J Vet Sci, 2004 Dec, 5(4), 359 - 67 Effect of probiotic containing Saccharomyces boulardii on experimental ochratoxicosis in broilers: hematobiochemical studies; B AS et al.; In the present investigation, the toxicopathological effects of ochratoxin A of 0.5 ppm on hematobiochemical parameters of broilers were studied with efficacy of dietary concentration of probiotic containing yeast culture Saccharomyces boulardii of 10 mg/kg of feed . One hundred twenty day old chicks were randomly divided into four groups, thirty chicks each . Groups A and C chicks were offered normal feed and that added with probiotic Saccharomyces boulardii respectively . The birds in group B were fed ochratoxin A of 0.5 ppm of feed . Where as, the birds of group D, were fed with ochratoxin A of 0.5 ppm along with probiotic Saccharomyces boulardii of 10 mg/kg of diet . Hematological studies carried revealed significant decrease in the haemoglobin and packed cell volume in birds of group B and reduced effect in birds of group D due to probiotic . Biochemical profiles revealed significant improvement in probiotic treated group D when compared with decreased values of Total protein, albumin, globulin and increased levels of serum creatinine and SGPT in birds of groups B. Biotechnol Lett, 2004 Oct, 26(19), 1521 - 7 Inferring ethanol tolerance of Saccharomyces and non-Saccharomyces yeasts by progressive inactivation; Pina C et al.; The kinetics of cell inactivation in the presence of ethanol at 20, 22.5% and 25% (v/v), was measured by progressive sampling and viable counting, and used as an inference of the ethanol resistance status of five non-Saccharomyces strains and one strain of Saccharomyces cerevisiae . The capacity of standard inocula of the same strains to establish growth at increasing initial ethanol concentrations was employed as a comparison . The effect of various different pre-culture conditions on the ethanol resistance of the 6 strains was analysed by the cell inactivation method and by the cell growth method . Exposing cells to 25% (v/v) ethanol for 4 min enabled the differentiation of the yeasts in terms of their resistance to ethanol . The results suggest that the two methods are generally concordant and that the cell inactivation method can, thus, be used to infer ethanol resistance of yeast strains. Pol Merkuriusz Lek, 2004, 17 Suppl 1, 22 - 6 {Assessment of usefulness of anti-Saccharomyces cerevisiae and anti-neutrophil cytoplasmic antibodies in patients with unspecific inflammatory bowel diseases}; Mokrowiecka A et al.; Ulcerative colitis (UC) and Crohn's disease (CD) are the nonspecific inflammatory bowel diseases with unknown etiology . Existing diagnostic methods are in many cases insufficient for proper diagnosis and choice of treatment method . In 10% cases of inflammatory bowel disease indeterminate colitis (IC) is described . It includes cases with diverse clinical manifestations, range and histopathological picture . New methods to distinguish inflammatory bowel diseases, chose proper treatment and monitor their activity are searched . Anti-Saccharomyces cerevisiae antibodies (ASCA) and antineutrophil cytoplasmatic antibodies (pANCA) seem to be a new markers used in this end . OBJECTIVE: Comparison of ASCA and pANCA appearance in CD and UC searching of correlation between serum levels of antibodies, activity and clinical picture of disease and appearance of ASCA and pANCA in IC . MATERIAL AND METHODS: We examined 63 patients with UC aged 24 to 74 (30 men and 33 women), 14 patients with CD aged 27 to 64 (10 men and 4 women) and 11 patients with IC aged 21 to 53 (4 men and 7 women) . Mean duration time of diseases respectively for UC, CD and IC was 7.9 (from 1.0 to 32), 9 (from 1 to 20) and 3.04 (0.5 to 7) years . Diagnosis in specific clinical picture was established with endoscopic and histopathologic examination . Disease activity was described according to Rachmilewitz scale in UC and in CD according to Crohn's disease activity index (CDAI) . Control group consisted of 24 healthy men . Examination of pANCA, IgA and IgG ACSA was performed with ELISA kits respectively by Congent Diagnostics and Genesis Diagnostic . RESULTS: There were 39 of UC patients and of CD patients with active disease according to Rachmilewitz scale (above 3 points) and CDAI . We observed statistically more often pANCA in patients with UC (58%; n=31) than in patients with CD (28%; n=4) (p<0.05) . Both IgA and IgG ACSCA occurred more often in patients with CD (57%; n=8) than in patients with UC (24%; n=15) (p<0.05) . There were no significant differences between antibodies according to duration time, activity, location and treatment method of diseases . Obtained results besides the help with understanding of pathologic mechanisms, indicate for use of antibodies as an diagnostic tool in inflammatory bowel diseases. Aliment