EMBO J, 1991 Jan, 10(1), 221 - 6 S . pombe pac1+, whose overexpression inhibits sexual development, encodes a ribonuclease III-like RNase; Iino Y et al.; The Schizosaccharomyces pombe pac1 gene is a multicopy suppressor of the pat1 temperature-sensitive mutation, which directs uncontrolled meiosis at the restrictive temperature . Overexpression of the pac1 gene had no apparent effect on vegetative growth but inhibited mating and sporulation in wild type S . pombe cells . In such cells, expression of certain genes required for mating or meiosis was inhibited . The pac1 gene is essential for vegetative cell growth . The deduced pac1 gene product has 363 amino acids . Its C-terminal 230 residues revealed 25% amino acid identity with ribonuclease III, an enzyme that digests double-stranded RNA and is involved in processing ribosomal RNA precursors and certain mRNAs in Escherichia coli . The pac1 gene product could degrade double-stranded RNA in vitro . These observations establish the presence of a RNase III homolog in eukaryotic cells . The pac1 gene product probably inhibits mating and meiosis by degrading a specific mRNA(s) required for sexual development . It is likely that mRNA processing is involved in the regulation of sexual development in fission yeast. EMBO J, 1991 Jan, 10(1), 123 - 32 Identification and cloning of TCF-1, a T lymphocyte-specific transcription factor containing a sequence-specific HMG box; van de Wetering M et al.; CD3-epsilon expression is controlled by a downstream T lymphocyte-specific enhancer element . We report the identification of a T cell-specific transcription factor, TCF-1, binding to this element . The multimerized recognition motif of TCF-1 constituted a T cell-specific enhancer . Subsequent cloning of TCF-1 identified three splice alternatives . TCF-1 contained a single DNA-binding HMG box most closely related to similar boxes in the putative mammalian sex-determining gene SRY and in the Schizosaccharomyces pombe Mc mating type gene . TCF-1 mRNA was expressed uniquely in T lymphocytes . Upon cotransfection into non-T cells, TCF-1 could transactivate through its cognate motif . These results identify TCF-1 as a T cell-specific transcription factor, which might play a role in the establishment of the mature T cell phenotype. J Cell Biol, 1991 Jan, 112(2), 191 - 201 The chromatin structure of centromeres from fission yeast: differentiation of the central core that correlates with function; Polizzi C et al.; We have examined the chromatin structure of centromere regions from the fission yeast Schizosaccharomyces pombe . The large and complex centromere regions of the S . pombe chromosomes encompass many kilobase pairs of DNA and contain several classes of tandemly repeated DNA sequences . The repeated sequences are further organized into a large inverted repeat flanking a central core, a conserved structural feature among all three centromeres in S . pombe . The nucleosomal configuration of the centromere regions is nonuniform and highly varied . Most of the centromere-specific repeated DNA sequences are packaged into nucleosomes typical of bulk chromatin . However, the central core and core-associated repeated sequences from the centromere regions of chromosomes I (cen1) and II (cen2), when present in S . pombe, show an altered chromatin structure, with little or no evidence of regular nucleosomal packaging . The atypical chromatin organization of the cen2 central core is not due to transcription, as no transcripts from this region were detected . These same DNA sequences, however, are packaged into nucleosomes typical of bulk chromatin when present in a nonfunctional environment on a minichromosome in the budding yeast Saccharomyces cerevisiae . Because the cen2 central core sequences themselves do not preclude regular nucleosomal packaging, we speculate that in S . pombe they constitute a specialized site of kinetochore protein assembly . The atypical nucleosomal pattern of the cen2 central core remains constant during the cell cycle, with only minor differences observed for some sequences . We propose that the unusual chromatin organization of the core region forms the basis of a higher order structural differentiation that distinguishes the centromere from the chromosome arms and specifies the essential structure for centromere function. Mol Cell Biol, 1991 Jan, 11(1), 289 - 98 The strong ADH1 promoter stimulates mitotic and meiotic recombination at the ADE6 gene of Schizosaccharomyces pombe; Grimm C et al.; The effect of the strong promoter from the alcohol dehydrogenase gene on mitotic and meiotic intragenic recombination has been studied at the ade6 locus of the fission yeast Schizosaccharomyces pombe . A 700-bp fragment containing the functional adh1 promoter was used to replace the weak wild-type promoter of the ade6 gene . Analysis of mRNA showed that strains with this ade6::adh1 fusion construct had strongly elevated ade6-specific mRNA levels during vegetative growth as well as in meiosis . These increased levels of mRNA correlated with a 20- to 25-fold stimulation of intragenic recombination in meiosis and a 7-fold increased prototroph formation during vegetative growth . Analysis of flanking marker configurations of prototrophic recombinants indicated that simple conversions as well as conversions associated with crossing over were stimulated in meiosis . The strongest stimulation of recombination was observed when the adh1 promoter was homozygous . Studies with heterologous promoter configurations revealed that the highly transcribed allele was the preferred acceptor of genetic information . The effect of the recombinational hot spot mutation ade6-M26 was also investigated in this system . Its effect was only partly additive to the elevated recombination rate generated by the ade6::adh1 fusion construct. Mol Cell Biol, 1991 Jan, 11(1), 281 - 8 Regulation of heat shock factor in Schizosaccharomyces pombe more closely resembles regulation in mammals than in Saccharomyces cerevisiae; Gallo GJ et al.; The heat shock response appears to be universal . All eucaryotes studied encode a protein, heat shock factor (HSF), that is believed to regulate transcription of heat shock genes . This protein binds to a regulatory sequence, the heat shock element, that is absolutely conserved among eucaryotes . We report here the identification of HSF in the fission yeast Schizosaccharomyces pombe . HSF binding was not observed in extracts from normally growing S . pombe (28 degrees C) but was detected in increasing amounts as the temperature of heat shock increased between 39 and 45 degrees C . This regulation is in contrast to that observed in Saccharomyces cerevisiae, in which HSF binding is detectable at both normal and heat shock temperatures . The S . pombe factor bound specifically to the heat shock element, as judged by methylation interference and DNase I protection analysis . The induction of S . pombe HSF was not inhibited by cycloheximide, suggesting that induction occurs posttranslationally, and the induced factor was shown to be phosphorylated . S . pombe HSF was purified to near homogeneity and was shown to have an apparent mobility of approximately 108 kDa . Since heat-induced DNA binding by HSF had previously been demonstrated only in metazoans, the conservation of heat-induced DNA binding by HSF among S . pombe and metazoans suggests that this mode of regulation is evolutionarily ancient. J Basic Microbiol, 1991, 31(2), 149 - 56 Use of alpha-aminoadipate and lysine as sole nitrogen source by Schizosaccharomyces pombe and selected pathogenic fungi; Ye ZH et al.; alpha-Aminodipate, an intermediate of the lysine biosynthetic pathway of fungi, or lysine when used as the sole nitrogen source in the medium was growth inhibitory and toxic to Saccharomyces cerevisiae . The fission yeast Schizosaccharomyces pombe and pathogenic fungi Candida albicans, Filobasidiella neoformans and Aspergillus fumigatus grew in the medium containing alpha-aminoadipate as the sole nitrogen source . C . albicans, A . fumigatus, and one of the strains of F . neoformans also grew in the medium containing lysine as the sole nitrogen source . When grown in the alpha-aminoadipate medium, only S . pombe accumulated a significant amount of alpha-ketoadipate in the culture supernatant . Also, 14C-alpha-aminoadipate was converted to 14C-alpha-ketoadipate in vivo . In the ammonium sulfate medium, S . pombe cells converted 14C-alpha-aminoadipate to lysine . The levels of glutamate-alpha-ketoadipate transaminase, an enzyme responsible for the conversion of alpha-aminoadipate to alpha-ketoadipate, and alpha-aminoadipate reductase, an enzyme required for the conversion of alpha-aminoadipate to lysine, were similar in S . pombe cells grown in the alpha-aminoadipate or ammonium sulfate medium . However, the level of homoisocitrate dehydrogenase, an enzyme before the alpha-ketoadipate step, was twelvefold lower in S . pombe cells grown in the alpha-aminoadipate medium compared to the level in cells grown in the ammonium sulfate medium . Pathogenic fungi used in this study did not accumulate alpha-ketoadipate and alpha-aminoadipate-delta-semialdehyde when grown in medium containing alpha-aminoadipate and lysine, respectively, as sole nitrogen source . However, only pathogenic fungi used both lysine and alpha-aminoadipate as sole nitrogen source . This unique metabolic property could be useful for the identification of these pathogens. Genes Dev, 1991 Jan, 5(1), 60 - 73 Fission yeast genes that confer resistance to staurosporine encode an AP-1-like transcription factor and a protein kinase related to the mammalian ERK1/MAP2 and budding yeast FUS3 and KSS1 kinases; Toda T et al.; Staurosporine, a potent inhibitor of protein kinase C, arrests fission yeast cell elongation specifically at a stage immediately after cell division . We isolated two genes, which, when carried on multicopy plasmids, confer drug resistance in fission yeast . One, spk1+, encodes a protein kinase highly similar (54% identity) to those encoded by the mammalian ERK1/MAP2 kinase and the budding yeast KSS1 and FUS3 genes . It is not essential for vegetative growth of Schizosaccharomyces pombe cells but is required for conjugation . The spk1+ gene product is a 45-kD protein enriched in the nucleus, and its level increases 10-fold after addition of staurosporine . The other gene pap1+ encodes an AP-1-like transcription factor that contains a region rich in basic amino acids followed by a "leucine zipper" motif . The pap1+ gene is required for spk1(+)-conferred staurosporine resistance . These two genes appear to function as a part of the fission yeast growth control pathway. Princess Takamatsu Symp, 1991, 22, 231 - 8 Mammalian G2 regulatory genes and their possible involvement in genetic instability in cancer cells; Okayama H et al.; In the fission yeast Schizosaccharomyces pombe, mitosis is initiated following the activation of the cdc2+/cyclin B kinase . The cdc2+/cyclin B kinase is positively regulated by cdc25+ tyrosine phosphatase and negatively regulated by wee1+/mik1+ tyrosine kinases . This regulatory system is evolutionarily conserved throughout higher eukaryotes . Drosophila and humans contain a cdc25+ gene homolog called String and CDC25Hs (hereafter referred to as CDC25Hul), respectively . We recently cloned a wee1+ homolog (WEE1Hu) and two additional cdc25+ homologs (CDC25Hu2 and CDC25Hu3) from human cells . Consequently, human cells contain at least one wee1+ and three cdc25+ homologs . Both CDC25Hu1 and CDC25Hu2 resemble the cdc25+ gene not only in structure and function but also in the mode of expression . They are expressed mostly in G2 . On the other hand, CDC25Hu3 is expressed mostly in early S, indicating that it has some novel function in the early phase of the cell cycle . In all the cell lines examined, CDC25Hu2 is expressed to a greater extent than CDC25Hu1 or CDC25Hu3 . The expression of CDC25Hu2 is particularly high in various cancer cells including those transformed by SV40 or human papilloma virus type 16 E6, E7, both of which are well known for their ability to induce genomic instability . In addition, there is a noticeable correlation between the extent of aneuploidy and the level of CDC25Hu2 expression in the cancer cells examined . In view of the fact that overexpression of cdc25+ under certain conditions induces genomic instability in the fission yeast, overexpression of CDC25Hu2 associated with many cancer cells might play at least a role in the induction of their chromosomal abnormalities. Folia Microbiol (Praha), 1991, 36(2), 153 - 7 Formation and reversion of Schizosaccharomyces pombe cells with apical protoplast protuberances; Zemanova Z et al.; Lytic enzymes from the hepatopancreas of Helix pomatia do not induce a uniform digestion of the cell wall of Schizosaccharomyces pombe over the entire cell surface . Perforations are formed in growth zones through which a protoplast can locally protrude . Conditions were found under which the frequency of formation of apical protoplast protuberances is higher than 90% cells with such protuberances can reverse to normally multiplying cells. Cold Spring Harb Symp Quant Biol, 1991, 56, 605 - 11 New elements in the mitotic control of the fission yeast Schizosaccharomyces pombe; Fantes PA et al.; The p107wee1 protein kinase plays a central role in regulating the cell cycle of fission yeast . It mediates transmission of signal(s) related to the nutritional status of the cell to the p34cdc2 protein kinase, which is an active component of the MPF complex driving cells into mitosis . p107wee1 is itself subject to control by the products of other genes such as nim1+/cdr1+, win1+, and perhaps wis1+ and other wis+ genes . At present, the relationships between these genes and their possible roles in the mitotic control are unclear and must await further analysis (Fig . 