3553 - 6
Cloning of opal suppressor tRNA genes of a filamentous fungus reveals two tRNASerUGA genes with unexpected structural differences; Debuchy R et al.; The informational suppressors su4-1 and su8-1 of Podospora anserina were isolated by transformation of Schizosaccharomyces pombe UGA mutants . The DNA sequence revealed that they were opal (UGA) suppressor tRNAs . Wild-type alleles were also isolated by hybridization . The DNA sequence showed that they both encode species of tRNASerUGA . The gene SU8 has an 18-bp intervening sequence and its primary sequence is very different from that of SU4.

Biochemistry, 1985 Dec 3, 24(25), 7418 - 23
Primary structures of ribosomal protein YS25 from Saccharomyces cerevisiae and its counterparts from Schizosaccharomyces pombe and rat liver; Itoh T et al.; Protein YS25 and its counterparts, SP-S28 and rat S21 {nomenclature according to Sherton, C . C., & Wool, I . G . (1972) J . Biol . Chem . 247, 4460-4467}, from Saccharomyces cerevisiae, Schizosaccharomyces pombe, and rat liver cytoplasmic ribosomes, respectively, were sequenced by a combination of various enzymatic digestions and/or chemical cleavage . Proteins YS25 and SP-S28 consist of 87 amino acid residues, and rat S21 consists of 83 . The amino termini are all N alpha-acetylated . The amino-terminal halves of the protein molecules are highly conserved (73-85% homologies) in contrast to the carboxy-terminal parts . Overall, rat S21 is 54% homologous to YS25 and 57% to SP-S28, despite a 76% homology between YS25 and SP-S28 . Direct comparison with the available prokaryotic ribosomal protein sequences did not reveal any significant homology.

Nature, 1985 Nov 7-13, 318(6041), 78 - 80
Fission yeast Schizosaccharomyces pombe correctly excises a mammalian RNA transcript intervening sequence; Kaufer NF et al.; Study of heterologous gene expression in the budding yeast Saccharomyces cerevisiae has shown that this organism is incapable of correctly removing intervening sequences from transcripts of higher eukaryotic genes . This is probably due to the stringent requirement for the presence of a TACTAAC box close to the 3' end of the intervening sequence if splicing in S . cerevisiae is to occur . Comparison of the introns found in the fission yeast Schizosaccharomyces pombe has identified conserved sequences similar to those found in higher eukaryotes . Therefore, we have investigated whether Schiz . pombe is capable of accurately excising intervening sequences from the transcripts of higher eukarotic genes . We show here that both the 5' and 3' splice sites of the simian virus 40 (SV40) small-T antigen transcript are accurately utilized when cloned viral DNA is expressed in Schiz . pombe cells . These data suggest that Schiz . pombe may be a better model system than S . cerevisiae for the genetic study of RNA splicing and for expressing higher eukaryotic genes.

J Antibiot (Tokyo), 1985 Nov, 38(11), 1573 - 80
Leptomycins A and B, new antifungal antibiotics . III . Mode of action of leptomycin B on Schizosaccharomyces pombe; Hamamoto T et al.; Mode of action of leptomycin B (LMB), a new antifungal antibiotic, was studied with Schizosaccharomyces pombe . A low concentration of LMB caused inhibition of cell division, producing elongated cells with morphologically altered nuclei and several cell plates, while it inhibited nucleic acid synthesis in intact cells at 100-fold higher concentration . Addition of LMB during G2 phase in synchronous culture blocked following events in cell cycle . Analysis of the effect of LMB on cdc mutants suggested the antibiotic inhibited some specific step, possibly in M phase just prior to nuclear division.

Mol Cell Biol, 1985 Nov, 5(11), 3261 - 9
Organization, primary structure, and evolution of histone H2A and H2B genes of the fission yeast Schizosaccharomyces pombe; Choe J et al.; The histone H2A and H2B genes of the fission yeast Schizosaccharomyces pombe were cloned and sequenced . Southern blot and sequence analyses showed that, unlike other eucaryotes, Saccharomyces cerevisiae included, S . pombe has unequal numbers of these genes, containing two histone H2A genes (H2A-alpha and -beta) and only one H2B gene (H2B-alpha) per haploid genome . H2A- and H2B-alpha are adjacent to each other and are divergently transcribed . H2A-beta has no other histone gene in close proximity . Preceding both H2A-alpha and -beta is a highly conserved 19-base-pair sequence (5'-CATCAC/AAACCCTAACCCTG-3') . The H2A DNA sequences encode two histone H2A subtypes differing in amino acid sequence (three residues) and size (H2A-alpha, 131 residues; H2A-beta, 130 residues) . H2B-alpha codes for a 125-amino-acid protein . Sequence evolution is extensive between S . pombe and S . cerevisiae and displays unique patterns of divergence . Certain N-terminal sequences normally divergent between eucaryotes are conserved between the two yeasts . In contrast, the normally conserved hydrophobic core of H2A is as divergent between the yeasts as between S . pombe and calf.

J Mol Biol, 1985 Sep 5, 185(1), 65 - 81
Yeast promoters URA1 and URA3 . Examples of positive control; Losson R et al.; Transcription of the two unlinked structural genes URA1 and URA3 of Saccharomyces cerevisiae is positively regulated by the gene product PPR1 . We have used S1 digestion and primer extension mapping to investigate the RNAs produced in different genetic backgrounds: wild-type, ppr1 deletion mutants, constitutively induced and non-inducible ppr1 mutants . Results show that each structural gene specifies multiple messenger RNA classes with different 5'-terminal sequences . The basal level of these transcripts does not require a functional PPR1 gene . Induction of URA1 results from an even increase of the level of synthesis of all the transcripts in contrast to that of URA3 which is effected by selectively increasing the levels of synthesis of one subset of transcripts . The PPR1-mediated control was also studied in the foreign genetic background of Schizosaccharomyces pombe using autonomously replicating hybrid plasmids carrying the gene URA1 or URA3 along with the regulatory gene PPR1, either in a constitutive or non-inducible allelic form . The 5' ends of the transcripts URA1 and URA3 made in S . pombe map upstream from the initiation sites used in S . cerevisiae . In contrast to S . cerevisiae, in S . pombe the URA3 but not URA1 transcripts respond to the PPR1-induction . We have identified a minimal control region for the PPR1-specific induction of URA1, that includes sequences located between the T-A-T-A box and the translation start codon . This region contains sequence features in common with URA3 . There is an extensive alternating Pu:Py region including the T-A-T-A box of both promoters and an eight base-pair exact homology; further downstream, there is another 11 base-pair highly conserved sequence which either overlaps or lies in close proximity to the unregulated start sites of URA1 in S . pombe and of URA3 in S . cerevisiae . A positive regulatory model taking into accounts all these observations is presented.

Arch Microbiol, 1985 Sep, 142(4), 370 - 4
Subcellular localization and glycoprotein nature of the invertase from the fission yeast Schizosaccharomyces pombe; Moreno S et al.; The subcellular localization of the enzyme invertase in Schizosaccharomyces pombe cells, both repressed and derepressed for synthesis of the enzyme, was studied . Most of the invertase was found to be located outside the plasma membrane and only a small percentage was found to be associated to membranes . A substantial portion of the external enzyme remained firmly bound to cell-wall material . All of the invertase recovered in soluble form from cellular extracts reacted with concanavalin A and with the lectin from Bandeiraea simplicifolia seeds, indicating the presence in the enzyme of a carbohydrate moiety which probably contains terminal mannosyl (or structurally related) and galactosyl residues . The possibility of the presence of two different forms of invertase in S . pombe was considered . An intracellular, soluble form of invertase, devoid of carbohydrate, similar to the small invertase of the budding yeast Saccharomyces cerevisiae, was not found in S . pombe . However, the Michaelis constant for sucrose of the enzyme present in repressed cells was smaller than that of the invertase synthesized under derepressing conditions, although this difference could also be the result of a different pattern of glycosylation of the invertase synthesized under different growth conditions.

Mol Cell Biol, 1985 Sep, 5(9), 2389 - 98
The two alpha-tubulin genes of Chlamydomonas reinhardi code for slightly different proteins; Silflow CD et al.; Full-length cDNA clones corresponding to the transcripts of the two alpha-tubulin genes in Chlamydomonas reinhardi were isolated . DNA sequence analysis of the cDNA clones and cloned gene fragments showed that each gene contains 1,356 base pairs of coding sequence, predicting alpha-tubulin products of 451 amino acids . Of the 27 nucleotide differences between the two genes, only two result in predicted amino acid differences between the two gene products . In the more divergent alpha 2 gene, a leucine replaces an arginine at amino acid 308, and a valine replaces a glycine at amino acid 366 . The results predicted that two alpha-tubulin proteins with different net charges are produced as primary gene products . The predicted amino acid sequences are 86 and 70% homologous with alpha-tubulins from rat brain and Schizosaccharomyces pombe, respectively . Each gene had two intervening sequences, located at identical positions . Portions of an intervening sequence highly conserved between the two beta-tubulin genes are also found in the second intervening sequence of each of the alpha genes . These results, together with our earlier report of the beta-tubulin sequences in C . reinhardi, present a picture of the total complement of genetic information for tubulin in this organism.

J Mol Biol, 1985 Aug 5, 184(3), 353 - 66
The mitochondrial genome of the fission yeast Schizosaccharomyces pombe . The cytochrome b gene has an intron closely related to the first two introns in the Saccharomyces cerevisiae cox1 gene; Lang BF et al.; The DNA sequence of the cob region of the Schizosaccharomyces pombe mitochondrial DNA has been determined . The cytochrome b structural gene is interrupted by an intron of 2526 base-pairs, which has an open reading frame of 2421 base-pairs in phase with the upstream exon . The position of the intron differs from those found in the cob genes of Saccharomyces cerevisiae, Aspergillus nidulans or Neurospora crassa . The Sch . pombe cob intron has the potential of assuming an RNA secondary structure almost identical to that proposed for the first two cox1 introns (group II) in S . cerevisiae and the p1-cox1 intron in Podospora anserina . It has most of the consensus nucleotides in the central core structure described for this group of introns and its comparison with other group II introns allows the identification of an additional conserved nucleotide stretch . A comparison of the predicted protein sequences of group II intronic coding regions reveals three highly conserved blocks showing pairwise amino acid identities of 34 to 53% . These regions comprise over 50% of the coding length of the intron but do not include the 5' region, which has strong secondary structural features . In addition to the potential intron folding, long helical structures involving repetitive sequences can be formed in the flanking cob exon regions . A comparison of the Sch . pombe cytochrome b sequence with those available from other organisms indicates that Sch . pombe is evolutionarily distant from both budding yeasts and filamentous fungi . As was seen for the Sch . pombe cox1 gene (Lang, 1984), the cob exons are translated using the universal genetic code and this distinguishes Sch . pombe mitochondria from all other fungal and animal mitochondrial systems.

Genetika, 1985 Aug, 21(8), 1266 - 71
{Specificity of replicating instability in Schizosaccharomyces pombe haploid yeasts}; Kurennaia ON et al.; UV-induced genetic instability in haploid Schizosaccharomyces pombe does not appear to be very locus-specific . This conclusion contradicts the data previously published by other authors . The possible causes for this discrepancy are discussed.