Pharmacol Ther, 2004 Nov 15, 20(10), 1143 - 52 Anti-Saccharomyces cerevisiae mannan antibodies in inflammatory bowel disease: comparison of different assays and correlation with clinical features; Annese V et al.; BACKGROUND: Anti-Saccharomyces cerevisiae mannan antibodies have been proposed as a new serological marker associated with Crohn's disease . However, their clinical value is still unclear; furthermore, a standardization of anti-S . cerevisiae mannan measurements is lacking . AIM: In this study, we aimed to assess the correlation between anti-S . cerevisiae mannan detection and specific clinical features in Crohn's disease and ulcerative colitis . Moreover, we tested the concordance of four different anti-S . cerevisiae mannan assays . MATERIALS AND METHODS: Serum samples from 196 patients with Crohn's disease, 197 patients with ulcerative colitis and 100 unrelated healthy controls were tested for anti-S . cerevisiae mannan with a standard enzyme-linked immunosorbent assay method (Lille) by one of the authors (VP) . Subsequently, 60 randomly selected serum samples (27 Crohn's disease, 28 ulcerative colitis and five healthy controls) were tested for anti-S . cerevisiae mannan with three different commercial kits . RESULTS: With the Lille assay, anti-S . cerevisiae mannan were detected in 100 of 196 patients with Crohn's disease (51%; P < 0.0001 vs . controls), 32 of 197 patients with ulcerative colitis (16%; P < 0.02 vs . controls), and six of 100 controls (6%) . No correlation between presence of anti-S . cerevisiae mannan and specific clinical features was found in both ulcerative colitis and Crohn's disease patients . The percentages of anti-S . cerevisiae mannan detected with four different assays ranged from 28 (Bouty) up to 43% (Inova), but these differences did not reach statistical significance . The concordance rate of anti-S . cerevisiae mannan detection in the four assays was very low (11 concordant results of 60 samples, 18.3%) (k = 0.15) . No improvement of the concordance rate was obtained by modifying the suggested cut-off values (k = 0.20) . CONCLUSION: In this study, we confirm that anti-S . cerevisiae mannan are significantly more frequent in Crohn's disease patients compared with ulcerative colitis patients (P < 0.0001) and controls . However, no correlation with clinical features was found in both ulcerative colitis and Crohn's disease . The low prevalence of anti-S . cerevisiae mannan, at least in our population, and the low concordance rate between different assays, makes the clinical role of this marker questionable. Biochem J . 2004 Nov 24; {Epub ahead of print} Inositol phosphoryl ceramide synthase a regulator of intracellular levels of Diacylglcerol and ceramide during the G1 to S transition in Saccharomyces cerevisae; Cerbon J et al.; We recently reported that diacylglycerol generated during sphingomyelin synthesis plays an important role in protein kinase C activation and cell proliferation in Madin-Darby canine kidney cells (Cerbon, J . and Lopez-Sanchez, R.C . (2003) Biochem . J . 373, 917-924) . In yeast cells, Inositolphosphorylceramide synthase catalyzes the transfer of phosphoinositol from phosphatidylinositol to ceramide to form Inositol-phosphoryl ceramide and generates diacylglycerol . Here we show that during G1 to S transition after nitrogen-starvation, there was a significant increase in the synthesis of inositol-phosphoryl ceramide accompanied by a progressive elevation (up to 6 fold) in the level of DAG . The increased diacylglycerol levels were coincident with decrements in ceramide and sphingoid base levels, conditions that are adequate for the activation of the putative protein kinase C required for the G1 to S transition and proliferation of yeast cells . To separate the role of diacylglycerol generated during Inositol phosphorylceramide synthesis from that originated of other sources, we utilized beta-chloroalanine and myriocin, inhibitors of serine-palmitoyl CoA transferase, the first committed step in sphingolipid synthesis, to avoid accumulation of sphingolipid intermediates . When the synthesis of sphingolipids was inhibited, diacylglycerol accumulation was significantly reduced and the G1 to S transition was blocked; such blockage was avoided by metabolic complementation with phytosphingosine . The ratio diacylglycerol / ceramide was 0.27 and changed to 2.0 during growth reinitiation suggesting that the synthesis of phosphosphingolipids could act to switch a growth arrest (increased ceramide) to a mitogenic signal (increased diacylglycerol) and this signaling process is preserved in yeast and mammalian cells. Yeast, 2004 Oct 30, 21(14), 1195 - 203 Developing methods and strains for genetic studies in the Saccharomyces bayanus var . uvarum species; Talarek N et al.; For years, Saccharomyces cerevisiae has been used as a model organism to gain