5) . It is likely that some of the gene products are concerned with the sensing and/or transmission of nutritional signals . p107wee1 negatively regulates the activity of p34cdc2, probably by direct tyrosine phosphorylation, and also appears to regulate the activities of the cdc1+ and cdc27+ gene products . The effects of nitrogen starvation and of wee1 mutations on conditional lethal mutations at the cdc1, cdc2, and cdc27 loci, taken together, support the largely speculative model shown in Figure 5 . During the normal cycle, the balance between phosphorylated and dephosphorylated p34cdc2 changes such that at the appropriate time, p34cdc2 is activated and the cell enters mitosis . We suggest that the cdc1+ and cdc27+ products may be regulated in a similar way . Such a mechanism would ensure coordinated activation of these and perhaps other proteins required for the G2/M transition . There are, of course, many uncertainties, and these must await elucidation by biochemical and genetic analysis. Cold Spring Harb Symp Quant Biol, 1991, 56, 599 - 604 stf1: a new suppressor of the mitotic control gene, cdc25, in Schizosaccharomyces pombe; Hudson JD et al.; A novel element in the mitotic control, stf1, has been identified genetically by its ability to rescue cdc25-22 as well as a gene disruption of cdc25 . This is the first phenotypically non-wee mutation shown to do so . stf1-1 functions additively with cdc2-1w, cdc2-3w, or wee1-6 to rescue cdc25 . The available data are consistent with the wild-type gene product operating either on the same pathway as cdc25 or to stimulate cdc2 by a pathway independent of cdc25 or wee1 . The stf1 gene has been cloned and sequenced and encodes a putative protein of 50-65 kD, depending on whether a potential intron is present . It is a novel protein with no homology detected in the current data bases . When challenged with hydroxyurea, stf1-1 acts additively with cdc2-3w in rescuing cdc25 mutants and in allowing mitosis to occur without DNA synthesis . It does not appear to play a role in the nutritional sensing pathway nor in the pathway mediating radiation-induced G2 delay. Oncogene, 1991 Jan, 6(1), 3 - 9 Identification and characterization of a human homolog of the Schizosaccharomyces pombe ras-like gene YPT-3; Drivas GT et al.; The Polymerase Chain Reaction was used to amplify ras and ras-like sequences from two human cDNA libraries . Members corresponding to each of the three major ras-subfamilies (ras, rho, and rab/YPT) were identified . The one homologous to rab/YPT, referred to here as YL8, appears to be the human homolog of the recently reported Schizosaccharomyces pombe YPT3 gene . The YL8 gene could encode a guanine nucleotide binding protein of 216 amino acids with about 70% amino acid sequence identity to S . pombe YPT3, and is transcriptionally active in a variety of human cell lines. Cell Motil Cytoskeleton, 1991, 18(2), 86 - 93 Microtubules in the fission yeast Schizosaccharomyces pombe contain only the tyrosinated form of alpha-tubulin; Alfa CE et al.; The state of tubulin tyrosination in the fission yeast Schizosaccharomyces pombe was investigated using a combination of indirect immunofluorescence microscopy and Western blotting . Antibodies specific for the tyrosinated form of alpha-tubulin stained all microtubule arrays in wild type cells and recognised the two alpha-tubulin polypeptides in Western blots of cell extracts enriched for tubulin by DEAE-Sephadex chromatography . Antisera that specifically recognised the detyrosinated, glu, form, on the other hand, gave consistently negative results, both in cells undergoing rapid exponential growth and in those allowed to accumulate in stationary phase . Neither the "ageing" of microtubules, by arresting cells at different points (late G1 or G2/M) in the cell division cycle, nor stabilising them, using D2O, lead to any detectable tubulin detryrosination . These results suggest that S . pombe lacks the carboxypeptidase that carries out the tubulin detyrosination reaction . This is the first report of an organism that possesses the correct C-terminal alpha-tubulin sequence yet fails to carry out this post-translational modification . The implication of this novel finding for the biological role of these events is discussed. Princess Takamatsu Symp, 1991, 22, 137 - 44 Protein phosphatases in cell division: how vital are they? Yanagida M, Yamano H, Stone EM, Kinoshita N, Yoshida T, Shiozaki K. In contrast to the wealth of information on cellular function of protein kinases, many of which are known to be the products of proto-oncogenes, little is known about how protein dephosphorylation is involved in growth control of normal and malignant cells . In the present study, roles of protein phosphatases in cell division cycle control were examined by molecular genetic approaches using a lower eukaryote, the fission yeast Schizosaccharomyces pombe . Nine protein phosphatase genes have been so far identified and characterized in this organism . Each of two (dis2+, sds21+, and ppa1+, ppa2+) gene products is highly similar to mammalian type 1 and 2A ser/thr phosphatases, respectively . The ppx1+ product is an intermediate of type 1 and 2A, while the ppb1+ product is similar to Ca(2+)-dependent type 2B . At least two protein tyrosine phosphatase genes (pyp1+ and pyp2+) exist . The cdc25 protein is now established to be a tyrosine phosphatase that activates cdc2 kinase . Some of these phosphatase genes are interrelated but have distinct, essential functions in cell cycle control . Missense mutations, deletions or high dosage expression of these phosphatase genes affect entry into and exit from mitosis, mitotic chromosome disjunction, cell size and cell shape . They seem to interact with the main regulators of mitosis, cdc2, cdc13/cyclin, cdc25 and weel, or with mitotic structural components, such as condensed chromosomes or the spindle apparatus . We show that the product of an essential gene, sds22+, is an important, positive factor in controlling the expression and modulating the activity of dis2 phosphatase. Cell Motil Cytoskeleton, 1991, 20(1), 47 - 54 Segregation of the nucleolus during mitosis in budding and fission yeast; Granot D et al.; The segregation of the nucleolus during mitosis was examined in Saccharomyces cerevisiae and Schizosaccharomyces pombe by indirect immunofluorescence using antibodies directed to highly conserved anti-nucleolus antigens . In mitotic S . pombe cells, the nucleolus appears to trail the bulk of the DNA . In wild-type cells of S . cerevisiae, the nucleolus segregates alongside the bulk of the genomic DNA . Based on its distance from the centromere, we would expect the rDNA in both organisms to segregate behind the majority of the genomic DNA, if telomeric regions trail centromeric regions as in other eukaryotes . We therefore suggest that in S . cerevisiae the nucleolus is attached to other parts of the nucleus which enable it to segregate along with the bulk of the DNA . The segregation of the nucleolus in topoisomerase mutants and nuclear division mutants of S . cerevisiae was also investigated . In cdc14 mutants which arrest at late anaphase, the vast majority of the DNA is separated, but the nucleolar antigens remain extended between the mother and daughter cells . Thus, the CDC14 gene of S . cerevisiae appears to be important for the separation of the nucleolus at mitosis. Verh K Acad Geneeskd Belg, 1991, 53(4), 365 - 85 {From ovocyte to biochemistry of the cell cycle}; Ozon R; The cell division cycle in eukaryotes contains up to three major transition points; the conversion of quiescent cells to a stage of active proliferation, the initiation of DNA synthesis (S phase) and the induction of mitosis in cells with newly replicated genome (M phase) . Within the past years two strategies, have converged to identify, genetically and biochemically a key protein kinase p34 cdc2 that governs the entry into mitosis . In the fission yeast Schizosaccharomyces pombe a number of mutants in the mitotic regulatory circuit have been isolated . A central gene in the network is cdc2 which is essential for the proper execution of mitosis . The cdc2 gene interacts with a number of other genes for correct mitotic control . The Amphibian oocyte, the oocyte from Xenopus laevis particularly, is arrested at the G2 phase of the first meiotic division; when it enters M phase, it contains a dominant regulatory factor known as MPF (M-phase or maturation promoting factor) . Purified MPF is an heterodimer formed of two polypeptides p34cdc2 an homologue of the product of the gene cdc2 and p45cdc13 or cyclin an homologue of the product of the gene cdc13 . Biochemical studies have revealed that p34cdc2 is a phosphotyrosine protein during the G2 phase of the cell cycle, both mitotic and meiotic . The tyrosine phosphorylation of p34cdc2 is regulated by the gradual accumulation of cyclin . At the onset of M phase, the complex p34cdc2/cyclin is activated as an histone H1 kinase, and p34cdc2 is tyrosine dephosphorylated . The mechanism of activation of p34cdc2 is negatively regulated by a form of protein phosphatase 2A . Ovulated vertebrate oocytes are arrested at metaphase of the second meiotic division (M II) under the control of the proto-oncogene c-mos a protein kinase . The exit of M II phase and the initiation of early embryonic mitotic cell cycles are physiologically induced by the spermatozoa at the time of fertilization . They requires the degradation of c-mos by a Ca2+ dependent proteolytic enzyme and the destruction of cyclin by an ubiquitin dependent pathway . The Xenopus oocyte has led to the molecular elucidation of MPF and identified links between cell cycle control, protein phosphorylation and proto-oncogenes . Despite the impresive progess of recent years, there is still much to be learned about the control of meiosis in Xenopus oocytes. Science, 1990 Dec 14, 250(4987), 1573 - 6 Complementation of the mitotic activator, p80cdc25, by a human protein-tyrosine phosphatase; Gould KL et al.; The onset of M phase requires the activation of the pp34 protein kinase in all eukaryotes thus far examined . In Schizosaccharomyces pombe, pp34 is phosphorylated on Tyr15, and dephosphorylation of this residue regulates the initiation of mitosis . In this study, it is shown that dephosphorylation of Tyr15 triggered activation of the pp34-cyclin complex from fission yeast, that a human protein-tyrosine phosphatase can catalyze this event both in vitro and in vivo, and that activation of fission yeast pp34 does not require threonine dephosphorylation . The complementary DNA that encoded the tyrosine phosphatase replaced the mitotic activator p80cdc25, closely associating the cdc25(+)-activating pathway with tyrosine dephosphorylation of pp34. Genes Dev, 1990 Dec, 4(12A), 2146 - 56 Evidence for base-pairing between mammalian U2 and U6 small nuclear ribonucleoprotein particles; Hausner TP et al.; Intramolecular and intermolecular snRNA cross-links were generated by irradiating HeLa nuclear extract with 365 nm light in the presence of the psoralen derivative AMT . After deproteinization, cross-linked RNAs were resolved by gel electrophoresis and identified as anomalously migrating species by Northern blotting . In addition to the U4/U6 snRNA cross-link, we detected an intermolecular U2/U6 cross-link, as well as several apparently intramolecular U1, U2, and U5 cross-links . Photoreversal of the U2/U6 cross-link with 254 nm irradiation released stoichiometric amounts of U2 and U6 snRNA . To localize the U2/U6 cross-link, the 3' end of U2 in the purified U2/U6 complex was labeled selectively using a novel oligonucleotide "splint" technique . The labeled U2/U6 complex was then subjected to rapid enzymatic RNA sequencing or to targeted digestion of the U2 and U6 components of the complex by RNase H and a panel of complementary oligonucleotides . The U2/U6 cross-link is located upstream of nucleotide 15 in U2 and downstream of nucleotide 85 in U6, suggesting that the phylogenetically conserved base-pairing between these regions (6 consecutive base pairs in human, Drosophila melanogaster, and Caenorhabditis elegans, 7 in Schizosaccharomyces pombe and Trypanasoma brucei, 8 in Pisum sativum, 11 in Saccharomyces cerevisiae) is significant. Mol Cell Biol, 1990 Dec, 10(12), 6791 - 8 Two related families of retrotransposons from Schizosaccharomyces pombe; Levin HL et al.; Two related families of transposons were isolated from schizosaccharomyces pombe, an organism which has been the object of extensive genetic studies which had previously produced no evidence for the existence of such elements . These two classes of repeated DNAs, dubbed Tf1 (transposon of fission yeast 1) and Tf2 have many properties of retrotransposons . Tf1 and Tf2 both possess long terminal repeats and predicted protein sequences that resemble the protease, reverse transcriptase, and integrase domains of retroviruses . The chromosomal locations and total numbers of Tf1 and Tf2 differ greatly in various isolates of S . pombe . The Tf elements are expressed in the form of 4.5-kb mRNAs . The complete sequence of Tf1 was determined and suggests that a novel mechanism for regulating its gene expression may be used. EMBO J, 1990 Dec, 9(12), 4007 - 16 Nuclear scaffold attachment stimulates, but is not essential for ARS activity in Saccharomyces cerevisiae: analysis of the Drosophila ftz SAR; Amati B et al.; Nuclei isolated from eukaryotic cells can be depleted of histones and most soluble nuclear proteins to isolate a structural framework called the nuclear scaffold . This structure maintains specific interactions with genomic DNA at sites known as scaffold attached regions (SARs), which are thought to be the bases of DNA loops . In both Saccharomyces cerevisiae and Schizosaccharomyces pombe, genomic ARS elements are recovered as SARs . In