Exp Cell Res, 1985 Aug, 159(2), 495 - 509
Changes in phosphoprotein pattern in Schizosaccharomyces pombe; Querengesser LD et al.; A variety of evidence suggests that protein phosphorylation (pp) may be important in cell-cycle control . Phosphorylated proteins from S . pombe have been examined for phosphorylation changes under several conditions: known triggers of the division control (low nitrogen, low phosphate), cell size mutants (WEE1 and CDC2 alleles) and cell cycle mutants (CDC2, CDC10, CDC17, CDC25 alleles) . Three major phosphorylated proteins (pp38, pp45 and pp54) showed the greatest response to nutritional shifts . The changes in the phosphorylated states of these proteins correlated with growth rate . Some phosphorylations (e.g . pp53) occurred transiently following a stimulus to cell division suggesting a possible involvement with the division mechanism . An allele-specific alteration of charge was noted for pp45 suggesting that this protein is the product of the CDC2 gene . The wee1-6 phosphoprotein pattern is similar to wild-type indicating that this mutant cell line accurately senses its nutritional environment and that the mutation likely affects the transfer of this information to the division control . Cells blocked by various temperature-sensitive cell cycle mutants did not show an alteration of phosphoprotein pattern.

EMBO J, 1985 Aug, 4(8), 2093 - 9
Identification and molecular analysis of a third Aspergillus nidulans alcohol dehydrogenase gene; McKnight GL et al.; An Aspergillus nidulans functional cDNA encoding an alcohol dehydrogenase (ADH) was isolated by its ability to complement an adh1 mutation in Saccharomyces cerevisiae . Alignment of the cDNA and cloned genomic DNA sequences indicated that the ADH gene contains two small introns . The presence of ethanol in the growth medium was shown to result in ADH mRNA accumulation presumably due to transcriptional induction of the gene . However, ADH mRNA accumulation was at most only partially repressed by the presence of glucose . The ADH gene characterized here is designated ADH3 since it is distinct from the alcA gene which encodes ADH I and appears distinct from the gene which encodes ADH II . We demonstrated that the first intron in the A . nidulans ADH3 gene was not efficiently spliced in S . cerevisiae whereas the promoter region was utilized weakly . We also present a comparison of the primary structure of A . nidulans ADH III with the alcohol dehydrogenases of S . cerevisiae and Schizosaccharomyces pombe.

Exp Cell Res, 1985 Jun, 158(2), 544 - 53
Novel cell cycle regulation in the yeast Schizosaccharomyces pombe . The DNA-division sequence modulates mass accumulation; Johnston GC et al.; For wee1 mutant cells of Schizosaccharomyces pombe the DNA-division sequence of the cell cycle can be differentially slowed by the presence of low concentrations of the S-phase inhibitor hydroxyurea, or by semipermissive temperatures for certain wee1 cdc double mutants . Under these conditions the rate of proliferation is decreased, yet still exponential . Relief of these constraints slowing the DNA-division sequence resulted in prompt increases in the exponential rates of mass accumulation, to rates greater than those normally found . These observations suggest that mass accumulation by this yeast is always modulated by performance of the DNA-division sequence.

Exp Cell Res, 1985 Jun, 158(2), 533 - 43
Indirect suppression of the wee1 mutant phenotype in Schizosaccharomyces pombe; Singer RA et al.; For S . pombe cells mutations in the wee1 regulatory gene have been shown previously to allow cells to be smaller than normal at cell division, to endow the cell with a significantly long G1 cell cycle interval, and to alter the timing in the cell cycle of certain mutationally-defined cell cycle steps in G2 . We show here that situations which lengthen S phase in proliferating wee1 mutant cells 'suppress' to varying degrees these wee1-mediated cell cycle alterations . Conditions chosen to protract S phase were use of cdc22.M45 mutant cells at semipermissive temperatures, and the presence of sub-arresting concentrations of the S phase inhibitors hydroxyurea or deoxyadenosine . Proliferation in the presence of each of these inhibitors was shown directly to result in protracted S phase . Residual cell division measurements were used to measure the cell cycle timing of G1 and G2 cell-cycle steps . The indirect suppression of the wee1 phenotype shown here can be understood in terms of the proposed role of the wee1+ gene product in coordinating cell division with cellular growth.

J Bacteriol, 1985 Jun, 162(3), 902 - 4
Buoyant density constancy of Schizosaccharomyces pombe cells; Kubitschek HE et al.; Buoyant densities of cells from exponentially growing cultures of the fission yeast Schizosaccharomyces pombe 972h- with division rates from 0.14 to 0.5 per h were determined by equilibrium centrifugation in Percoll gradients . Buoyant densities were independent of growth rate, with an average value (+/- standard error) of 1.0945 (+/- 0.00037) g/ml . When cells from these cultures were separated by size, mean cell volumes were independent of buoyant density, indicating that buoyant densities also were independent of cell age during the division cycle . These results support the suggestion that most or all kinds of cells that divide by equatorial fission may have similar, evolutionarily conserved mechanisms for regulation of buoyant density.

EMBO J, 1985 Jun, 4(6), 1569 - 73
Transformation of Saccharomyces cerevisiae and Schizosaccharomyces pombe with linear plasmids containing 2 micron sequences; Guerrini AM et al.; Linear plasmids were constructed by adding telomeres prepared from Tetrahymena pyriformis rDNA to a circular hybrid Escherichia coli-yeast vector and transforming Saccharomyces cerevisiae . The parental vector contained the entire 2 mu yeast circle and the LEU gene from S . cerevisiae . Three transformed clones were shown to contain linear plasmids which were characterized by restriction analysis and shown to be rearranged versions of the desired linear plasmids . The plasmids obtained were imperfect palindromes: part of the parental vector was present in duplicated form, part as unique sequences and part was absent . The sequences that had been lost included a large portion of the 2 mu circle . The telomeres were approximately 450 bp longer than those of T . pyriformis . DNA prepared from transformed S . cerevisiae clones was used to transform Schizosaccharomyces pombe . The transformed S . pombe clones contained linear plasmids identical in structure to their linear parents in S . cerevisiae . No structural re-arrangements or integration into S . pombe was observed . Little or no telomere growth had occurred after transfer from S . cerevisiae to S . pombe . A model is proposed to explain the genesis of the plasmids.

J Biol Chem, 1985 May 25, 260(10), 6348 - 53
Isolation of the fructose-1,6-bisphosphatase gene of the yeast Schizosaccharomyces pombe . Evidence for transcriptional regulation; Vassarotti A et al.; The fructose-1,6-bisphosphatase structural gene (FBP+) of Schizosaccharomyces pombe has been isolated by genetic complementation of a deficient mutant, which is characterized by the inability to grow on a nonfermentable carbon source such as glycerol . Growth on glycerol-containing medium was restored in a S . pombe fructose-1,6-bisphosphatase-deficient mutant (fbp-16) when it was transformed with a plasmid (pAVO4) carrying FBP+ . The transformant displayed a 5-fold increase in enzymatic activity when compared to the parental S . pombe strain . Subcloning of DNA fragments from the 8.5-kilobase (kb) insert of pAVO4 defined a 4-kb DNA fragment which contained the functional FBP+ gene and its regulatory region . When this gene was placed under the control of the lac promoter-operator, functional expression in Escherichia coli was obtained, as deduced by complementation of bacterial fructose-1,6-bisphosphatase mutants . The FBP+ gene encodes a 1.9-kb glucose-repressible transcript whose appearance in S . pombe is correlated with fructose-1,6-bisphosphatase derepression in glycerol-containing medium . We suggest that the regulation of the S . pombe FBP+ gene is exerted at the transcriptional level . The S . pombe FBP+ gene gave rise to a 1.9-kb transcript in Saccharomyces cerevisiae, but not to measurable enzymatic activity.

Nucleic Acids Res, 1985 May 24, 13(10), 3711 - 22
Evaluation of heterologous ARS activity in S . cerevisiae using cloned DNA from S . pombe; Maundrell K et al.; Cloned segments of Schizosaccharomyces pombe genomic DNA were screened for ARS activity in the native host, S . pombe, using high frequency transformation, phenotypic instability and extrachromosomal maintenance of unrearranged plasmid sequences as criteria for ARS function . This analysis revealed 12 ARS elements in a total of 230 kb of chromosomal DNA, indicating an average frequency of one ARS every 19 kb of genomic DNA . We then used these clones to assess the reliability of the S . cerevisiae assay for detecting ARS elements in heterologous DNA . The results show that not only does the S . cerevisiae assay fail to detect a large proportion of true ARS elements but it also wrongly identifies a significant proportion of clones which did not display ARS activity in the native host . We would therefore recommend restraint when extrapolating from observed ARS function of heterologous DNA in S . cerevisiae to a presumed analogous role in the original host.

J Cell Sci, 1985 Apr, 75, 357 - 76
Growth in cell length in the fission yeast Schizosaccharomyces pombe; Mitchison JM et al.; The cylindrical cells of Schizosaccharomyces pombe grow in length by extension at the ends and not the middle . At the beginning of the cell cycle, growth is restricted to the 'old end', which existed in the previous cycle . Later on, the 'new end', formed from the septum, starts to grow at a point in the cycle that we have called NETO ('new end take-off') . Fluorescence microscopy on cells stained with Calcofluor has been used to study NETO in size mutants, in blocked cdc mutants and with different growth temperatures and media . In wild-type cells (strain 972) NETO happens at 0.34 of the cycle with a cell length of 9.5 microns . With size mutants that are smaller at division, NETO takes place at the same size (9.0-9.5 microns) but this is not achieved until later in the cycle . Another control operates in larger size mutants since NETO occurs at the same stage of the cycle (about 0.32) as in wild type but at a larger cell size . This control is probably a requirement to have completed an event in early G2, since most cdc mutant cells blocked before this point in the cycle do not show NETO whereas most of those blocked in late G2 do show it . We conclude that NETO only happens if: (1) the cell length is greater than a critical value of 9.0-9.5 microns; and (2) the cell has traversed the first 0.3-0.35 of the cycle and passed early G2 . NETO is delayed in poor media, in which cell size is also reduced . Temperature has little effect on NETO under steady-state conditions, but there is a transient delay for some hours after a temperature shift . NETO is later in another wild-type strain, 132 . Time-lapse photomicrography was used to follow the rates of length growth in single cells . Wild-type cells showed two linear segments during the first 75% of the cycle . There was a rate-change point (RCP), coincident with NETO, where the rate of total length extension increased by 35% . This increase was not due simply to the start of new-end growth, since old-end growth slowed down in some cells at the RCP . cdc 11.123 is a mutant in which septation and division is blocked at 35 degrees C but nuclear division continues.(ABSTRACT TRUNCATED AT 400 WORDS)

Mol Cell Biol, 1985 Apr, 5(4), 808 - 15
Mutations preventing expression of sup3 tRNASer nonsense suppressors of Schizosaccharomyces pombe; Pearson D et al.; Suppression of nonsense codons in Schizosaccharomyces pombe by sup3-e tRNASerUGA or sup3-i tRNASerUAA is reduced or abolished by mutations within the suppressor locus . Twenty-five suppressor-inactive sup3-e genes and thirteen mutant sup3-i genes were isolated from S . pombe genomic clone banks by colony hybridization . Sequence analysis of these revertant alleles corroborates genetic evidence for mutational hotspots within the sup3 tRNA gene . Fifteen types of point mutations or insertions were found . Many of these replace bases which are highly or completely conserved in eucaryotic tRNA genes . Transcription of the altered sup3 genes in a Saccharomyces cerevisiae extract enabled the identification of mutations which affect the rate of 5'-end maturation or splicing of the tRNA precursors or both . A total of seven mutations were found which alter transcriptional efficiencies . Of these, five are located outside the internal transcription control regions.