insight into complex biological processes . The study of closely related yeast species may be critical for understanding the molecular mechanism of evolution . Among those species, S . bayanus var . uvarum could be particularly pertinent because of the availability of its genome sequence . However, to date, in that species genetic studies are problematical due to the lack of standard strains collection and genetic methods . Here, we have developed heterothallic S . bayanus var . uvarum strains and obtained stable haploid strains . We further used UV-induced mutation and gene disruption to create a collection of auxotrophic derivatives . Finally, we have elaborated or improved methods to cultivate cells, obtain zygotes and spores and to transform this species . All these tools can now be used by the scientific community to study the biology of this species . copyright 2004 John Wiley & Sons, Ltd. In Silico Biol . 2004 Jun 30;4(3):0034 {Epub ahead of print} A search tool for identification and analysis of conserved sequence patterns in Saccharomyces spp . orthologous promoter; Kohli DK et al.; We describe a web-based resource to identify, search and analyze sequence patterns conserved in the multiple sequence alignments of orthologous promoters from closely related / distant Saccharomyces spp . The webtool interfaces with a database where conserved sequence patterns (greater than 4 bp) have been previously extracted from genome-wide promoter alignments, allowing one to carry out user-defined genome-wide searches for conserved sequences to assist in the discovery of novel promoter elements based on comparative genomics . The web-based server can be accessed at anand/sacch_prom_pat.html. Inflamm Bowel Dis, 2004 Sep, 10(5), 521 - 8 Disease location, anti-Saccharomyces cerevisiae antibody, and NOD2/CARD15 genotype influence the progression of disease behavior in Crohn's disease; Smith BR et al.; BACKGROUND: Crohn's disease (CD) is characterized by heterogeneity of phenotype . The Vienna classification can be used to classify CD, and recent data illustrate that behavior evolves over the course of the disease . Clinical and biological influences on disease progression remain unclear . We examined the associations of CD disease progression at diagnosis and for up to 20 years of follow-up . METHODS: Two hundred thirty-one well-characterized CD patients were studied . Demographic, clinical, and NOD2/CARD15 data were collected . Disease behavior according to the Vienna classification was assessed at diagnosis and for up to 20 years following diagnosis . RESULTS: At diagnosis, 70% of patients had inflammatory disease, 9% stricturing, and 21% penetrating . Early age at diagnosis was associated with ileocolonic and upper GI disease (p = 0.015), and positive anti-Saccharomyces cerevisiae antibody (ASCA) was associated with ileal involvement (p = 0.008) . Smoking was relatively protective against colonic, rather than ileal involvement at diagnosis (p < 0.02) . At 20 years, 92% had progressed to a more severe disease type . Patients who progress to a more severe disease type require more frequent surgery (p < 0.00001) . Multivariate analysis found disease progression to be associated with ileal disease location (p = 0.001) and positive ASCA (p = 0.003) . Variant NOD2/CARD15 alleles were protective against rapid progression of disease phenotype (p = 0.04) . The presence of perianal disease was independent of intestinal penetrating disease . CONCLUSIONS: The progression of disease type in CD is associated with the need for more frequent surgery . Rapid progression is associated with ileal disease and positive ASCA, and delayed progression is associated with variant NOD2/CARD15 alleles . Consideration should be given to a separate Vienna classification for perianal disease . Dig Dis Sci, 2004 Aug, 49(7-8), 1311 - 7 Studies on the time course of the effects of the probiotic yeast Saccharomyces boulardii on electrolyte transport in pig jejunum; Schroeder B et al.; Orally administered Saccharomyces boulardii (synonym Saccharomyces cerevisiae Hansen CBS 5926) has already been shown to affect relevant functions of the mucosa in pig jejunum such as lowering the secretory response to theophylline or stimulating sodium/glucose cotransport, but knowledge of time-dependent relationship is minimal . In this study we examined the effects of S . boulardii on sodium (Na+) and chloride (Cl-) transport in pig jejunum under nonstimulated (basal) and stimulated (secretory) conditions . For this purpose the conventional Ussing chamber method was used for measuring electrical parameters (short circuit currents, Isc; tissue conductances, G(T)) and electrolyte transport of isolated intact jejunal epithelia in the absence and presence of the secretagogue theophylline (10 mM, serosal side) . Time profiles of the mucosa response were assessed