addition, SARs from Drosophila melanogaster bind to yeast nuclear scaffolds in vitro and a subclass of these promotes autonomous replication of plasmids in yeast . In the present report, we present fine mapping studies of the Drosophila ftz SAR, which has both SAR and ARS activities in yeast . The data establish a close relationship between the sequences involved in ARS activity and scaffold binding: ARS elements that can bind the nuclear scaffold in vitro promote more efficient plasmid replication in vivo, but scaffold association is not a strict prerequisite for ARS function . Efficient interaction with nuclear scaffolds from both yeast and Drosophila requires a minimal length of SAR DNA that contains reiteration of a narrow minor groove structure of the double helix. Biochem Cell Biol, 1990 Dec, 68(12), 1297 - 330 Protein kinase cascades in meiotic and mitotic cell cycle control; Pelech SL et al.; Eukaryotic cell cycle progression during meiosis and mitosis is extensively regulated by reversible protein phosphorylation . Many cell surface receptors for mitogens are ligand-stimulated protein-tyrosine kinases that control the activation of a network of cytoplasmic and nuclear protein-serine (threonine) kinases . Over 30 plasma membrane associated protein-tyrosine kinases are encoded by proto-oncogenes, i.e., genes that have the potential to facilitate cancer when disregulated . Proteins such as ribosomal protein S6, microtubule-associated protein-2, myelin basic protein, and casein have been used to detect intracellular protein-serine (threonine) kinases that are activated further downstream in growth factor signalling transduction cascades . Genetic analysis of yeast cell division control (cdc) mutants has revealed another 20 or so protein-serine (threonine) kinases . One of these, specified by the cdc-2 gene in Schizosaccharomyces pombe, has homologs that are stimulated during M phase in maturing sea star and frog oocytes and mammalian somatic cells . Furthermore, during meiotic maturation in these echinoderm and amphibian oocytes, this is followed by activation of many of the same protein-serine (threonine) kinases that are stimulated when quiescent mammalian somatic cells are prompted with mitogens to traverse from G0 to G1 phase . These findings imply that a similar protein kinase cascade may oversee progression at multiple points in the cell cycle. Curr Genet, 1990 Dec, 18(6), 501 - 9 A mutated swi4 gene causes duplications in the mating-type region of Schizosaccharomyces pombe; Fleck O et al.; Efficient mating-type (MT) switching in homothallic strains of Schizosaccharomyces pombe is significantly reduced if they have a mutation in any of the eleven known swi genes . The swi4 mutation causes heterothallic as well as homothallic segregants, both of which have duplications in the MT region . In contrast to homothallic strains, h+ swi4 strains yield only a few duplications . The duplications originate in the process of MT switching, presumably by mistakes in the resolution of DNA intermediates . They always consist of one cassette and one of the intervening sequences, L and K respectively . Strains with up to seven cassettes in the MT region were found . The possible modes of their origins are discussed. Curr Genet, 1990 Dec, 18(6), 511 - 6 Use of the Tn903 neomycin-resistance gene for promoter analysis in the fission yeast Schizosaccharomyces pombe; Lang-Hinrichs C et al.; The bacterial neo gene from transposon Tn903 (Tn601) was used for dominant transformation of the fission yeast Schizosaccharomyces pombe . It was found that high transformation efficiency was dependent on a high level of promoter activity, mediated by the strong promoter of the Schizosaccharomyces pombe alcohol dehydrogenase gene (adh1), as shown by comparing the efficiency of transformation to G418-resistance, the resistance levels of transformed cells, and the in vitro amino-glycoside phosphotransferase activity . On the other hand, the heterologous promoter of the Saccharomyces cerevisiae alcohol dehydrogenase I gene (adc1) is shown to be a weak promoter in Schizosaccharomyces pombe, though its activity is significantly enhanced in cells grown on glycerol as a carbon source . This system for selection and detection of promoter-active sequences may provide a useful basis for the analysis of promoter elements in fission yeast. Nucleic Acids Res, 1990 Nov 25, 18(22), 6485 - 9 High-frequency transformation method and library transducing vectors for cloning mammalian cDNAs by trans-complementation of Schizosaccharomyces pombe; Okazaki K et al.; We describe a highly efficient alkali cation method and library transducing vectors for cloning mammalian cDNAs by trans-complementation of fission yeast Schizosaccharomyces pombe mutants . cDNA libraries constructed with the pcD or pcD2 vector are transduced into yeast by cotransfection with a linearized vector, which allows an enhanced homologous recombination between the yeast vector and the library plasmid leading to the efficient formation of concatemers containing pcD molecules . The transformation frequencies obtained by the method are 10(6) colonies per 10(8) cells transfected with 2 micrograms of library and 1 microgram of vector, 50-60% of which contain pcD molecules . The high-efficiency alkali cation method circumvents many of the shortcomings of the spheroplast method generally used for Schiz . pombe transfection . The vectors are maximized for the efficiency of library transduction and minimized for the rearrangements of pcD molecules during propagation in yeast . This system allows rapid screening of multi-million cDNA clone libraries for rare cDNAs in a routine scale of experiments . Using this system, various mammalian cDNAs that are extremely difficult, time-consuming, or unclonable to clone by other methods have been cloned. Gene, 1990 Nov 15, 95(2), 179 - 86 Expression and characterization of infectious bursal disease virus polyprotein in yeast; Jagadish MN et al.; Various expression vectors containing a cDNA fragment encoding all but the first five amino acids (aa) of the large polyprotein (N-VP2-VP4-VP3-C) of infectious bursal disease virus were transformed into yeasts . In both Saccharomyces cerevisiae and Schizosaccharomyces pombe, co- or post-translational processing of the unfused large polyprotein occurred, generating a stable C-terminal product (VP3) or correct size, but without any detectable N-terminal product (VP2) . Furthermore, when the processing of the polyprotein was interrupted, because of an engineered in-frame site-specific insertion of 4 aa, even VP3 (as part of the unprocessed polyprotein) was undetected . VP2 was detected in S . cerevisiae only when fused to yeast pre-sequences at the N terminus, suggesting that in yeast, VP2 or the unprocessed polyprotein, in the absence of its native N terminus or proper protection of its N-terminal aa residues is susceptible to proteolytic degradation . The first 8 aa of a modified pre-sequence of the CUP1 gene product and the pre-pro sequence of MF alpha 1 gene product have been used for stable intra- and extra-cellular production of VP2, respectively. Science, 1990 Nov 9, 250(4982), 786 - 91 Cell cycle control of DNA replication by a homologue from human cells of the p34cdc2 protein kinase; D'Urso G et al.; The regulation of DNA replication during the eukaryotic cell cycle was studied in a system where cell free replication of simian virus 40 (SV40) DNA was used as a model for chromosome replication . A factor, RF-S, was partially purified from human S phase cells based on its ability to activate DNA replication in extracts from G1 cells . RF-S contained a human homologue of the Schizosaccharomyces pombe p34cdc2 kinase, and this kinase was necessary for RF-S activity . The limiting step in activation of the p34 kinase at the G1 to S transition may be its association with a cyclin since addition of cyclin A to a G1 extract was sufficient to start DNA replication . These observations suggest that the role of p34cdc2 in controlling the start of DNA synthesis has been conserved in evolution. J Biol Chem, 1990 Nov 5, 265(31), 19351 - 5 A guanosine diphosphatase enriched in Golgi vesicles of Saccharomyces cerevisiae . Purification and characterization; Yanagisawa K et al.; We have recently described a luminal guanosine diphosphatase activity in Golgi-like vesicles of Saccharomyces cerevisiae (Abeijon, C., Orlean, P., Robbins, P . W., and Hirschberg, C . B . (1989) Proc . Natl . Acad . Sci . U . S . A . 86, 6935-6939) . The presumed in vivo role of this enzyme is to convert GDP into GMP . GDP is a reaction product following outer-chain mannosylation of luminal proteins and a known inhibitor of mannosyltransferases . It is hypothesized that GMP then returns to the cytosol . We have purified this enzyme to apparent homogeneity . Following solubilization from a membrane pellet using a buffer containing Triton X-100, the enzyme was purified on a concanavalin A-Sepharose column followed by Mono Q fast protein liquid chromatography (FPLC) and Superose-12 FPLC columns . After treatment with endoglycosidase H, the deglycosylated active enzyme was applied to a second Mono Q FPLC column and a phenyl-Superose FPLC column . The final enzyme activity was enriched 6500-fold over that of the Triton X-100 extract . The apparant molecular mass of the deglycosylated enzyme is 47 kDa . The purified enzyme is highly specific for guanosine diphosphate, requires Ca2+ for maximal activity, and has a broad pH optimum between 7.4 and 8.2 . The apparent Km for GDP is 0.1 mM; the Vmax is 4.9 mmol/min/mg of protein . An enzyme activity with similar substrate specificity has also been detected in membranes of Schizosaccharomyces pombe. Mol Gen Genet, 1990 Nov, 224(2), 264 - 8 An electrophoretic karyotype of Aspergillus niger; Debets AJ et al.; An electrophoretic karyotype of Aspergillus niger was obtained using contour-clamped homogeneous electric field (CHEF) gel electrophoresis . Chromosome-sized DNA was separated into four bands . Seven of the eight linkage groups could be correlated with specific chromosomal bands . For this purpose DNA preparations from seven transformant strains of A . niger each carrying the heterologous amdS gene of Aspergillus nidulans on a different chromosome were analysed . Some of the assignments were confirmed with linkage group-specific A . niger probes . The estimated sizes of the A . niger chromosome range from 3.5 to 6.6 Mb, based on gel migration relative to the chromosomes of Schizosaccharomyces pombe strains, Saccharomyces cerevisiae and A . nidulans . The total genome size of A . niger significantly exceeds that of A . nidulans and is estimated to be about 35.5-38.5 Mb . Electrophoretic karyotyping was used to allocate non-mutant rRNA genes and to estimate the number of plasmids integrated in a high copy number transformant. Proc Natl Acad Sci U S A, 1990 Nov, 87(22), 8711 - 5 Leucine-rich repeats and carboxyl terminus are required for interaction of yeast adenylate cyclase with RAS proteins; Suzuki N et al.; A Saccharomyces cerevisiae gene encoding adenylate cyclase has been analyzed by deletion and insertion mutagenesis to localize regions required for activation by the Sa . cerevisiae RAS2 protein . The NH2-terminal 657 amino acids were found to be dispensable for the activation . However, almost all 2-amino acid insertions in the middle 600 residues comprising leucine-rich repeats and deletions in the COOH-terminal 66 residues completely abolished activation by the RAS2 protein, whereas insertion mutations in the other regions generally had no effect . Chimeric adenylate cyclases were constructed by swapping the upstream and downstream portions surrounding the catalytic domains between the Sa . cerevisiae and Schizosaccharomyces pombe adenylate cyclases and examined for activation by the RAS2 protein . We found that the fusion containing both the NH2-terminal 1600 residues and the COOH-terminal 66 residues of the Sa . cerevisiae cyclase rendered the catalytic domain of the Sc . pombe cyclase, which otherwise did not respond to RAS proteins, activatable by the RAS2 protein . Thus the leucine-rich repeats and the COOH terminus of the Sa . cerevisiae adenylate cyclase appear to be required for interaction with RAS proteins. Proc Natl Acad Sci U S A, 1990 Nov, 87(21), 8272 - 6 A putative protein kinase gene (kin1+) is important for growth polarity in Schizosaccharomyces pombe; Levin DE et al.; Mixed synthetic oligonucleotides encoding a sequence conserved among tyrosine-specific protein kinases were used to probe the genome of the fission yeast Schizosaccharomyces pombe . A single gene (kin1+) was isolated that encodes a putative protein kinase closely related to the KIN1- and KIN2-encoded serine/threonine-specific protein kinases of Saccharomyces cerevisiae . kin1+ is transcribed into a 3.5-kilobase mRNA that contains an uninterrupted open reading frame encoding a polypeptide of 98 kDa . In contrast to results obtained with kin mutants of S . cerevisiae, disruption of the Sc . pombe kin1+ gene resulted in recessive morphological and growth defects . kin1-disrupted cells grew slowly on enriched medium and grew as spheres, in contrast to wild-type Sc . pombe cells, which grow as rods . Relative to kin1+ cells, kin1-disrupted cells were differentially sensitive to lysis by treatment with alpha- and beta-glucanases, suggesting an alteration in either the composition or the organization of their cell walls. J Gen Microbiol, 1990 Nov, 136 ( Pt 11), 2251 - 9 Glycoprotein molecules in the walls of Schizosaccharomyces pombe wild-type cells and a morphologically altered mutant resistant to papulacandin B; de Mora JF et al.; Schizosaccharomyces pombe cell walls contain two major glycoprotein species, I and II, with molecular masses of 2 x 10(6) and 5 x 10(5) Da respectively, as determined by gel