Cell, 1985 Apr, 40(4), 879 - 86
Concerted evolution of tRNA genes: intergenic conversion among three unlinked serine tRNA genes in S . pombe; Amstutz H et al.; In many cases the multiple genes coding for one specific tRNA are dispersed throughout the genome . The members of such a gene family nevertheless maintain a common nucleotide sequence during evolution . A major mechanism contributing to this concerted evolution is intergenic conversion . Here we show that it occurs between three tRNA genes of related sequence residing on different chromosomes of Schizosaccharomyces pombe . Sequence analysis of converted genes indicates that blocks of a minimal length of 18-33 bp and of a maximal length of 190 bp can be transferred from one gene to the other . During meiosis the frequency of these transfers lies in the order of 10(-5) per progeny spore . Information transfer between any two members of the gene family occurs in both directions.

Eur J Biochem, 1985 Apr 1, 148(1), 35 - 9
Essential arginyl residues in the H+-translocating ATPase of plasma membrane from the yeast Schizosaccharomyces pombe; Di Pietro A et al.; The H+-translocating adenosine-5'-triphosphatase (ATPase) purified from the yeast Schizosaccharomyces pombe is inactivated upon incubation with the arginine modifier 2,3-butanedione . The inactivation of the enzyme is maximal at pH values above 8.5 . The modified enzyme is reactivated when incubated in the absence of borate after removal of 2,3-butanedione . The extent of inactivation is half maximal at 10 mM 2,3-butanedione for an incubation of 30 min at 30 degrees C at pH 7.0 . Under the same conditions, the time-dependence of inactivation is biphasic in a semi-logarithmic plot with half-lives of 10.9 min and 65.9 min . Incubation with 2,3-butanedione lowering markedly the maximal rate of ATPase activity does not modify the Km for MgATP . These data suggest that two classes of arginyl residues play essential role in the plasma membrane ATPase activity . Magnesium adenosine 5'-triphosphate (MgATP) and magnesium adenosine 5'-diphosphate (MgADP), the specific substrate and product, protect partially against enzyme inactivation by 2,3-butanedione . Free ATP or MgGTP which are not enzyme substrates do not protect . Free magnesium, another effector of enzyme activity, exhibits partial protection at magnesium concentrations up to 0.5 mM, while increased inactivation is observed at higher Mg2+ concentrations . These protections indicate either the existence of at least one reactive arginyl in the substrate binding site or a general change of enzyme conformation induced by MgATP, MgADP or free magnesium.

Nucleic Acids Res, 1985 Mar 25, 13(6), 1855 - 69
The linear extrachromosomal DNA of Physarum polycephalum replicates and is maintained under non-selective conditions in two different lower eukaryotes; Kunzler P; The slime mould Physarum polycephalum contains 100 to 200 molecules of extrachromosomal linear DNA (PeDNA) . Two sets of the 19S and 26S ribosomal genes are located on each molecule of PeDNA . In the nonmitotic phase of the cell cycle PeDNA is localised in the nucleolus . The molecules are maintained throughout vegetative growth . In order to study the signals responsible for its maintenance, PeDNA was purified and introduced into the two distantly related yeasts Saccharomyces cerevisiae and Schizosaccharomyces pombe . Surprisingly, intact PeDNA transforms both yeasts with high frequency and PeDNA sequences are maintained in the absence of selective pressure.

J Gen Microbiol, 1985 Mar, 131 ( Pt 3), 527 - 32
The all2 gene is required for the induction of the purine deamination pathway in Schizosaccharomyces pombe; Fluri R et al.; Five mutants were isolated at the all2 gene on the basis of their inability to utilize hypoxanthine as a sole source of nitrogen . These mutants failed to utilize the purines adenine, hypoxanthine, xanthine, uric acid, allantoin and allantoic acid, although they could utilize urea and ammonium . The all2 mutants appeared to be defective in purine induction of uricase, allantoinase, allantoicase and ureidoglycollase activities but retained wild-type activity of the constitutively synthesized urease . The all2 mutations were recessive.

EMBO J, 1985 Mar, 4(3), 687 - 91
Molecular cloning and sequence analysis of a ras gene from Schizosaccharomyces pombe; Fukui Y et al.; We have cloned a ras gene homologue from fission yeast Schizosaccharomyces pombe and determined its nucleotide sequence . A putative coding sequence for 219 amino acids was found . The sequence contained one set of splicing signals: GTAAGT for a donor sequence, ACTAA for a unique sequence found in introns of yeast genes and TAG for an acceptor sequence, indicating the existence of an intron . The amino-terminal one third of the predicted S . pombe ras protein was nearly perfectly homologous and the next one third moderately homologous to those of mammalian ras proteins . The carboxy-terminal one third showed no homology but terminated with a short conserved sequence Cys-X-X-Z (X being a hydrophobic amino acid) as in other ras proteins . The result of Southern analysis of S . pombe DNA under nonstringent hybridization conditions using our clone as a probe indicated that no other closely related gene may be present in the S . pombe genome . The transcript of this gene could be detected by Northern analysis.

EMBO J, 1985 Feb, 4(2), 457 - 63
Cloning, sequencing and transcriptional control of the Schizosaccharomyces pombe cdc10 'start' gene; Aves SJ et al.; The cdc10 'start' gene from the fission yeast Schizosaccharomyces pombe has been cloned by rescue of mutant function . It is present as a single copy in the haploid genome . Hybridisation of the gene to Northern blots has identified a low abundance 2.7-kb polyadenylated RNA . Study of RNA extracted from cells both entering stationary phase and undergoing synchronous cell divisions suggests that commitment to the cell cycle is not controlled by regulation of cdc10 transcript level . DNA sequence analysis of the gene has identified an open reading frame capable of encoding a protein of mol . wt . 85 400 . The putative cdc10 gene product shows no significant primary structure similarity with products of other fission and budding yeast cell cycle genes, or with other protein sequences in several databases.

J Gen Microbiol, 1985 Feb, 131 ( Pt 2), 309 - 16
A correlation between mode of growth and regional ultrastructure of the plasma membrane of Schizosaccharomyces pombe as revealed by freeze-fracturing before and after filipin treatment; Takeo K; The ultrastructure of the plasma membrane of Schizosaccharomyces pombe was studied by freeze-fracture using invaginations of the plasma membrane as natural markers and filipin-induced deformations as artificial markers . In accord with the mode of growth of this organism, ultrastructural aspects of the plasma membrane were related to the following ring zones: the growing pole, adjacent regions, proximal regions, the new cell pole, and the middle in dividing cells . The growing pole and adjacent regions had no or only a few invaginations . Filipin induced numerous deformations in these regions . By contrast, the proximal regions of the plasma membrane had several invaginations and resisted filipin-induced deformation . Concomitantly with commitment to cytokinesis, both the invaginations and the resistance to filipin-induced deformation disappeared in the middle . The results presented here strongly suggest the existence of two states of the plasma membrane of S . pombe, a fact which correlates well with the mode of growth of this organism.

Proc Natl Acad Sci U S A, 1985 Feb, 82(4), 1113 - 5
Presence and source of free isopentenyladenosine in yeasts; Laten HM et al.; Cytokinins are a class of naturally occurring compounds that regulate growth and differentiation in tissues of higher plants . Many cytokinins are isopentenylated derivatives of adenine and its riboside, adenosine . By virtue of the post-transcriptional isopentenylation of specific anticodon loop adenosine residues in certain tRNA sequences, cytokinins are nearly universal, but tRNA-independent (de novo) cytokinin synthesis has been demonstrated in a few species . Using a radioimmunoassay, we have demonstrated that haploid strains of Saccharomyces cerevisiae and Schizosaccharomyces pombe contain, respectively, 0.8 and 0.9 microgram of the free cytokinin, N6-(delta 2-isopentenyl)adenosine, per g of cells (wet weight) . Strains of both species characterized by mutations that result in deficiencies of isopentenylated tRNAs have somewhat elevated levels of free N6-(delta 2-isopentenyl)adenosine . These findings lead to the conclusion that the major, if not exclusive, source of free cytokinins in these two yeasts is a synthetic pathway independent of isopentenylated RNA turnover.

Gene, 1985, 39(2-3), 223 - 30
Cloning and characterization of two genes restoring acid phosphatase activity in pho1- mutants of Schizosaccharomyces pombe; Maundrell K et al.; Schizosaccharomyces pombe acid phosphatase (APh) is a secreted cell surface glycoprotein which is deficient in pho1 mutants . By screening an S . pombe gene bank for sequences which can functionally rescue the pho1-44 mutation, we have isolated two genomic clones carried in plasmids pSp4B and pSp4C/2 . These two sequences map of different genetic loci and show no cross hybridization by Southern blotting . pSp4C/2 was found to contain the PHO1 gene, and cells transformed with this plasmid produce a protein which cross-reacts with antibodies raised against the protein moiety of APh . Data from Northern blotting experiments show that pSp4C/2 encodes a 1.6-kb transcript, and that mRNA levels are increased when cells are grown in low concentrations of inorganic phosphate . The results indicate that pSp4C/2 contains the structural gene for APh, PHO1, whereas pSp4B appears to carry a gene coding for a minor species of APh, PHO4 which is not regulated by extracellular phosphate.

Curr Genet, 1985, 9(2), 123 - 6
Rapid alkaline preparation for yeast circular covalently closed DNA molecules; Filetici P et al.; The alkaline preparation of prokaryotic plasmids (Birnboim and Doly, 1979) has been here adapted to yeast . By simple denaturation and renaturation steps we recovered, from Saccharomyces cerevisiae and Schizosaccharomyces pombe, a population of nucleic acid molecules highly enriched in circular forms . In S . cerevisiae killer strains it is possible to copurify double stranded RNA molecules . The overall recovery was estimated to be 10-30% of the total circular molecules.

Gene, 1985, 40(1), 125 - 30
Transcription of the triose-phosphate-isomerase gene of Schizosaccharomyces pombe initiates from a start point different from that in Saccharomyces cerevisiae; Russell PR; Gene tpi, encoding the glycolytic enzyme triose phosphate isomerase (TPI) from the fission yeast Schizosaccharomyces pombe was cloned by complementation of a Saccharomyces cerevisiae tpil mutant . Nucleotide sequence analysis of the cloned gene revealed a single open reading frame (ORF) encoding a protein 59% homologous to S . cerevisiae TPI . The gene has a very high codon usage bias . Messenger RNA synthesis initiates at two points located 38 and 44 nucleotides downstream from a TATA box promoter sequence . In S . cerevisiae, transcription of this S . pombe gene initiates about 26 nucleotides downstream from the S . pombe start points . This observation indicates that the two yeasts have diverged in the mechanism which determines the 5' end of the messenger RNA relative to the TATA box . It appears that in some respects the transcription initiation mechanism of S . pombe more closely resembles that of higher eukaryotes than does the S . cerevisiae mechanism.