by treating animals perorally with S . boulardii for 0 (control), 3, 8, and 16 days . Intestinal tissues were obtained from growing pigs in the weight range between 25 and 40 kg . All animals were fed twice daily and received 1.0-1.6 kg/day of a standard diet avoiding probiotics as food additives . After a 9- to 10-day adaptation period the diets for treated animals were supplemented with approximately 1.8 x 10(7) colony forming units (CFU)/g feed of the probiotic . Whereas basal tissue conductances were not affected by treatment duration, basal Isc values decreased significantly during 8 days of treatment, by 26%, indicating a lower electrogenic net ion transport, which, however, was reconstituted after 16 days . This effect could be explained by almost the same reduction of basal Jms of Na+ during 8 days of treatment, whereas respective flux rates in the opposite direction remained stable . Under basal conditions unidirectional and net flux rates of Cl- were not affected by S . boulardii . Induction of secretory conditions by theophylline revealed pronounced increases in net Cl- secretion but this effect was more than 60% lower after 8-day S . boulardii application, and this was reflected by a respectively lower Isc stimulation . Interestingly, this inhibitory effect on the secretory response could no longer be observed in the 16-day group . And this was reflected by a respectively lower Isc stimulation . A similar effect could be observed regarding net Na+ flux rates . Residual fluxes were affected neither by S . boulardii nor by theophylline, therefore, Isc values can be explained completely by respective Na+ and Cl- fluxes . In conclusion, S . boulardii has specific duration-dependent effects on the secretory response of the pig jejunal mucosa which developed during 8-day treatment but disappeared during further application . Thus, this study supports the concept that probiotics may exert beneficial effects in the gastrointestinal tract. Genetics . 2004 Sep 15; {Epub ahead of print} X-ray survival characteristics and genetic analysis for nine Saccharomyces deletion mutants that affect radiation sensitivity; Game JC et al.; The availability of a genome-wide set of Saccharomyces deletion mutants provides a chance to identify all the yeast genes involved in DNA repair . Using X-rays, we are screening these mutants to identify additional genes that show increased sensitivity to the lethal effects of ionizing radiation . For each mutant identified as sensitive, we are confirming that the sensitivity phenotype co-segregates with the deletion allele and are obtaining multipoint survival-versus-dose assays in at least two haploid and one homozygous diploid strains . We present data for deletion mutants involving the genes DOT1, MDM20, NAT3, SPT7, SPT20, GCN5, HFI1, DCC1 and VID21/EAF1, and discuss their potential roles in repair . Eight of these genes have a clear radiation-sensitive phenotype when deleted, but the ninth, GCN5, has at most a borderline phenotype . None of the deletions confer substantial sensitivity to ultra-violet radiation, although one or two may confer marginal sensitivity . The DOT1 gene is of interest because its only known function is to methylate one lysine residue in the core of the histone H3 protein . We find that histone H3 mutants (supplied by K . Struhl) in which this residue is replaced by other amino-acids are also X-ray sensitive, seeming to confirm that methylation of the lysine-79 residue is required for effective repair of radiation damage. Inflamm Bowel Dis, 2004 May, 10(3), 234 - 9 Effects of disease activity on anti-Saccharomyces cerevisiae antibodies: implications for diagnosis and follow-up of children with Crohn's disease; Canani RB et al.; BACKGROUND: To determine diagnostic accuracy of anti-Saccharomyces cerevisiae antibodies (ASCA) in identifying children with inflammatory bowel disease (IBD) and to differentiate Crohn's disease (CD) from other IBD forms; and to determine the effect of medical or surgical treatment and of disease location and activity on ASCA titers . METHODS: Serum samples were obtained from 196 IBD children and 142 controls . ASCA IgA and IgG titers were measured by ELISA . Measurements were repeated during the follow up of CD children . RESULTS: ASCA titers were significantly higher in CD than in other IBD and in control patients . Combination of IgA and IgG ASCA positivity was highly specific for CD . Medical treatment and disease location did not influence assay results . Significantly lower ASCA titers were obtained in CD children with intestinal resection compared to CD-affected children who did not undergo surgical resection . ASCA titers correlated significantly with disease activity, and children with severe active disease showed higher ASCA values compared to those in remission . A significant reduction of ASCA was observed