filtration chromatography and PAGE . The ratio of sugar to protein is higher in species I than in species II . Much of the sugar in both glycoproteins (about 85% in wild-type cells) is O-linked to the peptide moiety . The morphological sph1 mutant is resistant to papulacandin B, and its cell wall contains less glycoprotein II (but not less glycoprotein I) than the parental wild-type strain, although glycoprotein II is still synthesized and released into the growth medium . Papulacandin B largely reverses the morphological alteration of the mutant, and returns the ratio between species I and II to about that found in the parental strain, although the absolute amount of species II is still lower in the mutant . The results point to the importance of the relative amounts of the different wall polymers in determining cell morphology. EMBO J, 1990 Nov, 9(11), 3573 - 81 Drosophila cdc2 homologs: a functional homolog is coexpressed with a cognate variant; Lehner CF et al.; Using probes obtained by PCR amplification, we have cloned Drosophila cDNAs encoding structural homologs of the p34cdc2 cell cycle kinase . Southern blot experiments and in situ hybridization to polytene chromosomes demonstrated that the isolated cDNAs, were derived from two distinct genes, Dm cdc2 (31E) and Dm cdc2c (92F) . Northern blot and in situ hybridization experiments revealed that these two genes are coexpressed during embryogenesis and that expression is correlated with cell proliferation . However, despite the similarity in structure and expression, the two gene products differed in functional assays in yeasts . Expression of Dm cdc2 in Schizosaccharomyces pombe and Saccharomyces cerevisiae rescued cell cycle arrest caused by mutations in cdc2+ and CDC28, the genes encoding the p34cdc2 kinase homologs of these yeasts . In contrast, the Dm cdc2c gene product did not restore cell cycle progression . Thus, in addition to the identification of a functional homolog in Drosophila, our results indicate the presence of a closely related cognate of the p34cdc2 cell cycle kinase. EMBO J, 1990 Nov, 9(11), 3565 - 71 Complementation of fission yeast cdc2ts and cdc25ts mutants identifies two cell cycle genes from Drosophila: a cdc2 homologue and string; Jimenez J et al.; We have exploited the universality of the molecular mechanisms that control entry into mitosis to clone the Drosophila melanogaster homologues of fission yeast Schizosaccharomyces pombe cell division control (cdc) genes by the complementation of temperature sensitive mutations . The Drosophila genes were expressed in S.pombe as cDNAs from the SP6 promoter . Successful recovery of complementing plasmids required that we first 'adapt' pooled plasmids from a Drosophila embryonic cDNA library for propagation in fission yeast by introducing an ars1-LEU2 DNA fragment into the vector . This library was introduced into S.pombe cdc2 and cdc25 mutants, and plasmids isolated carrying cDNAs that complement these mutations . The gene that encodes the Drosophila cdc2 homologue maps to a single locus in the Drosophila genome at 31E on chromosome 2 . It is expressed maternally to provide mRNA in syncytial embryos, and appears to be zygotically expressed in mitotically active regions of the cellularized embryo . We have isolated two different cDNAs that complement cdc25-22 . One corresponds to a transcript of string, previously described as the Drosophila homologue of cdc25, and the other to a gene that has not been previously characterized. J Gen Microbiol, 1990 Nov, 136 ( Pt 11), 2261 - 5 The occurrence of glucosaminoglycan in the wall of Schizosaccharomyces pombe; Sietsma JH et al.; The major part of the wall of Schizosaccharomyces pombe consists of (1----3)-alpha-glucan and (1----3)-beta-glucan with some (1----6)-beta-linkages . Although in hydrolysed samples only a minute amount of glucosamine could be detected, this amino sugar may play an essential role as an integral part of a glucosaminoglycan/glucan complex . Treatment of the wall with either nitrous acid or chitinase changed the solubility properties of the beta-glucan, which suggests that the glucosaminoglycan/glucan complex is essentially similar to that found in walls of other fungi . An enzyme with properties similar to that of chitin synthase of other fungi, and probably responsible for the synthesis of the glucosaminoglycan, was detected in a mixed-membrane fraction. J Cell Sci, 1990 Nov, 97 ( Pt 3), 509 - 16 Mitochondrial growth and DNA synthesis occur in the absence of nuclear DNA replication in fission yeast; Sazer S et al.; Cell growth and division require the doubling of cellular constituents followed by their equal distribution to the two daughter cells . Within a growing population, the ratio of mitochondrial to cellular volume is maintained, as is the number of mitochondrial genomes per cell . The mechanisms responsible for coordinating nuclear and mitochondrial DNA synthesis, and for balancing increases in cell and mitochondrial size are not well understood . In studies of the fission yeast Schizosaccharomyces pombe we quantified cellular and mitochondrial DNA content by both Southern blot analysis and flow cytometry of cells stained with a variety of DNA-binding fluorochromes, which we show are able to detect nuclear and mitochondrial DNA with different efficiencies . In the conditional cell division cycle mutant cdc10, which is unable to initiate nuclear DNA synthesis, we found that there was an increase in the mitochondrial DNA content in the absence of nuclear DNA replication . This demonstrates that mitochondrial and nuclear DNA synthesis are not obligately linked . We also show that mitochondrial DNA replication is not required for the increase in mitochondrial size that occurs as cells elongate, although this results in a decrease in the ratio of mitochondrial DNA to mitochondrial volume. Gene, 1990 Oct 30, 95(1), 91 - 8 The ADE2 gene from Saccharomyces cerevisiae: sequence and new vectors; Stotz A et al.; We have determined the sequence of a DNA fragment encoding the ADE2 gene from Saccharomyces cerevisiae . A DNA fragment of 2241 bp capable of complementing ade2 mutations was modified so it is available as a single BglII fragment for use in yeast vectors or for gene disruptions . The minimal fragment codes for a putative protein which is highly similar to the protein encoded by the ADE6 gene from Schizosaccharomyces pombe and to the proteins encoded by the purEK operon of Escherichia coli. Gene, 1990 Oct 30, 95(1), 105 - 10 Isolation of a gene encoding a mitochondrial HSP70 protein from Schizosaccharomyces pombe; Powell MJ et al.; We have isolated cDNA and genomic clones encoding a mitochondrial HSP70 protein from Schizosaccharomyces pombe . Nucleotide sequence analysis indicates that the encoded protein is homologous to the HSP70s of other organisms . The highest degree of amino acid conservation is with the proteins encoded by the Escherichia coli dnaK gene, the SSC1 gene of Saccharomyces cerevisiae and the MTP70 gene of Trypanosoma cruzi, the latter two having recently been shown to be located in the mitochondria . Western-blot analysis with immunoglobulin G raised against a peptide corresponding to the C terminus of the SSP1 protein indicates a 70-kDa protein which is associated with the mitochondria. FEBS Lett, 1990 Oct 29, 273(1-2), 107 - 10 Heat shock induces enzymes of trehalose metabolism, trehalose accumulation, and thermotolerance in Schizosaccharomyces pombe, even in the presence of cycloheximide; De Virgilio C et al.; Exponentially growing cells of the fission yeast, Schizosaccharomyces pombe, contained virtually no trehalose at 27 degrees C but rapidly accumulated large quantities during heat shock at 40 degrees C . Activities of trehalose-6-phosphate synthase and trehalase also increased upon heat shock . Thermotolerance of the cells, measured as survival at 52 degrees C, increased in parallel to trehalose accumulation and decreased in parallel to the trehalose levels when cells were shifted back to 27 degrees C . Trehalose levels, activities of enzymes of trehalose metabolism and thermotolerance strongly increased upon heat shock even in the presence of cycloheximide, indicating that none of these effects requires protein synthesis . The data support the hypothesis that trehalose acts as a thermoprotectant in Schizosaccharomyces pombe. J Biol Chem, 1990 Oct 25, 265(30), 18400 - 7 Calcium homeostasis and transport are affected by disruption of cta3, a novel gene encoding Ca2(+)-ATPase in Schizosaccharomyces pombe; Ghislain M et al.; A new P-type ATPase gene, cta3, has been identified in Schizosaccharomyces pombe . The deduced amino acid sequence presents a 45% identity with the Saccharomyces cerevisiae putative Ca2(+)-ATPase encoded by the PMR2 gene . The cta3 protein contains 7 out of the 8 amino acid residues involved in high affinity Ca2+ binding in the sarcoplasmic reticulum Ca2(+)-ATPase from muscles . It also contains a region similar to the phospholamban-binding domain that characterizes this Ca2+ pump . A null mutation of cta3 leads to higher levels of cytosolic free Ca2+ and to lower amounts of sequestered and bound Ca2+ . Cellular Ca2+ efflux and rates of uptake into intracellular compartments are reduced by the loss of cta3 function . The sequence analysis and the physiological results strongly support the conclusion that the cta3 gene encodes a Ca2(+)-ATPase, probably located in intracellular membranes. Nature, 1990 Oct 18, 347(6294), 680 - 2 Distinct nuclear and spindle pole body population of cyclin-cdc2 in fission yeast; Alfa CE et al.; Cyclins, as subunits of the protein kinase encoded by the cdc2 gene are major controlling elements of the eukaryotic cell cycle . The fission yeast Schizosaccharomyces pombe has a B-type cyclin, which is a nuclear protein encoded by the cdc13 gene . Here we demonstrate the presence of two spatially distinct cdc13 cyclin populations in the nucleus of S . pombe, one of which is associated with the mitotic spindle poles . Both populations colocalize with the product of the cdc2 gene (p34cdc2) . Treatment of cells with the antimicrotubule drug thiabendazole prevents cyclin degradation and blocks the tyrosine dephosphorylation and activation of cdc2 . These results suggest a key regulatory role of the cdc2-cyclin complex in the initiation of mitotic spindle formation and also that mitotic microtubule function is required for cdc2 activation. J Biol Chem, 1990 Oct 5, 265(28), 16856 - 62 Protein transport into mitochondria is conserved between plant and yeast species; Chaumont F et al.; Protein targeting into plant mitochondria was investigated by in vitro translocation experiments . The precursor of the mitochondrial F1-ATPase beta subunit from Nicotiana plumbaginifolia was synthesized in vitro, translocated to, processed, and assembled in purified Vicia faba mitochondria . Transport (but not binding) required a membrane potential and external nucleotides and was conserved among plant species . beta subunit precursors from the yeasts Saccharomyces cerevisiae and Schizosaccharomyces pombe were imported and correctly processed in plant mitochondria . This translocation used protease-sensitive components of the outer membrane . Conversely, the N . plumbaginifolia beta subunit precursor was efficiently translocated and cleaved in yeast mitochondria . However, a precursor for a chloroplast protein was not targeted to plant or yeast mitochondria . We conclude that the machinery for protein import into mitochondria is specific and conserved in plant and yeast organisms . These results are discussed in the context of a poly- or monophyletic origin of mitochondria. Mol Cell Biol, 1990 Oct, 10(10), 5548 - 52 Evolutionary origin of the U6 small nuclear RNA intron; Reich C et al.; U6 is the most conserved of the five small nuclear RNAs known to participate in pre-mRNA splicing . In the fission yeast Schizosaccharomyces pombe, the single-copy gene encoding this RNA is itself interrupted by an intron (T . Tani and Y . Ohshima, Nature (London) 337:87-90, 1989) . Here we report analysis of the U6 genes from all four Schizosaccharomyces species, revealing that each is interrupted at an identical position by a homologous intron; in other groups, including ascomycete and basidiomycete fungi, as well as more distantly related organisms, the U6 gene is colinear with the RNA . The most parsimonious interpretation of our data is that the ancestral U6 gene did not contain an intron, but rather, it was acquired via a single relatively recent insertional event. J Appl Bacteriol, 1990 Oct, 69(4), 569 - 77 Growth inhibitory and biocidal activity of some isothiazolone biocides; Collier PJ et al.; Similar patterns of growth inhibition were observed for the three biocides, benzisothiazol-3-one (BIT), 5-chloro-N-methylisothiazol-3-one (CMIT) and N-methylisothiazol-3-one (MIT) against Escherichia coli ATCC 8739 and Schizosaccharomyces pombe NCYC 1354 . After periods of induced stasis, proportional to biocide concentration, growth proceeded at an inhibited rate . Extrapolation of the static periods and inhibited growth rates against biocide concentration gave minimum growth inhibitory concentration estimates of 0.1-0.5 micrograms/ml for CMIT, 15-20 micrograms/ml for BIT and 40-250 micrograms/ml for MIT . Patterns of growth inhibition by CMIT and induced morphological changes in inhibited cultures suggested this compound to also inhibit initiation of DNA replication . Growth inhibitory activity was rapidly quenched by the addition of thiol-containing materials such as glutathione and cysteine . The activity of CMIT was additionally quenched by the presence of the non-thiol amino acids valine and/or histidine . These results suggest that the chlorinated isothiazolones can react with amines as well as with essential thiol groups. Curr Genet, 1990 Oct, 18(3), 269 - 72 The