Mol Gen Genet, 1985, 201(1), 82 - 7
First identification of an amber nonsense mutation in Schizosaccharomyces pombe: major differences in the efficiency of homologous versus heterologous yeast suppressor tRNA genes; Krupp G et al.; In Saccharomyces cerevisiae ochre and opal, as well as amber mutations are known, whereas in the fission yeast Schizosaccharomyces pombe no amber alleles have been described . We have characterized trp1-566, an amber allele in the trp1 locus of S . pombe . The identification of trp1-566 as an amber allele is based on the following results: (a) The nonsense allele can be converted to an ochre allele by nitrosoguanidine mutagenesis . (b) trp1-566 is suppressed by a bona fide S . pombe amber suppressor tRNA, supSI . The supSI gene was obtained by primer-directed in vitro mutagenesis of a tRNASer from S . pombe . Unexpectedly, an S . cerevisiae amber suppressor tRNASer, supR21, transformed into S . pombe, failed to suppress trp1-566 . Northern analysis of S . pombe transformants, containing supRL1 or S . cerevisiae tRNALeu or tRNATyr genes reveals that these genes are not transcribed in the fission yeast . As an additional tool for the analysis of nonsense mutations in S . pombe, we obtained by nitrosoguanidine mutagenesis two unlinked amber suppressor alleles, sup13 and sup14, which act on trp1-566.

Mol Gen Genet, 1985, 199(1), 152 - 3
Replicating instabilities in yeast: occurrence in different mutational systems; Nasim A et al.; Following mutagenesis of yeast cells with nitrosoguanidine, primary mosaic colonies exhibiting prototrophic/auxotrophic phenotypes were obtained . Upon replating of these primary mosaics, numerous secondary mosaics were present in the progeny . This study shows that replicating instabilities occur at many different loci within the Schizosaccharomyces pombe genome . In addition, the ade-1 gene of Saccharomyces cerevisiae (causing red pigmentation) was used to show that the phenomenon also occurs in this yeast.

Mol Gen Genet, 1985, 200(2), 252 - 7
Characterization of meiosis-deficient mutants by electron microscopy and mapping of four essential genes in the fission yeast Schizosaccharomyces pombe; Shimoda C et al.; Meiosis-deficient mutants of the fission yeast Schizosaccharomyces pombe carrying mei1, mei2, mei3, mei4 and mes1 mutant alleles were characterized by electron microscopy and staining of the nucleus with 4', 6-diamidino-2-phenylindole . Zygotes of either mei1, mei2 or mei3 mutants contained one round nucleus with a single spindle pole body (SPB) . These mutants were arrested before premeiotic DNA synthesis . Zygotes of mei4 mutants had one elongated nucleus containing thick electron-dense filaments (linear elements) . In the mes1 mutant, the first meiotic division was completed but the SBPs did not duplicate . Modification of the SPB (outer plaque formation) was also blocked and the forespore membrane was not assembled . By haploidization, random spore and tetrad analyses, four essential genes for meiosis (mei2, mei3, mei4 and mes1) were mapped . Gene mei2 was located on chromosome I 14.2 cM distant from ura2 . Gene mei3 was linked to ade7 (45.4 cM) on chromosome II . Gene mei4 was linked to cdc2 (0.6 cM) on chromosome II . Gene mes1 was linked to ura3 (25.3 cM) on chromosome I.

Mol Gen Genet, 1985, 199(3), 365 - 71
Direct selection of mutants influencing gene conversion in the yeast Schizosaccharomyces pombe; Thuriaux P; In Schizosaccharomyces pombe, a suppressor-active mutation at the anticodon site of the tRNASerUCA gene sup3 leads to opal (UGA)-specific suppression . Second-site mutations (rX) in sup3 inactivate the suppressor . The sup3-UGA, rX double mutants are genetically unstable in meiotic selfings, due to the intergenic transfer of information between sup3 and the unlinked genes sup9 and sup12 (Hofer et al . 1979; Munz and Leupold 1981; Munz et al . 1982) . These three genes have considerable sequence homology over about 200 base pairs (Hottinger et al . 1982) . Mutants showing a decrease or an increase of the meiotic instability at sup3 have been selected . One mutation (rec3-8) increases both the genetic instability and the frequency of intragenic recombination in sup3 by one order of magnitude . It has no effect on the stability of the nonsense alleles arg1-230 (UAA), ade6-704 and ural1-61 (UGA) or on the frequency of crossing-over between sup3 and the closely linked gene cdc8 . The existence of a common genetic control over intragenic recombination and genetic instability at sup3 provides a direct way of selecting for rec mutants in homothallic haploid strains of S . pombe carrying a suppressor-inactive allele of sup3 . It also supports the hypothesis that the instability of mutant alleles of this gene is due to chromosome mispairing at meiosis allowing sup3 to pair with sup9 or sup12 and then to undergo recombination by gene conversion restoring the suppressor-active allele sup3-UGA from the suppressor-inactive allele sup3-UGA, rX.(ABSTRACT TRUNCATED AT 250 WORDS)

Mol Gen Genet, 1985, 198(2), 360 - 3
The mitochondrial genome of the fission yeast Schizosaccharomyces pombe . 7 . Continuous gene for apocytochrome b in strain EF1 (CBS 356) and sequence variation in the region of intron insertion in strain ade 7-50h; Trinkl H et al.; The third BamHI fragment, containing most of gene for apocytochrome b, has been cloned and sequenced in the Schizosaccharomyces pombe strain EF1 (CBS 356) . In contrast to strain ade 7-50h- (50) from the Leupold collection, in which the gene is interrupted by an intron of group II (Lang et al . 1984), the homologous gene in strain EF1 is continuous . This demonstrates that the intron in the gene for apocytochrome b is optional . Aligning the EF1 sequence with the homologous regions in strain 50, 2 base pair changes were found in the leader and 14 in the coding region . These changes led to 12 altered triplets, but 9 of them specify the same amino acid . Seven base changes were clustered within a stretch of 30 base pairs in the region in which the intron is inserted in strain 50 . Five out of the resulting six triplet changes were also silent . These sequence variations around the highly conserved splice point region may be linked to the insertion or excision of the intron.

Gene, 1985, 39(2-3), 165 - 72
The RNA polymerase I initiation site and the external transcribed spacer of the fission yeast Schizosaccharomyces pombe ribosomal RNA genes; Balzi E et al.; A 5.45-kb fragment containing the 5' end of the ribosomal RNA transcriptional unit from the fission yeast Schizosaccharomyces pombe was cloned in the yeast-E . coli shuttle vector YEp13 . The transcription start point was mapped by R looping and S1 nuclease protection . The sequence of the entire external transcribed spacer (ETS) and its flanking regions was determined . Comparison of the sequence around the transcription start point with those of four budding yeasts (Saccharomycetoideae) reveals a consensus sequence from position -9 to -4 from the start . This sequence is likely to be an important element of the promoter for yeast RNA polymerase I (Pol.I) . Comparison of all known Pol.I promoter sequences reveals a strong bias for nucleotides (nt) at several positions between -16 and +10 . These nt may have a critical role in the transcription initiation process . The S . pombe ETS, which comprises 1355 bp, is significantly longer than those of the budding yeasts and lacks any significant sequence homology with the Saccharomyces cerevisiae ETS . R-loop analysis reveals a putative processing site within the ETS of S . pombe.

Int J Biochem, 1985, 17(7), 843 - 6
Purification and preliminary characterization of phosphoglycerate mutase from Schizosaccharomyces pombe; Price NC et al.; Phosphoglycerate mutase could be purified to over 95% homogeneity by a single step procedure involving elution from Cibacron Blue-Sepharose by a pulse of cofactor 2,3-bisphosphoglycerate . Although the enzyme has been isolated in only small quantities (c . 100 micrograms), gel filtration and sodium dodecylsulphate polyacrylamide gel electrophoresis indicated that it is monomeric with Mr approximately 23,000, an extremely low value for this enzyme . Preliminary investigations of the kinetic characteristics and the nature of important amino acid side chains have been undertaken.

Biol Cell, 1985, 55(1-2), 27 - 34
Electron microscopic studies of condensed mitotic chromosomes in the fission yeast Schizosaccharomyces pombe; Erard M et al.; By blocking cells in mitosis with the anti-fungal drug thiabendazole, it has been possible to carry out ultrastructural studies on the condensed chromosomes of the fission yeast, Schizosaccharomyces pombe . It is estimated that the DNA in these chromosomes is compacted approximately 1000-fold, and that the nucleoprotein density is similar to that of higher eukaryotic metaphase chromosomes . A basic structural component of the condensed chromosomes appears to be a 50-60 nm fibre, which is often visible in a loop configuration on the periphery of the chromatids . This is reminiscent of the 50-60 nm fibre loops which are frequently seen in preparations of metaphase chromosomes, and suggests that mechanisms of nucleoprotein folding may be similar in both lower and higher eukaryotes.

Biosystems, 1985, 18(3-4), 263 - 7
Probing fungal mitochondrial evolution with tRNA; Cedergren R et al.; Sequence data are now available for almost the entire complement of mitochondrial rRNAs from five fungi: Schizosaccharomyces pombe, Saccharomyces cerevisiae, Toropulis glabrata, Aspergillus nidulans and Neurospora crassa . Analysis of these data show that the five mitochondria can be related to a common ancestor . The unusually high similarity between some S . pombe mt tRNAs may be due to a process similar to gene conversion . Using the number of differences between tRNA pairs as a measure of the evolutionary rate the yeast-S . pombe branch has paradoxically a high nuclear rate and a low mt rate of evolution as compared with other branches in the phylogenetic tree . Finally the position of mt tRNA genes in S . pombe is abnormally distinct from gene orders in other mitochondria . All of the above factors must be taken into account when describing the relationship between these mitochondria.

Cell, 1984 Dec, 39(2 Pt 1), 349 - 58
The NDA3 gene of fission yeast encodes beta-tubulin: a cold-sensitive nda3 mutation reversibly blocks spindle formation and chromosome movement in mitosis; Hiraoka Y et al.; The cells of a cold-sensitive mutant nda3-KM311 of the fission yeast Schizosaccharomyces pombe were arrested highly synchronously at a step similar to mitotic prophase when incubated at a restrictive temperature . DAPI staining and indirect immunofluorescence microscopy showed three condensed chromosomes but no spindle . Six minutes after the temperature shifted to a permissive one, the spindle appeared and elongated . The chromosomes were separated at a constant speed (relative velocity 1 micron/min), and the spindle disappeared after the chromosomes reached opposite ends of the cell . The NDA3 gene of S . pombe was cloned by transformation . The 2.6 kb Hind III genomic DNA that complemented the nda3 mutations had only one coding frame split with five short introns . The predicted amino acid sequence contained 448 residues, and was 75% homologous to that of chicken beta-tubulin.

J Biochem (Tokyo), 1984 Nov, 96(5), 1511 - 23
O-Acetylhomoserine sulfhydrylase of the fission yeast Schizosaccharomyces pombe: partial purification, characterization, and its probable role in homocysteine biosynthesis; Yamagata S; A crude extract of Schizosaccharomyces pombe cells catalyzed sulfhydrylation of both O-acetyl-L-serine and O-acetyl-L-homoserine with H2S, but did not synthesize cystathionine from O-acetyl-L-homoserine and L-cysteine . The O-acetylhomoserine sulfhydrylase {EC 4.2.99.10} was very unstable; however, it could be stabilized by the addition of 25% (w/w) sucrose or glycerol . The optimal pH for activity was 8.0 and that for stability was 7.0 . The enzyme was purified approximately 300-fold from an ammonium sulfate-precipitated fraction . L-Methionine was the most effective inhibitor among the amino acids examined . It inhibited the enzyme competitively with respect to OAH with a Ki value of 2.6 mM . Sulfhydrylase activity was inhibited to various extents by some carbonyl reagents, but sulfhydryl reagents such as p-chloromercuribenzoic acid, 5,5'-dithio-bis(2-nitrobenzoic acid), and monoiodoacetic acid had no inhibitory effect . The enzyme also reacted with O-succinylhomoserine and L-homoserine to synthesize homocysteine directly, but could not utilize cysteine as a co-substrate in place of H2S . In the sulfhydrylation reactions, Km values for the substrates ranged from 10.4-12.5 mM . The enzyme was resolved to the apoenzyme by incubation with phenylhydrazine and reactivated by the addition of pyridoxal 5'-phosphate, whose Km value was 0.083 microM . The molecular weight of the enzyme was estimated to be approximately 186,000 by gel filtration and 170,000 by ultracentrifugation in sucrose density gradients . The isolectric point of the protein was pH 4.1 . The characteristics of this enzyme are compared with those of physiologically functional sulfhydrylases reported for other organisms, and the possibility of the enzyme functioning as a homocysteine synthase is discussed.