structural gene coding for thiamin-repressible acid phosphatase in Schizosaccharomyces pombe; Yang JW et al.; The pho4 gene of the fission yeast Schizosaccharomyces pombe is regulated by thiamin . The nucleotide sequence of this gene is given here and it is shown that it matches the amino acid sequence of thiamin-repressible acid phosphatase, corroborating genetic evidence that pho4 represents the structural gene of this enzyme . The gene codes for a protein of 463 amino acids in length and shows regions of strong similarity with the phosphate-repressible acid phosphatase of Schizosaccharomyces pombe . The enzyme has a cleavable signal sequence 18 amino acids long and carries nine potential N-glycosylation sites. Curr Genet, 1990 Oct, 18(3), 193 - 7 The recombinational hot spot mutation ade6-M26 of Schizosaccharomyces pombe stimulates recombination at sites in a nearby interval; Grimm C et al.; With the help of in vitro constructed intragenic double mutants, we investigated the influence of the recombinational hot spot mutation ade6-M26 on meiotic recombination between two additional ade6 mutations proximal to it . Recombination was stimulated four-fold when M26 was present in a heterozygous condition and ten-fold when homozygous . M26 itself remained unaffected in a substantial number of these events . This indicates that the stimulation can not only be due to a preferred conversion of M26 to wild-type with co-conversion of the second mutation in cis . A model is proposed in which M26 acts as an "entry site" for recombinational enzymes. Genetics, 1990 Oct, 126(2), 309 - 15 stf1: non-wee mutations epistatic to cdc25 in the fission yeast Schizosaccharomyces pombe; Hudson JD et al.; In Schizosaccharomyces pombe, cdc25 is a cell cycle regulated inducer of mitosis . wee1 and phenotypically wee alleles of cdc2 are epistatic to cdc25 . Mutant alleles of a new locus, stf1 (suppressor of twenty-five), identified in a reversion analysis of conditionally lethal cdr1-76 cdc25-22 and cdr2-96 cdc25-22 double mutant strains, also suppress both temperature-sensitive and gene disruption alleles of cdc25 . These mutants, by themselves, are phenotypically indistinguishable from wild type strains; hence they represent the first known mutations that are epistatic to cdc25 and do not display a wee phenotype . stf1 genetically interacts with other elements of mitotic control in S . pombe . stf1-1 is additive with wee1-50, cdc2-1w and cdc2-3w for suppression of cdc25-22 . Also, like wee1- and cdc2-w, stf1- suppression of cdc25 is reversed by overexpression of the putative type 1 protein phosphatase bws1+/dis2+ . Interaction with various mutants and plasmid overexpression experiments suggest that stf1 does not operate either upstream or downstream of wee1 . Similarly, it does not operate through cdc25 since it rescues the disruption . stf1 appears to encode an important new element of mitotic control. FEBS Lett, 1990 Oct 1, 271(1-2), 189 - 93 The RNA component of RNase P in Schizosaccharomyces species; Zimmerly S et al.; In the fission yeast Schizosaccharomyces pombe, the enzyme RNAse P copurifies with two RNAs, K1- and K2-RNA, which are identical except for their termini {1} and which are encoded by a single gene {2} . We have undertaken the cloning of the K-RNA genes in related organisms in order to gain comparative structural information . Because the K-RNA sequence is poorly conserved across species, we have cloned the K-RNA genes in the Schizosaccharomyces species S . malidevorans, S . japonicus, S . versatilis, and S . octosporus . Of the 4 species, only S . octosporus contains a K-RNA gene different from that in S . pombe; the gene diverges by 20% . Based on the two sequences, nuclease protection data and computer analysis, we have proposed a secondary structure model for the K-RNA . Northern analysis shows the K-RNA genes in all four Schizosaccharomyces species to be expressed as two RNAs, as in S . pombe. EMBO J, 1990 Oct, 9(10), 3045 - 50 Primary structure, genomic organization and heterologous expression of a glucose transporter from Arabidopsis thaliana; Sauer N et al.; Both genomic and full length cDNA clones of an Arabidopsis thaliana sugar carrier, STP1, have been obtained using a cDNA clone of the H+/hexose cotransporter from the green alga Chlorella kessleri as hybridization probe . The peptide predicted from these sequences in 522 amino acids long and has a molecular weight of 57,518 kd . This higher plant sugar carrier contains 12 putative transmembrane segments and is highly homologous to the H+/hexose cotransporter from Chlorella, with an overall identity in the amino acid sequence of 47.1% . It is also homologous to the human HepG2 glucose transporter (28.4%), and other sugar carriers from man, rat, yeast and Escherichia coli . The definite proof for the function of the STP1 protein as a hexose transporter and data on its substrate specificity were obtained by heterologous expression in the fission yeast Schizosaccharomyces pombe . Transformed yeast cells transport D-glucose with a 100-fold lower KM value than control cells . Moreover only the transformed cells were able to accumulate the non-metabolizable D-glucose analogue 3-O-methyl-D-glucose, indicating that the Arabidopsis carrier catalyses an energy dependent, active uptake of hexoses . Expression of STP1 mRNA is low in heterotrophic tissues like roots or flowers . High levels of expression are found in leaves. Proc Natl Acad Sci U S A, 1990 Oct, 87(20), 7814 - 8 Adenylyl cyclase is dispensable for vegetative cell growth in the fission yeast Schizosaccharomyces pombe; Maeda T et al.; Disruption of the cyr1 gene of Schizosaccharomyces pombe, which encodes adenylyl cyclase, did not confer lethality to fission yeast cells, although they grew 40% slower than wild-type strains in complete medium . These cells contained no measurable amount of cAMP and no adenylyl cyclase activity . When h+ and h- cyr1 disruptants were mixed, they underwent mating even in rich medium . Propagation of homothallic cyr1 disruptants was difficult, probably because such cells readily mate and produce asci and thus stop growing . A greater than 10-fold increase in the amount of cyr1 mRNA was observed when cloned cyr1+ was introduced into Sch . pombe cells on a multicopy plasmid . The total adenylyl cyclase activity was similarly high in these transformants . However, the level of intracellular cAMP was hardly affected . Evidence suggests that this was not due to increased phosphodiesterase activity . Thus, cAMP level in growing fission yeast cells appears to be regulated not by the amount of adenylyl cyclase protein but by a feedback mechanism at the enzyme level . The cAMP level fell by approximately 50% under nitrogen starvation, which triggers sexual development in Sch . pombe . We suggest that fission yeast controls the level of intracellular cAMP primarily to regulate sexual development rather than to drive or arrest the cell cycle. Mol Cell Biol, 1990 Oct, 10(10), 5442 - 54 Drosophila scaffold-attached regions bind nuclear scaffolds and can function as ARS elements in both budding and fission yeasts; Amati B et al.; Histone-depleted nuclei maintain sequence-specific interactions with genomic DNA at sites known as scaffold attachment regions (SARs) or matrix attachment regions . We have previously shown that in Saccharomyces cerevisiae, autonomously replicating sequence elements bind the nuclear scaffold . Here, we extend these observations to the fission yeast Schizosaccharomyces pombe . In addition, we show that four SARs previously mapped in the genomic DNA of Drosophila melanogaster bind in vitro to nuclear scaffolds from both yeast species . In view of these results, we have assayed the ability of the Drosophila SARs to promote autonomous replication of plasmids in the two yeast species . Two of the Drosophila SARs have autonomously replicating sequence activity in budding yeast, and three function in fission yeast, while four flanking non-SAR sequences are totally inactive in both. Bioessays, 1990 Oct, 12(10), 457 - 63 Yeast as a model system for understanding the control of DNA replication in Eukaryotes; Bartlett R et al.; In the yeasts Saccharomyces cerevisiae and Schizosaccharomyces pombe, the initiation of DNA replication is controlled at a point called START . At this point, the cellular environment is assessed; only if conditions are appropriate do cells traverse START, thus becoming committed to initiate DNA replication and complete the remainder of the cell cycle . The cdc2+/CDC28+ gene, encoding the protein kinase p34, is a key element in this complex control . The identification of structural and functional homologues of p34 suggests that it has a role in the control of DNA replication in all eukaryotes . The WHI1+, CLN1+ and CLN2+ gene products, identified in S . cerevisiae, are positive regulators that function at START and may interact with p34 . Determining how passing the START control point leads to the initiation of DNA replication is a major outstanding challenge in cell cycle studies. J Biol Chem, 1990 Sep 15, 265(26), 15503 - 5 Effects of phosphate and hydrophobic molecules on two mutations in the beta-strand sector of the H(+)-ATPase from the yeast plasma membrane; Goffeau A et al.; At concentrations from 10 to 100 mM, inorganic phosphate and sulfate stimulate the activity of the H(+)-ATPase purified from the wild type Schizosaccharomyces pombe plasma membranes . Compared to the wild type ATPase, the stimulation by phosphate is more pronounced in the mutant pma1-1 (Gly-268----Asp) and is much reduced in the mutant pma1-2 (Lys-250----Thr) enzymes . In contrast, the inhibition by trifluoperazine is less pronounced in the pma1-1 mutant than in the wild type or pma1-2 mutant . The mutant pma1-2 ATPase activity is markedly stimulated by 10-20% dimethyl sulfoxide, which has a limited effect on the wild type and pma1-1 enzymes . These data indicate that the protein domain located in the beta-strand sector, including Lys-250 and Gly-268, is located in the active site and that its hydrophobic character influences the interactions of the yeast H(+)-ATPase with inorganic phosphate, as well as with the hydrophobic inhibitor trifluoperazine or the hydrophobic solvent dimethyl sulfoxide. Gene, 1990 Sep 14, 93(2), 265 - 70 Synthesis, phosphorylation, and nuclear localization of human papillomavirus E7 protein in Schizosaccharomyces pombe; Tommasino M et al.; The complete E7 protein-encoding open reading frame of human papillomavirus type 16 (HPV-16) was expressed in the fission yeast Schizosaccharomyces pombe, under the control of a cloned yeast promoter . The HPV-16 E7 protein synthesized in S . pombe is a 17-kDa phosphoprotein which is recognized by anti-E7 antibodies (raised in rabbits against E7 fusion protein produced in Escherichia coli) . The mobility during sodium dodecyl sulfate-polyacrylamide-gel electrophoresis of native E7 phosphoprotein synthesized in S . pombe is identical to that of the E7 phosphoprotein immunoprecipitated from human CaSki cells . Immunofluorescence staining showed that HPV-16 E7 phosphoprotein is localized in the nuclei of transformed S . pombe . These results indicate that E7 protein synthesized by S . pombe is apparently indistinguishable from HPV-16 E7 protein synthesized in higher eukaryotic cells expressing genes of HPV-16, and also that the phosphorylated, nuclear HPV-16 E7 protein is synthesized in S . pombe in a form compatible with its biological activity. Nucleic Acids Res, 1990 Sep 11, 18(17), 5207 - 12 Immunoprecipitation distinguishes non-overlapping groups of snRNPs in Schizosaccharomyces pombe; Tollervey D et al.; The large number of snRNAs in the fission yeast Schizosaccharomyces pombe can be divided into four non-overlapping groups by immunoprecipitation with antibodies directed against mammalian snRNP proteins . 1) Of the abundant snRNAs, anti-Sm sera precipitate only the spliceosomal snRNAs U1, U2, U4, U5 and U6 . Surprisingly, three Sm-sera tested distinguish between U2, U4 and U5 and U1 from S.pombe; one precipitating only U1 and two precipitating U2, U4 and U5 but not U1 . 2) A group of 11 moderately abundant snRNAs are not detectably precipitated by human anti-Sm sera, but are specifically precipitated by monoclonal antibody H57 specific for the human B/B' polypeptides . From Aspergillus nidulans this antibody also precipitates at least 12 snRNAs . 3) Anti-(U3)RNP sera do not precipitate the above snRNAs, but precipitate at least 6 further snRNAs, including the homologues of U3 . Both the anti-(U3)RNP sera and H57 also efficiently precipitate a number of discrete non-capped RNAs . 4) A small number of additional snRNAs are not detectably precipitated by any anti-serum tested to date, further analysis may identify antisera specific for these snRNPs . Western blots of purified snRNP proteins were used to identify the S.pombe proteins responsible for these immunoprecipitations . Several Sm-sera decorate a 16.3kD protein which may be a D protein homologue, monoclonal H57 decorates a further protein of 16kD and an anti-(U3)RNP serum decorates the homologue of the 36kD U3-specific protein, fibrillarin. Mol Gen Genet, 1990 Sep, 223(3), 407 - 16 DNA sequence elements required for transcription initiation of the Schizosaccharomyces pombe ADH gene in Saccharomyces cerevisiae; Furter-Graves EM et al.; The roles of the TATA element and sequences near the mRNA initiation site in specifying the location of initiation sites in Saccharomyces cerevisiae were examined, using the Schizosaccharomyces pombe ADH gene . The importance of spacing was demonstrated by analysis of a series of deletions that removed from 8-50 bp between the TATA element and ATG translation initiation site of this gene . Primer extension mapping showed that increasing deletion length is associated with a progressive shift downstream in the location of the initiation sites . The distance of a given site from the promoter affected the relative ability of the site to be utilized for initiation . For this gene, a permissive region for transcription initiation exists between 55 and 125 bases downstream of the TATA element, and a zone of 75-115 bases allows maximal usage of an initiation site . The presence of a TATA sequence was shown to be necessary in S . cerevisiae for maintaining the location of this "window" of initiation . The TATA sequence is essential for function of the gene in S . pombe . This gene, as well as the majority of the 63 S . cerevisiae genes surveyed, uses initiation sites which fit a PyAA/T(Pu) consensus . Cis-acting mutations were recovered which restored ADH activity to a deletion allele that initiates its mRNAs downstream of the ATG . DNA sequence and transcript analysis with these mutants confirmed the requirement of proper spacing and conformity of initiation sites to the PyAA/T(Pu) consensus for efficient transcript initiation. Mutat Res, 1990 Sep-Nov, 236(2-3), 277 - 87 Eukaryotic DNA ligases; Lasko DD et al.; Recent studies on eukaryotic DNA ligases are briefly reviewed . The two distinguishable enzymes from mammalian cells, DNA ligase I and DNA ligase II, have been purified to homogeneity and characterized biochemically . Two distinct DNA ligases have also been identified in Drosophila melanogaster embryos . The genes encoding DNA ligases from Schizosaccharomyces pombe, Saccharomyces cerevisiae and vaccinia virus have been cloned and sequenced . These 3 proteins exhibit about 30% amino acid sequence identity; the 2 yeast enzymes share 53% amino acid sequence identity or conserved changes . Altered DNA ligase I activity has been found in cell lines from patients with Bloom's syndrome, although a causal link between the enzyme deficiency and the disease has not yet been proven. Proc Natl Acad Sci U S A, 1990 Sep, 87(17), 6679 - 83 Human DNA ligase I cDNA: cloning and functional expression in Saccharomyces cerevisiae; Barnes DE et al.; Human cDNA clones encoding the major DNA ligase activity in proliferating cells, DNA ligase I, were isolated by two independent methods . In one approach, a human cDNA library was screened by hybridization with oligonucleotides deduced from partial amino acid sequence of purified bovine DNA ligase I . In an alternative approach, a human cDNA library was screened for functional expression of a polypeptide able to complement a cdc9 temperature-sensitive DNA ligase mutant of Saccharomyces cerevisiae . The sequence of an apparently full-length cDNA encodes a 102-kDa protein, indistinguishable in size from authentic human DNA ligase I . The deduced amino acid sequence of the human DNA ligase I cDNA is 40% homologous to the smaller DNA ligases of S . cerevisiae and Schizosaccharomyces pombe, homology being confined to the carboxyl-terminal regions of the respective proteins . Hybridization between the cloned sequences and mRNA and genomic DNA indicates that the human enzyme is transcribed from a single-copy gene on chromosome 19. Cell Regul, 1990 Sep, 1(10), 763 - 9 Biological activity of the mammalian RAP genes in yeast; Xu HP et al.; We have screened expression libraries for mammalian cDNAs capable of suppressing defects in ras1- Schizosaccharomyces pombe . Both the RAP1A and RAP1B genes were identified in this manner . They suppress defects in cell morphology and sporulation, although not conjugation . In contrast, RAP genes do not suppress phenotypes in the yeast Saccharomyces cerevisiae that are deficient in RAS . Indeed, expression of RAP1A appears to antagonize the activated S . cerevisiae RAS2val19 gene . These results indicate that RAP proteins can interact with RAS targets, sometimes productively, sometimes nonproductively. Genes Dev, 1990 Aug, 4(8), 1332 - 44 Human cDNAs encoding homologs of the small p34Cdc28/Cdc2-associated protein of Saccharomyces cerevisiae and Schizosaccharomyces pombe; Richardson HE et al.; The Cks1 protein is a component of the Cdc28 protein kinase in the budding yeast Saccharomyces cerevisiae . This paper reports the cloning of two homologs of the S . cerevisiae CKS1 gene from human cells . These homologs, CKShs1 and CKShs2, both encode proteins of 79 amino acids that share considerable homology at the amino acid level with the products of CKS1 from S . cerevisiae and suc1+ from the fission yeast Schizosaccharomyces pombe . Both human homologs are capable of rescuing a null mutation of the S . cerevisiae CKS1 gene when expressed from the S . cerevisiae GAL1 promoter . S . pombe suc1+ expressed from the GAL1 promoter is also capable of rescuing a S . cerevisiae cks1 null mutation . Ckshs1 or Ckshs2 protein linked to Sepharose beads can bind the Cdc28/Cdc2 protein kinase from both S . cerevisiae and human cells . The CKShs1 and CKShs2 mRNAs are expressed in different patterns through the cell cycle in HeLa cells, which may reflect specialized roles for the encoded proteins. Proc Natl Acad Sci U S A, 1990 Aug, 87(15), 5697 - 701 Mitotic role for the Cdc28 protein kinase of Saccharomyces cerevisiae; Reed SI et al.; The Cdc28 protein kinase functions in the G1 to S phase transition of the cell cycle of the budding yeast Saccharomyces cerevisiae . This is in contrast with observations of the homologous protein kinase from a variety of metazoans, where activity and function are associated with the G2 to M phase transition . We present evidence that the Cdc28 protein kinase is also required for mitosis and that this function is executed in the G2 interval of the cell cycle . We show, in addition, that the protein kinase is highly active during this phase of the cell cycle . The dual role of the Cdc28 protein kinase in the S . cerevisiae cell cycle thus parallels that demonstrated for the cdc2 protein kinase of the fission yeast Schizosaccharomyces pombe. J Cell Sci, 1990 Aug, 96 ( Pt 4), 683 - 9 Cell cycle regulation of p34cdc2 kinase activity in Physarum polycephalum; Ducommun B et al.; The regulation of the mitotic histone H1 kinase activity has been analyzed during the naturally synchronous cell cycle of Physarum polycephalum plasmodia . The universal binding property of the p13suc1 Schizosaccharomyces pombe gene product was used to precipitate and assay the cdc2 histone H1 kinase activity . The kinase activity peaks at the beginning of metaphase and its decline, which requires protein synthesis, appears to be an early event during the metaphase process . Microtubular poisons, temperature shifts and DNA synthesis inhibitors were used to perturb cell cycle regulatory pathways and characterize their effects on cdc2 kinase activation . Our results suggest that the full activation of the mitotic kinase requires at least two successive triggering signals involving microtubular components and DNA synthesis. Mol Gen Mikrobiol Virusol, 1990 Aug, (8), 5 - 8 {Phenotypic manifestations of reverse transcriptase activity in yeast cells}; Reznik NL et al.; The yeast Saccharomyces cerevisiae and Schizosaccharomyces pombe transformed by plasmids containing retrotransposon from yeast or Drosophila under the control of a strong promoter show the remarkable reverse transcriptase activity . The activity results in the impaired yeast growth and decreased mitotic stability of the plasmids . The phenotypic expression of the reverse transcriptase activity is observed within 30 days. FEBS Lett, 1990 Jul 30, 268(1), 217 - 21 Human chorionic gonadotropin alpha and human cytomegalovirus promoters are extremely active in the fission yeast Schizosaccharomyces pombe; Toyama R et al.; We have investigated the transcriptional activity of human cytomegalovirus, herpes thymidine kinase, human chorionic gonadotropin alpha, somatostatin, immunoglobulin kappa chain, alpha crystallin, albumin and interferon-beta promoters in the fission yeast Schizosaccharomyces pombe . Among these, the human cytomegalovirus, human chorionic gonadotropin alpha, and somatostatin promoters were found to be very active, approximately 11-, 9-, and 0.9-fold as active as the SV40 early promoter, respectively . The remainder of the promoters studied were weak, having only 10-20% of the SV40 promoter activity . Primer extension analysis showed that the strong promoters initiated transcription in S . pombe at the same sites as in mammalian cells, indicating the high similarity between both transcriptional systems. Nature, 1990 Jul 19, 346(6281), 291 - 4 Striking conservation of TFIID in Schizosaccharomyces pombe and Saccharomyces cerevisiae; Fikes JD et al.; Eukaryotic promoters contain binding sites for basic transcription factors and gene-specific activator proteins . The transcription factors interact at the TATA box, which lies close to the position of transcription initiation . Activators typically bind to distant sites that can lie kilobases away from the initiation site . The factor TFIID binds specifically to the TATA box to initiate an ordered pathway of assembly of the basic transcription factors . Biochemical analyses have shown that human and Saccharomyces cerevisiae TFIID are functionally interchangeable in vitro . To study further the functional conservation of this critical factor, we are surveying proteins from divergent organisms that can substitute in vivo for the S . cerevisiae TFIID . We report here the isolation of a unique gene from Schizosaccharomyces pombe that fully complements a null mutation in SPT15, the gene that encodes TFIID in S . cerevisiae . The Schiz . pombe gene encodes a protein 93% identical (166/178) to S . cerevisiae TFIID in a region consisting of a direct repeat. Nature, 1990 Jul 19, 346(6281), 240 - 4 A gene from the human sex-determining region encodes a protein with homology to a conserved DNA-binding motif; Sinclair AH et al.; A search of a 35-kilobase region of the human Y chromosome necessary for male sex determination has resulted in the identification of a new gene . This gene is conserved and Y-specific among a wide range of mammals, and encodes a testis-specific transcript . It shares homology with the mating-type protein, Mc, from the fission yeast Schizosaccharomyces pombe and a conserved DNA-binding motif present in the nuclear high-mobility-group proteins HMG1 and HMG2 . This gene has been termed SRY (for sex-determining region Y) and proposed to be a candidate for the elusive testis-determining gene, TDF. Mol Cell Biol, 1990 Jul, 10(7), 3750 - 60 Cloning and analysis of a gene involved in DNA repair and recombination, the rad1 gene of Schizosaccharomyces pombe; Sunnerhagen P et al.; We have cloned the rad1 gene of Schizosaccharomyces pombe by complementation of the rad1-1 mutant, which is deficient in DNA repair and recombination . The coding region of the gene is 582 base pairs long and contains no introns . The predicted product is a strongly acidic, 22-kilodalton protein containing 194 amino acid residues . This gene does not exhibit significant homology to any other known repair gene . The major transcription start site is at 27 base pairs upstream of the putative start codon . Insertion mutagenesis revealed that besides the coding region, at least 151 base pairs of 5'-flanking sequence are required for full complementing activity . A strain carrying a null allele of rad1 was constructed and found to have a phenotype closely similar to that of the rad1-1 mutant . Expression in Escherichia coli of the coding region yielded a protein product of a size close to that predicted from the DNA sequence . This product reacted with antibodies raised against a synthetic peptide with a sequence from that predicted for the protein product . We have localized the rad1 gene to NotI fragment E of the S . pombe genome. Mol Gen Genet, 1990 Jul, 222(2-3), 169 - 75 Molecular basis for determining the sensitivity of eucaryotes to the antimitotic drug rhizoxin; Takahashi M et al.; Rhizoxin, an antibiotic, exhibits potent anti-mitotic activity against most eucaryotic cells including those of higher vertebrates, plants and fungi by binding to beta-tubulin . The benA gene of three independently isolated rhizoxin-resistant (Rhir) mutants of Aspergillus nidulans was cloned, sequenced and compared with that of the wild-type, rhizoxin-sensitive (Rhis) strain . In all three Rhir mutants, the AAC codon for Asn-100 of the benA beta-tubulin gene was altered to ATC, coding for Ile . Sequence displacement experiments confirmed that the substitution of Ile for Asn-100 confers resistance to rhizoxin in this organism . The amino acid sequences of beta-tubulin surrounding the 100th amino acid residue from the N-terminus including Asn-100 are highly conserved with a few exceptions . The fission yeast Schizosaccharomyces pombe and the budding yeast Saccharomyces cerevisiae are naturally occurring Rhir organisms whose beta-tubulin genes encode Ile and Val respectively at the 100th amino acid residue . The Ile-100 of S . pombe and the Val-100 of S . cerevisiae were altered to Asn using site-directed mutagenesis and gene displacement techniques . The resultant haploid strains of these two yeasts uniquely expressing beta-tubulin (Asn-100) instead of beta-tubulin (Ile-100 or Val-100) were found to be Rhis . Haploid yeast expressing beta-tubulin (Asn-100) is normal except for its sensitivity to rhizoxin . These results suggest that rhizoxin resistance has a common basis in both naturally occurring species and experimentally selected mutants in the substitution of Ile or Val for Asn-100 in beta-tubulin. J Gen Microbiol, 1990 Jul, 136 ( Pt 7), 1271 - 7 Effect of ethanol on the phospholipid and fatty acid content of Schizosaccharomyces pombe membranes; Koukou AI et al.; Ethanol at concentrations up to 5% (v/v) had no effect on the growth of Schizosaccharomyces pombe, whereas concentrations over 7.5% were inhibitory . The major membrane phospholipids in S . pombe cells growing aerobically