Gene, 1984 Nov, 31(1-3), 129 - 34
Sequence of the cell division gene CDC2 from Schizosaccharomyces pombe; patterns of splicing and homology to protein kinases; Hindley J et al.; The complete nucleotide sequence of a 2.9-kb DNA fragment containing the CDC2 gene-complementing activity from Schizosaccharomyces pombe has been determined . Within this region lies a 1.69-kb DNA sequence whose predicted amino acid sequence shows extensive homology to that previously deduced for the CDC28 gene product from Saccharomyces cerevisiae {Lorincz and Reed, Nature 307 (1984) 183-185} . Taken with the earlier observation that mutants in CDC2 can be rescued by the presence of the CDC28 gene {Beach, Durkacz and Nurse, Nature 300 (1982) 706-709}, these results strongly suggest that the two genes code for similar functions . In contrast to the CDC28 gene, however, which contains no introns, the CDC2 coding sequence is split by four introns and from a comparison of the two sequences a consensus sequence for intron splicing in S . pombe can be established . Both CDC2 and CDC28 contain the consensus sequences for the ATP binding and phosphorylation acceptor sites of protein kinases such as bovine cAMP-dependent protein kinase (bov PK) and the src family of viral oncogene products.

Nature, 1984 Nov 1-7, 312(5989), 61 - 3
Cell size control of development in Saccharomyces cerevisiae; Calvert GR et al.; Diploid cells of the yeast Saccharomyces cerevisiae in the G1 phase of the cell cycle are faced with the alternatives of either continuing vegetative cell division or undergoing the developmental processes of meiosis and subsequent ascospore formation, or adapting to starvation conditions if these apply . The course taken is influenced by the nutritional status of the culture medium, the presence of both MATa and MAT alpha mating-type alleles, and the need for cells to be in the G1 phase of the cell cycle . For those cells that continue cell division, size controls operate in both the budding yeast S . cerevisiae and the fission yeast Schizosaccharomyces pombe . In S . cerevisiae the 'start' event initiating the cell cycle is controlled in some way related to cell size because cells below a critical size fail to initiate cell division . The ability of cells to undergo the developmental process of sporulation is related to cell age, in that cells gain this ability just before the emergence of the first bud and the process of sporulation after initiation is altered in small cells . Here we report that the initiation of sporulation is subject to a size control related to absolute cell volume, which is distinct from control by cell age and also independent of the control operating on the initiation of cell division.

EMBO J, 1984 Sep, 3(9), 2129 - 36
The mitochondrial genome of the fission yeast Schizosaccharomyces pombe: highly homologous introns are inserted at the same position of the otherwise less conserved cox1 genes in Schizosaccharomyces pombe and Aspergillus nidulans; Lang BF; The DNA sequence of the second intron in the mitochondrial gene for subunit 1 of cytochrome oxidase (cox1), and the 3' part of the structural gene have been determined in Schizosaccharomyces pombe . Comparing the presumptive amino acid sequence of the 3' regions of the cox1 genes in fungi reveals similarly large evolutionary distances between Aspergillus nidulans, Saccharomyces cerevisiae and S . pombe . The comparison of exon sequences also reveals a stretch of only low homology and of general size variation among the fungal and mammalian genes, close to the 3' ends of the cox1 genes . The second intron in the cox1 gene of S . pombe contains an open reading frame, which is contiguous with the upstream exon and displays all characteristics common to class I introns . Three findings suggest a recent horizontal gene transfer of this intron from an Aspergillus type fungus to S . pombe . (i) The intron is inserted at exactly the same position of the cox1 gene, where an intron is also found in A . nidulans . (ii) Both introns contain the highest amino acid homology between the intronic unassigned reading frames of all fungi identified so far (70% identity over a stretch of 253 amino acids) . However, in the most homologous region, a GC-rich sequence is inserted in the A . nidulans intron, flanked by two direct repeats of 5 bp . The 37-bp insert plus 5 bp of direct repeat amounts to an extra 42 bp in the A . nidulans intron . (iii) TGA codons are the preferred tryptophan codons compared with TGG in all mitochondrial protein coding sequences of fungi and mammalia.(ABSTRACT TRUNCATED AT 250 WORDS)

EMBO J, 1984 Sep, 3(9), 2121 - 8
Three variant introns of the same general class in the mitochondrial gene for cytochrome oxidase subunit 1 in Aspergillus nidulans; Waring RB et al.; The oxiA gene of Aspergillus nidulans, coding for cytochrome oxidase subunit 1, is shown by DNA sequencing to contain three introns . An AUG start codon is not present at the beginning of the sequence, suggesting that either another codon, possibly the four base codon AUGA, is used for initiation or there is a further short intron between the true start codon and the beginning of the recognisable coding region . The second and third introns have long open reading frames, which could code for maturase proteins . The lack of conservation of amino acid sequence in the putative region of proteolytic cleavage for maturase formation suggests that the first conserved decapeptide may act as the recognition signal for protein processing . The third intron is remarkably (70%) homologous to the second intron of the cytochrome oxidase subunit 1 gene of Schizosaccharomyces pombe and both are located in exactly the same position . The third Aspergillus intron has an in-frame insertion of a 37-bp GC-rich DNA sequence which is now flanked by a 5-bp repeat, a well-known feature of transposable elements . All three introns in the oxiA gene have a 'core' RNA secondary structure found in a class of introns fitting the RNA splicing model of Davies et al . (1982) . This core RNA structure may play a catalytic as well as a structural role in intron splicing . A sequence within the intron could act as a guide to align the splice sites of two of the introns in accordance with the model of Davies et al.

EMBO J, 1984 Aug, 3(8), 1737 - 44
Isolation of type I and II DNA topoisomerase mutants from fission yeast: single and double mutants show different phenotypes in cell growth and chromatin organization; Uemura T et al.; We have isolated mutants defective in DNA topoisomerases and an endonuclease from the fission yeast Schizosaccharomyces pombe by screening individual extracts of mutagenized cells . Two type I topoisomerase mutants (top1) and three endonuclease mutants (end1) were all viable . The double mutant top1 end1 was also viable and, in its extract, Mg2+- and ATP- dependent type II activity could be detected . Three temperature-sensitive (ts-) mutants having heat-sensitive (hs-) type II enzymes were isolated, and the ts- marker cosegregated with the hs- type II activity . All the ts- mutations fell in one gene (top2) tightly linked to leul in chromosome II . The nuclear division of single top2 mutants was blocked at the restrictive temperature, but the formation of a septum was not inhibited so that the nucleus was cut across with the cell plate . In contrast, the double top1 top2 mutants were rapidly arrested at various stages of the cell cycle, showing a strikingly altered nuclear chromatin region . The type II topoisomerase may have an essential role in the compaction and/or segregation of chromosomes during the nuclear division but also complement the defect of the type I enzyme whose major function is the maintenance of chromatin organization throughout the cell cycle.

J Cell Sci, 1984 Jul, 69, 47 - 65
Septum pattern in ts mutants of Schizosaccharomyces pombe defective in genes cdc3, cdc4, cdc8 and cdc12; Streiblova E et al.; Septum-defective mutants of Schizosaccharomyces pombe impaired in cdc genes 3, 4, 8 and 12 were compared by fluorescence microscopy, freeze-etching and ultrathin sectioning . This approach made it possible to recognize the internal organization of defective phenotypes under restrictive conditions . Of special interest in this study was the pattern of unusual septum malformations found to be regular features of the terminal phenotypes of the mutants . Their overall topology was visualized at the cellular level by primulin fluorescence . The subcellular location of septum defects was found to be identical in origin to the compartment where normal septum was assembled in the wild type . Delocalized septation involved both microfibrillar and matrix components, which participated in the final assembly of malformations . Unique contour views of delocalized septa were exposed by freeze-fracturing . Cytoplasmic microtubules and microfilaments were detected in ultrathin sections of the cytoplasm of mutant cells . The internal organization of malformation-accumulating phenotypes suggested a disruption of the directional mechanism that steers septum material to the periplasm at the cell equator.

J Cell Sci, 1984 Jul, 69, 199 - 210
Protein synthesis and its relation to the DNA-division cycle in the fission yeast Schizosaccharomyces pombe; Creanor J et al.; The rate of protein synthesis has been measured with pulse labels of {3H}tryptophan in synchronous and asynchronous cultures of cdc mutants of Schizosaccharomyces pombe shifted up to the restrictive temperature . The cell cycle related fluctuations in rate that occur in normal synchronous cultures vanish when nuclear division is blocked in synchronous cultures of cdc2 and cdc10 . But they persist in cdc11 where nuclear division continues and cleavage is stopped . We conclude that nuclear division affects the rate of synthesis and that this effect is inhibitory and probably persists for the last 40% of the cycle . When nuclear division has been blocked, the rate of synthesis continues to increase until a plateau is reached where the rate remains constant . Three size mutants of cdc2 reach the plateau at the same average protein content per cell although their initial protein contents vary over a threefold range . Comparison of these results with those from cdc10 leads to the tentative conclusion that the plateau starts when the cells reach a critical protein/DNA ratio.

EMBO J, 1984 Jul, 3(7), 1573 - 80
Mutations affecting excision of the intron from a eukaryotic dimeric tRNA precursor; Willis I et al.; The nucleotide sequences of a Schizosaccharomyces pombe opal suppressor serine tRNA gene (sup9-e) and of 12 in vivo-generated mutant genes, which have lost the ability to suppress UGA mutations, have been determined . Analysis of the expression of these genes in Saccharomyces cerevisiae in vitro and in vivo systems has revealed defects in tRNA gene transcription and precursor tRNA processing . Single base changes in the D-loop, the intron and the extra arm affect the efficiency of splicing of the tRNA precursors while an anti-codon stem mutation may affect the accuracy of this process . Two mutations which occur in the intervening sequence of the sup9-e gene allow an alternate tRNA base pairing configuration . Transcription of the sup9-e gene and of the adjacent tRNAMet gene (located 7 bp downstream) is essentially abolished in vivo by a G----A19 mutation in the tRNASer gene, suggesting that tRNAMet may be derived solely via processing of the tRNASer-tRNAMet dimeric precursor.

Biochim Biophys Acta, 1984 Jun 19, 804(2), 221 - 9
Dielectrophoretic properties of yeast cells dividing by budding and by transversal fission; Iglesias FJ et al.; The dielectrophoretic behaviour of yeast cells dividing by budding or by transversal fission was analyzed . The results obtained show that the dielectrophoretic yield is a linear function of alternating voltage, cell concentration and the square root of the time of collection in all the species assayed . Dependence of the rate of collection on the frequency of the voltage applied (between 0.2 and 5 MHz) was also found . This behaviour is similar in the three microorganisms studied . The scale factor correlating the frequency spectrum for Saccharomyces cerevisiae and Saccharomycopsis lipolytica is proportional to cell size . However, these results can not be extended to Schizosaccharomyces pombe . A relationship between the dielectrophoretic yield and the age of the culture and the consumption of glucose has been established for the three yeast strains . Dielectrophoresis also permits the differentiation between viable and non-viable cells.