in the absence of added ethanol were phosphatidylinositol, phosphatidylcholine and phosphatidylethanolamine . Oleic acid (18:1) was the main fatty acid . When ethanol (7.5%) was added to aerobically growing cultures, the phosphatidylinositol content increased, whereas the 18:1 content decreased . Similar changes were observed in the membrane phospholipids of cells grown anaerobically without ethanol . However, the presence of ethanol in anaerobically growing cultures had an opposite effect on fatty acids, as the 18:1 content increased . The results support the idea that ethanol tolerance in S . pombe may be connected with a high content of 18:1 fatty acids, and with the ability to maintain a high rate of phospholipid biosynthesis. J Cell Sci, 1990 Jul, 96 ( Pt 3), 435 - 8 Continued DNA synthesis after a mitotic block in the double mutant cut1 cdc11 of the fission yeast Schizosaccharomyces pombe; Creanor J et al.; DNA synthesis is normally dependent on a cell having previously gone through mitosis . Hirano et al . (1986), however, found that DNA synthesis continued at the restrictive temperature in the double mutant cut1 cdc11 of Schizosaccharomyces pombe even though mitosis was blocked in some of the cells . We have confirmed this result with bulk DNA assays of asynchronous cultures . Synchronous cultures of a diploid double mutant at the restrictive temperature showed two peaks of incorporation with an interval between them that was approximately the same as the doubling time in cell length . Flow cytometry showed that the cells had increased their DNA content from 4C (the diploid value) to about 16C after 7h . The cytological appearance at this time was mixed, with uninucleate, binucleate and dead cells, but fluorescence measurements on single cells indicated that about half the population had single nuclei with about the 16C value and had therefore gone through two rounds of DNA synthesis without mitosis. Genes Dev, 1990 Jul, 4(7), 1141 - 8 Cloning of the Schizosaccharomyces pombe TFIID gene reveals a strong conservation of functional domains present in Saccharomyces cerevisiae TFIID; Hoffmann A et al.; The gene encoding the Schizosaccharomyces pombe TATA box-binding factor (TFIID) was cloned and sequenced . The gene contains three introns and codes for a polypeptide of 231 amino acids . The cDNA-expressed protein showed both TATA box-binding and basal transcription activities . The carboxy-terminal three-quarters of S . pombe TFIID shares an extraordinary degree of amino acid sequence homology with a corresponding region of Saccharomyces cerevisiae TFIID that has been shown to be necessary and sufficient for TATA box-binding and basal transcription activities . In contrast, the amino-terminal regions of the S . pombe and S . cerevisiae TFIIDs differ markedly in amino acid sequence and composition . Structure and function relationships of TFIID are discussed in light of these data. Mol Cell Biol, 1990 Jul, 10(7), 3524 - 34 Differential distribution of factors involved in pre-mRNA processing in the yeast cell nucleus; Potashkin JA et al.; The yeast cell nucleus has previously been shown to be divided into two regions by a variety of microscopic approaches . We used antibodies specific for the 2,2,7-trimethylguanosine cap structure of small nuclear ribonucleic acids (snRNAs) and for a protein component of small nuclear ribonucleoprotein particles to identify the distribution of small nuclear ribonucleoprotein particles within the yeast cell nucleus . These studies were performed with the fission yeast Schizosaccharomyces pombe and the budding yeast Saccharomyces cerevisiae . By using immunofluorescence microscopy and immunoelectron microscopy, most of the abundant snRNAs were localized to the portion of the nucleus which has heretofore been referred to as the nucleolus . This distribution of snRNAs is different from that found in mammalian cells and suggests that the nucleolar portion of the yeast nucleus contains functional domains in addition to those associated with RNA polymerase I activity. Proc Natl Acad Sci U S A, 1990 Jul, 87(13), 4917 - 21 Neurospora crassa a mating-type region; Staben C et al.; The a mating-type region of Neurospora crassa controls several major events in both the sexual and asexual phases of the fungal life cycle . This 3235-base-pair DNA segment is not homologous to the comparable genetic region of the A mating type . The unique a and A regions are bordered by nearly identical DNA sequences . The a genetic region contains at least two functional segments . One segment encodes a perithecium maturation function that is dependent on the second segment for phenotypic expression . This second a segment encodes a spliced mRNA that specifies the mt a-1 polypeptide . This polypeptide appears to be responsible for vegetative incompatibility, mating identity, and perithecium induction . The a-1 transcript is produced vegetatively and under conditions that induce sexual differentiation . The amino-terminal half of the mt a-1 polypeptide is homologous to the shorter Schizosaccharomyces pombe mat-Mc polypeptide . This homology and the properties of mt a-1 mutants suggest that the a-1 polypeptide segment that is homologous to the mat-Mc polypeptide may be primarily responsible for mating functions, while the distal segment is required for vegetative incompatibility. J Cell Sci, 1990 Jul, 96 ( Pt 3), 429 - 33 Changes in the rate of oxygen consumption in synchronous cultures of the fission yeast Schizosaccharomyces pombe; Novak B et al.; Oxygen consumption was measured with an oxygen electrode in synchronous cultures of S . pombe . There were changes during the cell cycle in the rate of oxygen uptake, which are most clearly shown as oscillations in acceleration curves (rate of the rate of uptake) . Under various conditions of selection and induction synchrony the acceleration curves are similar to those found earlier for CO2 production . As with CO2 production, the oscillations continued after a block to the DNA-division cycle . There were, however, two differences between oxygen uptake and CO2 production . The oxygen oscillations were more marked and also were out of phase by half a cycle . The respiratory coefficient therefore changes through the cycle. Can J Microbiol, 1990 Jun, 36(6), 390 - 4 Pattern of end growth of the fission yeast Schizosaccharomyces pombe; Miyata H et al.; The patterns of end growth of individual cells of Schizosaccharomyces pombe, wild-type cells (strain 972 h-), cells exposed to 8 mM hydroxyurea, and cdc mutants (cdc11-123 and cdc2-33), were investigated by time-lapse photomicrography . It was reconfirmed that there are three patterns of end growth: cells growing at the old end, at the new end, and at both ends from the beginning of the cell cycle . Cells that initiated growth at the old (new) end increased their growth rate at the new (old) end and became constant in their growth rate at the old (new) end when cells had their growth rate higher than a critical value: 0.08, 0.09, 0.08, and 0.11 microns/min in wild-type cells, cells exposed to hydroxyurea, cdc11-123 cells, and cdc2-33 cells, respectively . The critical value is proportional to the doubling time in length. Proc Natl Acad Sci U S A, 1990 Jun, 87(11), 4043 - 7 Introduction of large linear minichromosomes into Schizosaccharomyces pombe by an improved transformation procedure; Allshire RC; The efficiency of transformation of Schizosaccharomyces pombe has been increased 10- to 50-fold over previously reported methods . By using 1 microgram of plasmid, 7.0 x 10(5) transformants are regularly obtained . This increased transformation efficiency is mainly due to the inclusion of the cationic liposome-forming reagent Lipofectin in the protocol . Various parameters affecting transformation of Sc . pombe in the presence of Lipofectin have been examined . Lipofectin can also be used to increase transformation efficiency in Saccharomyces cerevisiae . It is also demonstrated that by using this improved transformation procedure, linear minichromosomes of greater than 500 kilobases can be introduced into Sc . pombe with relative ease . These minichromosomes can replicate as stable linear molecules upon reintroduction into Sc . pombe, demonstrating that Sc . pombe telomeres retain function when reintroduced as naked DNA . The ability of Sc . pombe to admit large DNA molecules indicates that it should be feasible to clone large DNA from other organisms in Sc . pombe. EMBO J, 1990 Jun, 9(6), 1929 - 37 Substrate recognition by RNase P and by the catalytic M1 RNA: identification of possible contact points in pre-tRNAs; Kahle D et al.; Modified bases were introduced into pre-tRNAs during in vitro RNA synthesis or by chemical modification . These RNAs were used as substrates for the catalytic M1 RNA and the RNase P holoenzyme from Schizosaccharomyces pombe . The synthetic approach permitted the insertion of 100% m7GTP into pre-tRNAs and this resulted in complete inhibition of the specific 5' processing reactions . Partially modified RNAs were obtained by chemical modifications of purines and uridines in the pre-tRNAs . This allowed detailed analyses of specific bases excluded in the products . With pre-tRNA(Ser) and initiator pre-tRNA(Met), strong effects were observed in the T arm and weaker effects in the anticodon stem . Only minor base exclusions were detected in the acceptor stem of pre-tRNA(Ser) and in the D arm of pre-tRNA(Met). Curr Genet, 1990 Jun, 17(6), 473 - 9 Evolutionary conservation of transcriptional machinery between yeast and plants as shown by the efficient expression from the CaMV 35S promoter and 35S terminator; Hirt H et al.; Complementation of fission yeast mutants by plant genomic libraries could be a promising method for the isolation of novel plant genes . One important prerequisite is the functioning of plant promoters and terminators in Schizosaccharomyces pombe and Saccharomyces cerevisiae . Therefore, we studied the expression of the bacterial beta-glucuronidase (GUS) reporter gene under the control of the Cauliflower Mosaic Virus (CaMV) 35S promoter and 35S terminator . We show here that S . pombe initiates transcription at exactly the same start site as was reported for tobacco . The 35S CaMV terminator is appropriately recognized leading to a polyadenylated mRNA of the same size as obtained in plant cells transformed with the same construct . Furthermore, the GUS-mRNA is translated into fully functional GUS protein, as determined by an enzymatic assay . Interestingly, expression of the 35S promoter in the budding yeast S . cerevisiae was found to be only moderate and about hundredfold lower than in S . pombe . To investigate whether different transcript stabilities are responsible for this enormous expression difference in the two yeasts, the 35S promoter was substituted by the ADH (alcohol dehydrogenase) promoter from fission yeast . In contrast to the differential expression pattern of the 35S promoter, the ADH promoter resulted in equally high expression rates in both fission and budding yeast, comparable to the 35S promoter in S . pombe . Since the copy number of the 35S-GUS constructs differs only by a factor of two in the two yeasts, it appears that differential recognition of the 35S promoter is responsible for the different transcription rates. Mol Cell Biol, 1990 Jun, 10(6), 2874 - 81 U1 small nuclear RNA from Schizosaccharomyces pombe has unique and conserved features and is encoded by an essential single-copy gene; Porter G et al.; We have cloned, sequenced, and disrupted the gene encoding U1 small nuclear RNA (snRNA) in the fission yeast Schizosaccharomyces pombe . This RNA is close in size and exhibits a high degree of secondary structure homology to human U1 RNA . There exist two regions of extended primary sequence identity between S . pombe and human U1 RNAs; the first comprises nucleotides involved in hydrogen bonding to 5' splice junctions, and the second is a single-stranded region which, in the human snRNA, forms part of the A protein binding site . S . pombe U1 lacks two nucleotides just following the 5' cap structure which are present in all other U1 homologs examined to date, and the region which corresponds to the binding site for the human 70K protein (molecular weight of 55,000) is more divergent than in other organisms . A putative upstream transcription signal is conserved in sequence and location among all loci encoding spliceosomal snRNAs in S . pombe with the exception of U6 . Disruption of the single-copy U1 gene, designated snu1, reveals that this RNA is indispensable for viability. EMBO J, 1990 Jun, 9(6), 1957 - 62 Structural and functional analysis of ypt2, an essential ras-related gene in the fission yeast Schizosaccharomyces pombe encoding a Sec4 protein homologue; Haubruck H et al.; Using the cloned Saccharomyces cerevisiae YPT1 gene as hybridization probe, a gene, designated ypt2, was isolated from the fission yeast Schizosaccharomyces pombe and found to encode a 200 amino acid long protein most closely related to the ypt branch of the ras superfamily . Disruption of the ypt2 gene is lethal . The bacterially produced ypt2 gene product is shown to bind GTP . A region of the ypt2 protein corresponding to but different from the 'effector region' of ras proteins is also different from that of ypt1 proteins of different species but identical to the 'effector loop' of the S.cerevisiae SEC4 gene product, a protein known to be required for vesicular protein transport . The S.pombe ypt2 gene under control of the S.cerevisiae GAL10 promoter is able to suppress the temperature-sensitive phenotype of a S . cerevisiae sec4 mutant, indicating a functional similarity of these GTP-binding proteins from the two very distantly related yeasts. EMBO J, 1990 Jun, 9(6), 1949 - 55 The ryh1 gene in the fission yeast Schizosaccharomyces pombe encoding