Proc Natl Acad Sci U S A, 1984 Jun, 81(11), 3481 - 5
Genes required for initiation and resolution steps of mating-type switching in fission yeast; Egel R et al.; The fission yeast Schizosaccharomyces pombe switches mating type by transposition of a copy of DNA derived from either of the two storage cassettes, mat2 -P and mat3 -M, into the expression locus, mat1 . The recombinational event of switching is initiated by a double-stranded DNA break present in approximately 20% of the molecules at mat1 . Fifty-three mutants defective in switching of mating type have been isolated previously, and each has been assigned to 1 of 10 linkage groups . One group consists of cis-acting mutations at mat1 , which reduce the amount of the DNA double-strand cut . The remaining nine groups are mutations in genes that are unlinked to the mating-type locus and are studied here . Three ( swi1 , -3, -7) are required for formation of the double-strand cut, whereas the others are not . Mutants of three genes ( swi4 , -8, -9) undergo high-frequency rearrangement of the mating-type locus indicative of errors of resolution of recombinational intermediates . The remaining three ( swi2 , -5, -6) have normal levels of cut, do not make errors of resolution, and possibly are required either for efficient utilization of the cut or determining the directionality of switching . The data suggest that the switching process can be dissected into genetically distinguishable steps.

Cell, 1984 May, 37(1), 233 - 42
Identification of the pleiotropic cell division cycle gene NDA2 as one of two different alpha-tubulin genes in Schizosaccharomyces pombe; Toda T et al.; Mutations in a cell-cycle gene NDA2 of Schizosaccharomyces pombe have pleiotropic effects on nuclear division, nuclear location, and thiabendazole sensitivity ( Toda et al., 1983) . By transformation and nucleotide sequence determination, we identified NDA2 as one of two alpha-tubulin genes present in the genome of S . pombe . Two cloned sequences complemented cold-sensitive and thiabendazole-supersensitive nda2 mutations; one was derived from NDA2 that encodes alpha 1-tubulin, the other from an unidentified locus encoding alpha 2-tubulin . The predicted amino acid sequences showed that the alpha 1- and alpha 2-tubulins had respective residues of 455 and 449 (molecular weights 51,200 and 50,600) . The homology to porcine alpha-tubulin was 76% in both cases . Frequent alterations took place in the two restricted regions . The alpha 1-tubulin (NDA2) clone had a 90 bp intervening sequence, the alpha 2-tubulin clone did not . RNA blot hybridization experiments indicated that both genes are transcribed . S . pombe tubulin was isolated by cycles of assembly and disassembly . Presumed alpha- and beta-tubulin polypeptide bands reacted with monoclonal antibodies specific for chicken alpha- and beta-tubulins.

Mol Cell Biol, 1984 Apr, 4(4), 651 - 6
High-frequency cotransformation by copolymerization of plasmids in the fission yeast Schizosaccharomyces pombe; Sakai K et al.; We have developed a high-frequency cotransformation system which is useful in introducing nonreplicating circular DNA plasmids into the fission yeast Schizosaccharomyces pombe . This system depends on two factors: the ability of the ural-complementing helper plasmids pFYM2 and pFYM225 to propagate autonomously in S . pombe, and the intensive recombination activity intrinsic to this yeast . If cotransformed with a helper plasmid, plasmids such as YIp5 or YIp32, Escherichia coli-Saccharomyces cerevisiae shuttle vectors incapable of replication in S . pombe, can enter S . pombe and express the gene carried on them at a frequency comparable to that of autonomously replicating plasmids (10(3) to 10(4) transformants per microgram of DNA) . Even if characters of the nonreplicating DNA are not selected directly, 50 to 70% of Ura+ cells transformed with the helper have also incorporated the nonreplicating plasmid . It is shown that these two plasmids have physically recombined at a site of common DNA sequence to form a heteropolymer in the fission yeast . Since any foreign DNA cloned in pBR322 or ColE1 derivatives can be incorporated into S . pombe by using pFYM2 or pFYM225 as a helper, this cotransformation system will serve as a convenient method to examine functional expression of such cloned DNA in S . pombe . This work also demonstrates that the kanamycin resistance gene carried by the bacterial transposon Tn903 can be expressed in S . pombe, as shown by its ability to inactivate the antibiotic G418.

J Biol Chem, 1984 Mar 10, 259(5), 2856 - 62
An antisuppressor mutation of Schizosaccharomyces pombe affects the post-transcriptional modification of the "wobble" base in the anticodon of tRNAs; Heyer WD et al.; The screening of antisuppressor mutants of the yeast Schizosaccharomyces pombe has been successfully accomplished with high resolution liquid chromatographic methods for the analysis of tRNA nucleosides . Antisuppressor mutations reduce or abolish the function of nonsense suppressor-tRNAs or other informational suppressors . Nonradioactive or 35S-labeled unfractionated tRNA from various strains was digested to nucleosides and analyzed by high performance liquid chromatography . The mutant sin3 has lost the nucleoside 5-(methoxycarbonylmethyl)-2-thiouridine from its tRNA in comparison to parental strains . In eukaryotes this nucleoside is found at the first position of the anticodon (wobble position) in several isoacceptor tRNAs that preferentially recognize codons ending with adenosine . The sin3 mutation reduces the efficiency of UGA and UAA suppressor tRNASer and suppressor tRNALeu . The genetic cosegregation of modification loss, antisuppressor phenotype, and a change in cell size is demonstrated . This indicates that a single mutation in the structural gene for a tRNA modification enzyme causes the three different phenotypes.

J Biol Chem, 1984 Mar 10, 259(5), 2845 - 9
Independent loci for the structural genes of the yeast mitochondrial alpha and beta ATPase subunits; Vassarotti A et al.; In the yeast Schizosaccharomyces pombe, the structural gene mutations A23-13 (alpha-) and B59-1 (beta-) which totally prevent the expression of either the alpha or the beta subunits of the mitochondrial ATPase, were shown by classical genetic mapping studies to be both located on chromosome I but genetically unlinked . It is concluded that the structural genes ATP1 and ATP2 for the alpha and beta subunits of the mitochondrial ATPase are not organized in a cluster . By both meiotic recombination frequency analysis and gene transfer studies, three single nuclear mutations affecting to different extents the electrophoretic mobility of the beta polypeptide were located on the chromosome I very close to the mutation B59-1 (beta-) . Two mutations involved a defective ATPase activity and the inability to grow on glycerol (gly) . One of these mutants E5-23 (beta") exhibited a beta subunit of slightly reduced electrophoretic mobility . The other mutation F1-10 (beta) was associated with a beta subunit of normal electrophoretic mobility . The plasmid pMa2 (Boutry, M., Vassarotti, A., Ghislain, M., Douglas, M., Goffeau, A . (1984) J . Biol . Chem . 259, 2840-2844) containing the structural gene for the beta subunit complemented the mutants E5-23 (beta") and F1-10 (beta) as well as B59-1 (beta-) . These three mutations are therefore likely to affect the beta structural gene itself or a very contiguous gene contained in the 5.4-kilobase genomic insert of pMa2 . The mutation F1-10 (beta) was mapped between E5-23 (beta") and B59-1 (beta-) by analysis of the meiotic recombination frequencies . Another mutation F25-28-11 (beta') was responsible for an appreciable decrease of electrophoretic mobility of the beta subunit which, however, did not affect either the ATPase activity or the ability to grow on glycerol (GLY) . This mutant transformed by pMa2 was able to express the structural gene for the wild type beta subunit and the resulting transformants synthesized and assembled both the beta and beta' subunits . It is concluded that the mutation F25-28-11 (beta') also affects the structural gene for the beta subunit and does not affect genes controlling the processing machinery.

J Biol Chem, 1984 Mar 10, 259(5), 2840 - 4
Isolation of the structural genes for the alpha and beta subunits of the mitochondrial ATPase from the fission yeast Schizosaccharomyces pombe; Boutry M et al.; The structural genes for the two major subunits of the mitochondrial ATPase were isolated among genomic clones from the yeast Schizosaccharomyces pombe by transformation and complementation of mutants unable to grow on glycerol and lacking either the alpha or the beta subunits . The plasmid pMa1 containing a 2.3-kilobase genomic insert transformed the mutant A23-13 lacking a detectable alpha subunit . The transformant grew on glycerol and contained an alpha subunit of normal electrophoretic mobility . The plasmid pMa2 containing a 5.4-kilobase genomic insert transformed the mutant B59-1 lacking the beta subunit . The transformant grew on glycerol and contained a beta subunit of normal mobility . The structural gene for the beta ATPase subunit for the fission yeast S . pombe was localized within the pMa2 insert by hybridization to a probe containing the beta ATPase gene from the budding yeast Saccharomyces cerevisiae (Saltzgaber, J., Kunapuli, S., and Douglas, M . G . (1983) J . Biol . Chem . 258, 11465-11470) . The mRNAs which hybridized to pMa1 and pMa2 were translated by a reticulocyte lysate into polypeptides of Mr = 59,000 and 54,000, respectively . These genes products reacted with an anti-F1-ATPase serum and therefore correspond most probably to precursors of the alpha and beta subunits.

EMBO J, 1984 Mar, 3(3), 603 - 10
Rearrangements of the transposable mating-type cassettes of fission yeast; Beach DH et al.; The fission yeast, Schizosaccharomyces pombe, switches mating type every few cell divisions . Switching is controlled by the genes of the mating-type locus, which consists of three components, mat1, mat2-P and mat3-M, each separated by approximately 15 kb . Copy transposition of P (Plus) or M (Minus) information from mat2-P or mat3-M into the expression locus mat1 mediates cell type switching . The mating-type locus undergoes events at high frequency (10(-2)-10(-6)) which stabilize one or other mating type . These events are shown to be rearrangements which result in either deletion or insertion of DNA between cassettes.

Stain Technol, 1984 Mar, 59(2), 79 - 82
The use of primuline to identify the septum polysaccharide of the fission yeast Schizosaccharomyces pombe; Duffus JH et al.; Treatment of cells and purified cell walls of the fission yeast Schizosaccharomyces pombe with primuline reveals the septum as a bright fluorescent band . When polysaccharides containing (1----3)-beta-, (1----6)-beta- or (1----3)-alpha-glucosidic linkages are treated with primuline, only those molecules containing chains of (1----3)-beta-glucosyl residues are stained . This implies that (1----3)-beta-glucan is present in the septum of Schiz . pombe as the main constituent.

Mutat Res, 1984 Feb, 125(2), 205 - 11
The effect of spermine on spontaneous and UV-induced mutations in Schizosaccharomyces pombe; Prendergast JA et al.; The effect of different concentrations of spermine on spontaneous and UV-induced mutation in the adenine forward mutation system of Schizosaccharomyces pombe was investigated . The effect of spermine on spontaneous mutation was studied in 5 mutator strains (mut 1-4, mut 1-23, mut 2-9, mut 2-20 and mut 3-21) and on UV-induced mutation in a pigmented adenine-requiring strain and its radiation-sensitive derivative (rad 13) . The effect of spermine exposure on mutation induction before and after UV irradiation was also investigated . Spermine increased spontaneous forward mutation in the mut 1-4 strain by 47%, and enhanced UV-induced forward mutation 2-fold in the rad 13 and normal pigmented strains . No antimutagenic effect of spermine was seen in any of the strains tested . This is in marked contrast to the antimutagenic effect of spermine observed with bacteria.

EMBO J, 1984 Feb, 3(2), 423 - 8
The Schizosaccharomyces pombe sup3-i suppressor recognizes ochre, but not amber codons in vitro and in vivo; Hottinger H et al.; The inefficient suppressor sup3-i of the fission yeast Schizosaccharomyces pombe is an ochre suppressor . Sup3-i was derived from the efficient serine inserting UGA suppressor sup3-e . The cloning and sequencing of the sup3-i gene indicate that the suppressor is different from the parent sup3-e by a C----T substitution in the sequence coding for the middle position of the anticodon . In vitro translation assays supplemented with purified sup3-i tRNA and programmed with Xenopus globin mRNAs lead to the accumulation of a readthrough product in response to UAA termination signals, but not in response to UGA termination codons . Transformation of Saccharomyces cerevisiae nonsense mutant strains with plasmid DNA carrying the S . pombe sup3-i gene, led to ochre, but not amber or UGA suppression in vivo.

Biochem J, 1984 Jan 15, 217(2), 585 - 8
A study of the maloalcoholic fermentation pathway in Schizosaccharomyces pombe; Maconi E et al.; The pathway of the maloalcoholic fermentation in Schizosaccharomyces pombe was investigated by a 1H-, 2H- and 13C-n.m.r.-spectroscopic study of hydrogen and deuterium distribution on the ethanol produced by S . pombe from L-malic acid in 2H2O and from L-{2-2H}malic acid . Our findings rule out a double-decarboxylation mechanism and agree with a pathway that involves acetaldehyde as intermediate.

Mikrobiologiia, 1984 Jan-Feb, 53(1), 48 - 9
{Cellular inequivalence in a culture of Schizosaccharomyces pombe}; Vrana D; The fission yeast Schizosaccharomyces pombe was grown in the chemostat at D = 0.03, 0.05, 0.1, 0.15 and 0.20 h-1 . The dry weight and substrate quantities, the number of cells and their morphological characteristics were determined in the steady state . The curves for the cell number and dry weight demonstrate changes in the coordination between the processes of cell growth and division at various growth rates . The cell division was shown to be asymmetric under the conditions of substrate limitation.

Mol Gen Genet, 1984, 197(3), 447 - 52
The sup8 tRNALeu gene of Schizosaccharomyces pombe has an unusual intervening sequence and reduced pairing in the anticodon stem; Sumner-Smith M et al.; We have cloned and sequenced the wild-type and suppressor alleles of the S . pombe sup8 tRNA gene . The wild-type allele has a leucine UAA anticodon and the suppressor (sup8-e) carries the opal suppressor anticodon UCA . The gene has a 16 base pair intervening sequence that, in the RNA, is predicted to form a secondary structure which involves base pairing to the 5', rather than the usual 3' side of the 5' splice site . When incubated in Saccharomyces cerevisiae cell-free extracts both alleles are efficiently transcribed, the 5' leader and 3' trailer sequences are removed and CCA is added to the 3' processed end; however, the intervening sequence is not excised . This finding implies that the structural requirements of the splicing endonucleases in the two yeasts have diverged . No other tRNA genes with related sequences were detected in S . pombe DNA by hybridization, suggesting that other UUA isoacceptors may be structurally dissimilar to sup8 or that the UUA codon may be decoded by a UUG leucine isoacceptor.

J Bacteriol, 1984 Jan, 157(1), 334 - 6
A meiotic mutant of the fission yeast Schizosaccharomyces pombe that produces mature asci containing two diploid spores; Nakaseko Y et al.; A mutant of the fission yeast Schizosaccharomyces pombe grew normally in the mitotic cycle but produced two-spored asci in the meiosis cycle . These spores were diploid, and the segregation of centromere-linked markers in the dyads was mostly reductional . Only the first meiotic division appears to occur in this tws1 mutant, resulting in enclosure of diploid nuclei into spores.

Mol Gen Genet, 1984, 196(3), 473 - 81
The mitochondrial genome of the fission yeast Schizosaccharomyces pombe . 3 . Gene mapping in strain EF1 (CBS 356) and analysis of hybrids between the strains EF1 and ade7-50h-; Zimmer M et al.; The Schizosaccharomyces pombe strain EF1 (CBS 356) is haploid, prototrophic, respiratory competent, and of homothallic mating type . From restriction enzyme analysis the length of the mitochondrial genome is 17.3 kilobase pairs, which is in good agreement with the value of 17.1 kilobase pairs determined by electron microscopy . The mitochondrial genome of strain EF1 is thus about 2.3 kilobase pairs shorter than that of strain ade7-50h- (about 19.4 kilobase pairs) . A restriction map was constructed for 11 enzymes: For most, but not all of them, the pattern is nearly identical to that of strain ade7-50h- . The genes for the large ribosomal RNA, the subunits 1, 2, and 3 of cytochrome c oxidase, subunits 6 and 9 of ATP synthetase, and cytochrome b were localized by hybridization with mitochondrial DNA probes from Saccharomyces cerevisiae . The gene order was found to be the same in both yeast strains . From Southern hybridization of strain ade7-50h- with nick-translated mitochondrial DNA from strain EF1 it is evident that strain EF1 does not possess the intron, which is present in the cytochrome b gene of Schizosaccharomyces pombe strain ade7-50h- . Crosses between strain ade7-50h- and EF1 demonstrate that both the nuclear and the mitochondrial genomes are able to recombine . The mitochondrial genomes of 2 out of 30 independently isolated hybrids between the two strains are described as the result of recombination between the two parental mitochondrial genomes.

Mol Gen Genet, 1984, 196(3), 465 - 72
The mitochondrial genome of the fission yeast Schizosaccharomyces pombe . 2 . Localization of genes by interspecific hybridization in strain ade7-50h- and cloning of the genome in small fragments; Lang BF et al.; A series of 18 small overlapping restriction fragments has been cloned, covering the complete mitochondrial genome of Schizosaccharomyces pombe . By hybridizing mitochondrial gene probes from Saccharomyces cerevisiae and Neurospora crassa with restriction fragments of Schizosaccharomyces pombe mitochondrial DNA, the following homologous genes were localized on the mitochondrial genome of S . pombe: cob, cox1, cox2 and cox3, ATPase subunit 6 and 9 genes, the large rRNA gene and both types of open reading frames occurring in mitochondrial introns of various ascomycetes . The region of the genome, hybridizing with cob exon probes is separated by an intervening sequence of about 2500 bp, which is homologous with the first two introns of the cox1 gene in Saccharomyces cerevisiae (class II introns according to Michel et al . 1982) . Similarly, in the cox1 homologous region, which covers about 4000 bp, two regions were detected hybridizing with class I intron probes, suggesting the existence of two cox1 introns in Schizosaccharomyces pombe . Hybridization with several specific exon probes with a determined order has revealed that cob, cox1, cox3 and the large rRNA gene are all transcribed from the same DNA strand . The low intensities of hybridization signals suggest a large evolutionary distance between Schizosaccharomyces pombe and Saccharomyces cerevisiae or Neurospora crassa mitochondrial genes . Considering the length of the mitochondrial DNA of Schizosaccharomyces pombe (about 19.4 kbp) and the expected length of the localized genes and intron sequences there is enough space left for encoding the expected set of tRNAs and the small rRNA gene . The existence of leader-, trailer-, ori- and spacer sequences or further unassigned reading frames is then restricted to a total length of about 3000 bp only.

Mol Gen Genet, 1984, 195(1-2), 164 - 9
Analysis of the structure and transcription of the aro3 cluster gene in Schizosaccharomyces pombe; Nakanishi N et al.; By selecting activities to complement Escherichia coli aro mutations, a gene responsible for the biosynthesis of aromatic amino acids in Schizosaccharomyces pombe (aro3) was cloned into pBR322 . Three independent clones named pFNA1, pFNA2 and pFNA3 were obtained . pFNA1 could complement E . coli aroD only, whereas the other two plasmids were able to complement both aroD and aroE . The aro3 locus of S . pombe was found to be a gene cluster which can be subdivided into five complons, A through E (Strauss 1979) . Transformation of S . pombe mutants defective in each complon with the pFNA plasmids indicated that pFNA1 carries at least a part of aro3A, the whole of B, C and D, as well as a part of E, whereas pFNA2 and pFNA3 carry a part of aro3C and the entire D and E complons . A physical map of the aro3 locus was constructed by analyzing the structure of these plasmids and their hybridization patterns to restricted genomic DNA . A transcript 4.5 kb long was detected as the sole aro3 mRNA encompassing all five complons . This study, in addition to the work of Strauss (1979), establishes that the arrangement of the aro3 complons in S . pombe in terms of their enzymatic coding activities is identical with that of Neurospora crassa, but different from that proposed for Saccharomyces cerevisiae.

J Biol Chem, 1983 Dec 25, 258(24), 15214 - 9
Complementation of a Schizosaccharomyces pombe mutant lacking the beta subunit of the mitochondrial ATPase by the ATP2 gene of Saccharomyces cerevisiae; Boutry M et al.; A chimeric plasmid carrying the structural gene (ATP2) for the mitochondrial ATPase beta subunit of Saccharomyces cerevisiae has been used to complement a mutant of Schizosaccharomyces pombe lacking the beta subunit (Boutry, M., and Goffeau, A . (1982) Eur . J . Biochem . 125, 471-477) . Transformation with ATP2 restored the growth rate of S . pombe mutant on glycerol as well as the mitochondrial ATPase and 32Pi-ATP exchange activities to approximately 20% of the parental strain . Mitochondria prepared from the transformant contained a normal amount of a hybrid F1-ATPase consisting of the S . cerevisiae beta subunit assembled with the remaining subunits of the S . pombe ATPase complex . The presence of the S . cerevisiae beta subunit in the S . pombe ATPase complex conferred a sensitivity to the energy transfer inhibitors citreoviridin and oligomycin which was like that of the intact S . cerevisiae enzyme . The S . cerevisiae beta subunit assembled into the hybrid ATPase complex was the same size as the mature subunit in S . cerevisiae . These data indicate that the mechanism of mitochondrial import and the assembly of the cytoplasmically synthesized subunits is similar or identical in these evolutionary divergent yeasts . In addition, this study provides a new approach for the construction of hybrid mitochondrial ATPase complexes which can be used to examine the function of selected subunits in energy transduction.

Nucleic Acids Res, 1983 Dec 20, 11(24), 8537 - 46
Six Schizosaccharomyces pombe tRNA genes including a gene for a tRNALys with an intervening sequence which cannot base-pair with the anticodon; Gamulin V et al.; We report the sequences of six S . pombe tRNA genes including two genes for tRNAArg, and one gene each for tRNAGlu, tRNAHis, tRNALys and tRNAPhe . All tRNA genes are found independently in the genome and represent individual transcription units . The gene for tRNALys has an 8 bp long intervening sequence which cannot base-pair with the tRNA anticodon . In vitro transcription studies indicate that all genes are faithfully transcribed in a yeast extract . Sequence comparison of the 5' flanking regions of the tRNA genes did not show significant homologies; however, they are very rich in AT base pairs.