a GTP-binding protein related to ras, rho and ypt: structure, expression and identification of its human homologue; Hengst L et al.; A gene, ryh1, of the fission yeast Schizosaccharomyces pombe encoding a GTP-binding protein of 201 amino acids and belonging to the ras superfamily was isolated using the protein-coding region of the cloned Saccharomyces cerevisiae YPT1 gene as hybridization probe . The ryh1 gene is interrupted by three introns . ryh1 null mutants are viable but unable to grow at temperatures greater than 35.5 degrees C . Invertase of ryh1- cells is properly secreted but has a faster electrophoretic mobility compared to that of wild-type cells . The temperature-sensitive phenotype of ryh1 null mutants is complemented by the human rab6 cDNA expressed either under transcriptional control of the S.pombe adh or the SV40 early promoter. Anal Biochem, 1990 May 15, 187(1), 94 - 7 A versatile microtiter assay for the universal cdc2 cell cycle regulator; Ducommun B et al.; A microassay for p34cdc2 based on the high affinity association between cdc2 and Schizosaccharomyces pombe p13suc1 has been developed . p13 purified from Escherichia coli was immobilized on microtiter plates and cellular lysate was incubated in the wells to allow the binding of cdc2 and its associated proteins . p34cdc2 was assayed either as a histone kinase or by immunological methods . The method was optimized for S . pombe cell extracts but can also be applied to other organisms such as Xenopus oocytes or HeLa cells . This rapid assay allows the specific determination of p34cdc2 histone H1 kinase activity in a very large number of samples. Mol Gen Genet, 1990 May, 221(3), 403 - 10 Effects of seven different mutations in the pho1 gene on enzymatic activity, glycosylation and secretion of acid phosphatase in Schizosaccharomyces pombe; Schwaninger R et al.; Structural gene mutants of the cell-surface glycoprotein acid phosphatase of Schizosaccharomyces pombe were analysed to define structural determinants that are responsible for enzymatic activity, N-glycosylation and secretion . All seven defined mutations cause a single amino acid substitution in the mature acid phosphatase protein and destroy the enzymatic activity . The mutational lesions are distributed throughout the pho1 gene . A ser to phe substitution at position 349 abolishes enzymatic activity only and does not affect glycosylation and secretion . Two mutations create a new N-glycosylation site by substitution of pro at position 56 by phe and ser, respectively . This new site is apparently used in the mutants . Their core-glycosylated acid phosphatase is slightly larger than that of the wild type . Overglycosylation seems not to affect secretion . Four different mutations (a gly to asp substitution at position 281 and ser to phe substitutions at positions 150, 271 and 277) cause intracellular accumulation of enzymatically inactive core-glycosylated acid phosphatase precursor . These mutational lesions apparently block transport of acid phosphatase from the endoplasmic reticulum to the Golgi apparatus. EMBO J, 1990 May, 9(5), 1417 - 22 Identification of ras-related, YPT family genes in Schizosaccharomyces pombe; Miyake S et al.; Screening for genes homologous to ras in Schizosaccharomyces pombe resulted in the isolation of a homolog of Saccharomyces cerevisiae YPT1 . This S . pombe gene, named ypt3, has a coding capacity of 214 amino acids interrupted by two introns, and is essential for cell growth . Two more YPT1 homologs were isolated from S . pombe using a part of the ypt3 gene as the probe . One of them, named ypt1, is highly homologous to S . cerevisiae YPT1 and mouse ypt1 and is essential for cell growth . This gene has four introns and encodes 203 amino acids . Its cDNA placed downstream of the S . cerevisiae GAL7 promoter could complement S . cerevisiae ypt1-, indicating that Sp ypt1 and Sc YPT1 are functionally homologous . The other isolate, named ryh1, and a fourth homolog, ypt2, have been characterized by Gallwitz and co-workers . The ypt1, ypt2 and ypt3 genes, but not ryh1, constitute a family, their products having double cysteine as their C terminus and serine in place of a glycine residue highly conserved in ras proteins (mammalian Gly-12 or S . pombe Gly-17) . The physiological roles of these genes appear to be distinct because each of them is indispensable for cell growth. EMBO J, 1990 May, 9(5), 1407 - 15 The developmental fate of fission yeast cells is determined by the pattern of inheritance of parental and grandparental DNA strands; Klar AJ; A key feature for development consists of producing sister cells that differ in their potential for cellular differentiation . Following two cell divisions, a haploid Schizosaccharomyces pombe cell produces one cell in four 'granddaughters' with a changed mating cell type, implying nonequivalence of sister cells in each of two consecutive cell divisions . The observed pattern of switching is analogous to the mammalian 'stem cell' lineage by which a cell produces one daughter like itself while the other daughter is advanced in its developmental program . It is tested here whether sisters differ because of unequal distribution of cytoplasmic and/or nuclear components to them or due to inheriting a specific parental DNA chain at the mating type locus . Only the DNA strand-segregation model predicts that those cells engineered to contain an inverted tandem duplication of the mating type locus should produce equivalent sisters . Consequently, two 'cousins' in four related granddaughter cells should switch . The results verified the prediction, thus establishing that all cells otherwise fully possess the potential to switch . Therefore, the program of cell type change in S.pombe cell lineages is determined by the pattern of DNA strand inheritance at the mating type locus . A specific DNA sequence present at the mating type locus is postulated to be the cause of developmental asymmetry between sister cells . A general model for cellular differentiation is proposed in which the act of DNA replication itself is hypothesized to produce developmentally nonequivalent sister genomes. Mol Cell Biol, 1990 May, 10(5), 2341 - 8 A gene family for acidic ribosomal proteins in Schizosaccharomyces pombe: two essential and two nonessential genes; Beltrame M et al.; We have cloned the genes for small acidic ribosomal proteins (A-proteins) of the fission yeast Schizosaccharomyces pombe . S . pombe contains four transcribed genes for small A-proteins per haploid genome, as is the case for Saccharomyces cerevisiae . In contrast, multicellular eucaryotes contain two transcribed genes per haploid genome . The four proteins of S . pombe, besides sharing a high overall similarity, form two couples of nearly identical sequences . Their corresponding genes have a very conserved structure and are transcribed to a similar level . Surprisingly, of each couple of genes coding for nearly identical proteins, one is essential for cell growth, whereas the other is not . We suggest that the unequal importance of the four small A-proteins for cell survival is related to their physical organization in 60S ribosomal subunits. Mol Cell Biol, 1990 May, 10(5), 1863 - 72 Functional analysis of a centromere from fission yeast: a role for centromere-specific repeated DNA sequences; Clarke L et al.; A circular minichromosome carrying functional centromere sequences (cen2) from Schizosaccharomyces pombe chromosome II behaves as a stable, independent genetic linkage group in S . pombe . The cen2 region was found to be organized into four large tandemly repeated sequence units which span over 80 kilobase pairs (kb) of untranscribed DNA . Two of these units occurred in a 31-kb inverted repeat that flanked a 7-kb central core of nonhomology . The inverted repeat region had centromere function, but neither the central core alone nor one arm of the inverted repeat was functional . Deletion of a portion of the repeated sequences that flank the central core had no effect on mitotic segregation functions or on meiotic segregation of a minichromosome to two of the four haploid progeny, but drastically impaired centromere-mediated maintenance of sister chromatid attachment in meiosis I . This requirement for centromere-specific repeated sequences could not be satisfied by introduction of random DNA sequences . These observations suggest a function for the heterochromatic repeated DNA sequences found in the centromere regions of higher eucaryotes. J Cell Sci, 1990 May, 96 ( Pt 1), 71 - 7 Distribution of tubulin and actin through the cell division cycle of the fission yeast Schizosaccharomyces japonicus var . versatilis: a comparison with Schizosaccharomyces pombe; Alfa CE et al.; Changes in the distribution of microtubules and F-actin through the cell division cycle of the fission yeast Schizosaccharomyces japonicus var . versatilis were investigated by fluorescence microscopy . The fluorescence images obtained with S . japonicus were markedly superior to those previously reported for S . pombe and revealed new details of cytoskeletal organization in this important genus . As in S . pombe, F-actin in S . japonicus was present as a concentration of 'dots' at the growing poles of interphase cells and as a filamentous equatorial ring directing the deposition of the cytokinetic septum . The transition between these two states occurred at late anaphase, in contrast to the situation in S . pombe where the appearance of the equatorial actin ring is tightly coupled to the early events of mitosis . During the course of cytokinesis in S . japonicus the actin ring constricted and broadened, suggesting that it is contractile . Microtubule organization in S . japonicus also revealed interesting differences from S . pombe . Whereas in S . pombe cytoplasmic microtubules are reinitiated from a pair of microtubule organizing centres (MTOCs) at the cell equator, in S . japonicus they arise by extensive microtubule growth from the spindle poles . Western blots of cell extracts enriched for tubulin by DEAE-Sephadex chromatography showed that, like S . pombe, S . japonicus contains two alpha-tubulins and a single beta-tubulin . Whilst the alpha 1- and beta-tubulins from the two species comigrated on one-dimensional polyacrylamide gels, the alpha 2 species were electrophoretically distinct . Although fundamental differences clearly exist between the two species, S . japonicus could prove to be a useful tool in basic studies of fission yeast cell biology. Trends Genet, 1990 May, 6(5), 150 - 4 Centromeres of budding and fission yeasts; Clarke L; Centromeres of the budding yeast Saccharomyces cerevisiae are structurally relatively simple, are specified by only about 125 base pairs of DNA, and contain no repeated DNA sequences . The centromere regions in the fission yeast Schizosaccharomyces pombe span many kilobase pairs of DNA and contain repeated DNA sequences that appear to be necessary for full centromere function . A portion of the repeated sequences is organized into a large inverted repeated structure in the centromere region of each S . pombe chromosome . Fission yeast provides an excellent model system for studying the role of repeated DNA sequences in centromere function. Biochem J, 1990 May 1, 267(3), 697 - 702 Purification and characterization of the invertase from Schizosaccharomyces pombe . A comparative analysis with the invertase from Saccharomyces cerevisiae; Moreno S et al.; Invertase (EC 3.2.1.26) was purified to homogeneity from exponentially growing cells of Schizosaccharomyces pombe fully de-repressed for synthesis of the enzyme, and was shown to be a high-molecular-mass glycoprotein that can be dissociated in the presence of 8 M-urea/1% SDS into identical subunits with an apparent molecular mass of 205 kDa . The carbohydrate moiety, accounting for 67% of the total mass, is composed of equimolar amounts of mannose and galactose . There is a small amount of glucosamine, which is probably involved in the linkage to the protein moiety, since the enzyme is sensitive to treatment with endoglycosidase H . The composition of the carbohydrate moiety resembles that found in higher-eukaryotic glycoproteins and differs from glycoproteins found in Saccharomyces cerevisiae . The protein portion of each subunit is a polypeptide of molecular mass 60 kDa, very similar to the invertase of Sacch . cerevisiae . Both proteins cross-react with antibodies raised against the protein fractions of the other, indicating that the two enzymes are similar. J Cell Biol, 1990 May, 110(5), 1617 - 21 Intramitotic controls in the fission yeast Schizosaccharomyces pombe: the effect of cell size on spindle length and the timing of mitotic events; Hagan IM et al.; We have used a new cinemicroscopy technique in combination with antitubulin immunofluorescence microscopy to investigate the timing of mitotic events in cells of the fission yeast Schizosaccharomyces pombe having lengths at division between 7 and 60 microns . Wild-type fission yeast cells divide at a length of 14 microns . Separation of daughter nuclei (anaphase B) proceeds at a rate of 1.6 +/- 0.2 microns min-1, until the spindle extends the length of the cell . Coincident with spindle depolymerization, the nuclei reverse direction and take up positions that will become the center of the two daughter cells . This post-mitotic nuclear migration occurs at a rate of 1.4 +/- 0.5 microns-1 . In cells in which the weel+ gene is overexpressed fivefold and that have an average length at mitosis of 28 microns, the rate of nuclear separation was only slightly reduced but, as spindles in these cells measure 20-22 microns, the duration of anaphase B was extended by approximately 40% . By contrast, in the mutant weel.50, which divides at 7 microns, both the rate and duration of anaphase B were indistinguishable from wild type . Nuclei reach the