Mol Cell Biol, 1983 Nov, 3(11), 1898 - 908
Isolation and characterization of sequences from mouse chromosomal DNA with ARS function in yeasts; Roth GE et al.; Fragments of chromosomal DNA from a variety of eucaryotes can act as ARSs (autonomously replicating sequence) in yeasts . ARSs enable plasmids to be maintained in extrachromosomal form, presumably because they function as initiation sites for DNA replication . We isolated eight different sequences from mouse chromosomal DNA which function as ARSs in Saccharomyces cerevisiae (bakers' yeast) . Although the replication efficiency of the different mouse ARSs in yeasts appears to vary widely, about one-half of them functions as well as the yeast chromosomal sequence ARS1 . Moreover, five of the ARSs also promote self replication of plasmids in Schizosaccharomyces pombe (fission yeast) . Each of the ARSs was cloned into plasmids suitable for transformation of mouse tissue culture cells . Plasmids were introduced into thymidine kinase (TK)-deficient mouse L cells by the calcium phosphate precipitation technique in the absence of carrier DNA . In some experiments, the ARS plasmid contained the herpes simplex virus type 1 TK gene; in other experiments (cotransformations), the TK gene was carried on a separate plasmid used in the same transformation . In contrast to their behavior in yeasts, none of the ARS plasmids displayed a significant increase in transformation frequency in mouse cells compared with control plasmids . Moreover, only 1 of over 100 cell lines contained the original plasmid in extrachromosomal form . The majority of cell lines produced by transformation with an ARS TK plasmid contained multiple copies of plasmid integrated into chromosomal DNA . In most cases, results with plasmids used in cotransformations were similar to those for plasmids carrying TK . However, cell lines produced by cotransformations with plasmids containing any one of three of the ARSs (m24, m25, or m26) often contained extrachromosomal DNAs.

Mutat Res, 1983 Nov, 111(3), 313 - 23
Mutagenic relevance of rat hepatocyte nuclei in the activation and inactivation of xenobiotica . Cyclophosphamide and epichlorohydrin activity on the yeasts S . pombe and S . cerevisiae; Migliore L et al.; The influence on the mutagenicity of cyclophosphamide (Cy) and epichlorohydrin (ECH), of liver nuclei and hepatic post-mitochondrial (S9) preparations from phenobarbital-treated rats, was examined . The study was conducted in vitro, with 2 yeasts, Schizosaccharomyces pombe (P1 strain) which allows the detection of forward mutations, and Saccharomyces cerevisiae (D5 strain), in which the induction of different genetic effects, such as mitotic recombination, can be evaluated . The results indicated that the nuclear fraction has a qualitative capacity for biotransformation of compounds, overlapping that of S9 . From a quantitative point of view, the Cy-activating capacity of the nuclear fraction was twice as high as that of S9 whereas the two fractions showed a similar ECH-inactivating ability . The present study strengthens the hypothesis of the relevance of nuclei as a site of metabolic activation and inactivation of exogenous compounds.

J Mol Biol, 1983 Aug 5, 168(2), 271 - 84
Cell division cycle genes nda2 and nda3 of the fission yeast Schizosaccharomyces pombe control microtubular organization and sensitivity to anti-mitotic benzimidazole compounds; Umesono K et al.; Two genes, nda2 and nda3, previously defined by cold sensitive nuclear division arrest (nda) mutations in the fission yeast Schizosaccharomyces pombe were studied . A mutant nda2-KM52 was found to be supersensitive (at the permissive temperature) to the tubulin-binding drugs such as thiabendazole, methylbenzimidazol-2yl carbamate and nocodazole . A single mutation in nda2 appears to cause both drug supersensitivity and cold sensitivity . The defective phenotypes of nda2-KM52 with a low concentration of the drugs were characterized by nuclear displacement and anomalously situated spindle pole bodies . The allele of the other mutant, nda3-KM311, was sh216 to be linked closely to the ben1 locus, which determines resistance to the drug . The identity of ben1 and nda3 genes was proved by a newly isolated mutant ben1-TB1005; it manifests ben1 resistance and the cold sensitive nda3 phenotype . At 22 degrees C, ben1-TB1005 showed cell branching and deformation characteristic of nda3-KM311 . Eleven mutants supersensitive to thiabendazole were newly isolated by replica plating . Four strains were mapped in nda2, while the other four were in nda3 . Most of the isolated mutants were blocked at nuclear division in the presence of a low concentration of the drug . Thus, the products of genes nda2 and nda3 (ben1) interact directly or indirectly with the drugs and control, in different ways, microtubular organization in the cells of S . pombe.

J Mol Biol, 1983 Aug 5, 168(2), 251 - 70
Cold-sensitive nuclear division arrest mutants of the fission yeast Schizosaccharomyces pombe; Toda T et al.; Thirteen recessive cold sensitive nuclear division arrest mutants were isolated from the fission yeast Schizosaccharomyces pombe . Twelve unlinked genes were defined; six in chromosome I, three in chromosome II and two in chromosome III . The map positions of three nuclear division arrest genes (nda1, nda2 and nda3) in chromosome II were determined precisely . Together with the previously obtained temperature-sensitive cell division cycle mutations, at least 20 genes appear to control the nuclear division of the fission yeast . Physiological studies indicated that most cold sensitive nda mutants incubated previously at 22 degrees C proceeded with a synchronously normal cell-cycle after temperature shift-up . The morphology of the nuclei and nuclear chromatin region was studied by the 4',6-diamidino-2-phenylindole staining method and by electron microscopy . Each mutant exhibited characteristic nuclear morphology at 22 degrees C, showing the specific blockages . The nda genes seem to control a pathway of structural alterations in the nuclear chromatin region with the order hemisphere, condensed ellipsoid, segregating U-form and separating hemispheres . Two genes, nda2 and nda3, pleiotropically control nuclear division, nuclear location and cell shape . The terminal phenotype of nda2-KM52 is characterized by the nuclear displacement, the absence of a spindle and abnormal locations of spindle pole bodies . The cells of nda3-KM311 were aberrant in shape and contained a partially separated chromatin region with a long spindle . Together with the results of the accompanying paper, we conclude that nda2 and nda3 genes control nuclear and cytoplasmic microtubular organization.

Mutat Res, 1983 Aug, 118(3), 213 - 26
Genotoxicity, metabolism and blood kinetics of epichlorohydrin in mice; Rossi AM et al.; Epichlorohydrin (ECH), a direct mutagen in vitro, did not induce chromosomal aberrations in bone-marrow cells of CD1 mice given single oral doses of 50 and 200 mg/kg in water . The ECH diol derivative (3-chloro-1,2-propanediol) was tested in vitro by a forward-mutation assay on the yeast Schizosaccharomyces pombe and showed a weak but significant mutagenic effect . The failure of ECH to induce mutagenic effects appears to be due to the rapid metabolic clearance of the compound in vivo . ECH blood kinetics at both doses, and at the same time the concentration of the diol, were determined . ECH rapidly disappeared from mouse blood, being no longer detectable 20 min after treatment . In contrast, 3-chloro-1,2-propanediol was measurable up to 5 h after dosage . No difference was observed in the kinetic and metabolic behavior of ECH after single and repeated doses (50 and 200 mg/kg/day for 7 days) . When 3-chloro-1,2-propanediol was tested, neither glutathione depletion nor epoxide hydrolase inhibition (evaluated with both styrene-7,8-oxide and ECH as substrates) could be detected in mouse liver . Finally, no difference in ECH blood kinetics or metabolism were observed in experiments in which the compound was administered (200 mg/kg) intraperitoneally in water or orally as a solution in dimethyl sulfoxide.

Eur J Biochem, 1983 Aug 1, 134(2), 315 - 9
Saccharomyces cerevisiae cdc9, a structural gene for yeast DNA ligase which complements Schizosaccharomyces pombe cdc17; Barker DG et al.; The Saccharomyces cerevisiae cdc9 gene has been cloned in the vector YRp12 by complementation of the temperature-sensitive lesion in vivo . The gene is contained within a 3300-base-pair fragment of DNA, which maps to the chromosomal locus of cdc9 and which is able to complement a DNA-ligase-deficient mutant of the fission yeast Schizosaccharomyces pombe.

Biochem J, 1983 Jun 15, 212(3), 749 - 54
Glycolysis and respiration in yeasts . The Pasteur effect studied by mass spectrometry; Lloyd D et al.; Simultaneous and continuous measurements of changes in CO2 and O2 concentrations in glucose-metabolizing yeast suspensions by mass spectrometry enabled a study of the Pasteur effect (aerobic inhibition of glycolysis) in Saccharomyces uvarum and Schizosaccharomyces pombe . A different control mechanism operates in Candida utilis to give a damped oscillation after the anaerobic-aerobic transition . The apparent Km values for respiration of the three yeasts were in the range 1.3-1.8 microM-O2 . The apparent Km values for O2 of the Pasteur effect were 5 and 13 microM for catabolite-repressed and derepressed S . uvarum respectively and 7 microM for Sch . pombe . These results are discussed with respect to currently accepted mechanisms for the control of glycolysis.

J Antibiot (Tokyo), 1983 Jun, 36(6), 639 - 45
Leptomycins A and B, new antifungal antibiotics . I . Taxonomy of the producing strain and their fermentation, purification and characterization; Hamamoto T et al.; A strain of Streptomyces was found to produce new antifungal antibiotics . The active compounds were purified and separated into two substances named leptomycin A and B by high performance liquid chromatography . The molecular formulae of leptomycins A and B are C32H46O6 and C33H48O6 respectively, and physicochemical and biological properties of them are very similar to each other . Leptomycins A and B exhibit strong inhibitory activity against Schizosaccharomyces and Mucor.

Mutat Res, 1983 Jun, 115(2), 215 - 23
Testing of chemicals for mutagenic activity with Schizosaccharomyces pombe; Loprieno N et al.; Two genetic end-points are used for testing mutagens in Schizosaccharomyces pombe: forward mutations of the loci which encode steps early in the adenine synthetic pathway and reversion of certain selected mutants . 54 chemicals have been tested for at least one of the genetic end-points . The relevant literature has been reviewed through 1979.

Mikrobiologiia, 1983 May-Jun, 52(3), 512 - 4
{Characteristics of the segregation of a new remote hybrid compared to an earlier remote hybrid obtained from crossing Saccharomyces cerevisiae with Schizosaccharomyces pombe}; Kosikov KV; The variability of two remote hybrids between Saccharomyces cerevisiae and Schizosaccharomyces pombe was compared in the course of their vegetative splitting . Hybrid 69 was obtained upon fusion of germinating spores, hybrid 92 was produced by copulation of vegetative cells . The variability of hybrid 92 was more diverse in terms of the size and shape of its cells; moreover, the hybrid yielded adenine-dependent haploid cells typical of S . cerevisiae with red colonies as well as dividing cells very similar to S . pombe with cross septa . Hybrid 69 did not produce such cultures.

Mutat Res, 1983 Apr, 117(1-2), 139 - 48
Mutagenicity of some organophosphorus compounds at the ade6 locus of Schizosaccharomyces pombe; Gilot-Delhalle J et al.; 12 organophosphorus insecticides were tested for toxicity and mutagenicity in the forward mutation test system ade6 of the yeast Schizosaccharomyces pombe . EMS and MMS were selected as positive controls . 3 compounds, dichlorvos, trichlorfon and paraoxon, showed a linear dose-response relationship . Among the other compounds investigated, methyl derivatives, though in general more toxic than ethyl derivatives, did not significantly increase the mutation frequency . Trichlorfon, tested in combination with malathion, methylparathion or methylazinphos (guthion), produced clearly synergistic effects for both toxicity and mutagenicity . The addition of S9 microsomal liver fraction decreased the efficiency of both single and combined treatments only where a dose-response relationship or a synergistic effect was obtained.

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