during the follow-up of CD children when clinical remission was achieved . CONCLUSIONS: The diagnostic accuracy of ASCA is influenced by disease activity and this suggests an additional use for the follow-up of CD children of this assay. Gut, 2004 Aug, 53(8), 1117 - 22 Anti-Saccharomyces cerevisiae antibody (ASCA) positivity is associated with increased risk for early surgery in Crohn's disease; Forcione DG et al.; BACKGROUND: Anti-Saccharomyces cerevisiae antibodies (ASCA) are a specific but only moderately sensitive diagnostic marker for Crohn's disease . We sought to explore the role of ASCA as a prognostic marker for aggressive disease phenotype in Crohn's disease . AIMS: To determine the role of ASCA status as a risk factor for early surgery in Crohn's disease . SUBJECTS: We performed a case control study in a cohort of patients, newly diagnosed with Crohn's disease, between 1991 and 1999 . All patients were followed for at least three years . Case subjects (n = 35) included those who had major surgery for Crohn's disease within three years of diagnosis . Controls (n = 35) included patients matched to cases for age, sex, disease location, and smoking status, and who did not undergo major surgery for Crohn's disease within three years of diagnosis . METHODS: Blinded assays were performed on serum for ASCA (immunoglobulin (Ig)A and IgG) . A paired analysis of cases-controls was performed to test for the association between ASCA status and risk of early surgery . RESULTS: ASCA IgA was strongly associated with early surgery (odds ratio (OR) 8.5 (95% confidence interval (CI) 2.0-75.9); p = 0.0013) . ASCA IgG+ and ASCA IgG+/IgA+ patients were also at increased risk for early surgery (OR 5.5 (95% CI 1.2-51.1), p = 0.0265; and OR 5.0 (95% CI 1.1-46.9), p = 0.0433, respectively) . The association between ASCA and early surgery was evident in patients requiring surgery for ileal or ileocolonic disease . CONCLUSIONS: Patients with Crohn's disease who are positive for ASCA IgA, IgG, or both, may define a subset of patients with Crohn's disease at increased risk for early surgery. J Biol Chem, 2004 Aug 27, 279(35), 36586 - 92 Epub 2004 Jun 25. Regulation of unsaturated fatty acid biosynthesis in Saccharomyces: the endoplasmic reticulum membrane protein, Mga2p, a transcription activator of the OLE1 gene, regulates the stability of the OLE1 mRNA through exosome-mediated mechanisms; Kandasamy P et al.; The Saccharomyces cerevisiae OLE1 gene encodes a membrane-bound Delta9 fatty-acid desaturase, whose expression is regulated through transcriptional and mRNA stability controls . In wild type cells grown on fatty acid-free medium, OLE1 mRNA has a half-life of 10 +/- 1.5 min (basal stability) that becomes highly unstable when cells are exposed to unsaturated fatty acids (regulated stability) . Activation of OLE1 transcription is dependent on N-terminal fragments of two membrane proteins, Mga2p and Spt23p, that are proteolytically released from the membrane by a ubiquitin-mediated mechanism . Surprisingly, disruption of the MGA2 gene also reduces the half-life of the OLE1 transcript and abolishes fatty acid regulated instability . Disruption of its cognate, SPT23, has no effect on the half-life of the mRNA . Mga2p appears to have two distinct functions with respect to the OLE1 mRNA stability: a stabilizing effect in cells grown in fatty acid-free medium and a destabilizing function in cells that are exposed to unsaturated fatty acids . These functions are independent of OLE1 transcription and can confer basal and regulated stability on OLE1 mRNAs that are produced under the control of the unrelated GAL1 promoter . Expression of soluble, N-terminal fragments of Mga2p stabilize the transcript but do not confer fatty acid-regulated instability on the mRNA suggesting that the stabilizing functions of Mga2p do not require membrane processing and that modifications to the protein introduced during proteolysis may play a role in the destabilizing effect . An analysis of mutants that are defective in mRNA degradation indicate that the Mga2p-requiring control mechanism that regulates the fatty acid-mediated instability of the OLE1 transcript acts by activating exosomal 3' --> 5'-exonuclease degradation activity. Microbiology, 2004 Jun, 150(Pt 6), 1983 - 90 Saccharomyces kluyveri FAD3 encodes an omega3 fatty acid desaturase; Oura T et al.; Fungi, like plants, are capable of producing the 18-carbon polyunsaturated fatty acids linoleic acid and alpha-linolenic acid . These fatty acids are synthesized by catalytic reactions of Delta12 and omega3 fatty acid desaturases . This paper describes the first cloning and functional characterization of a yeast omega3 fatty acid desaturase gene . The deduced protein encoded by the Saccharomyces kluyveri FAD3 gene (Sk-FAD3) consists of 419 amino acids, and shows 30-60 % identity with Delta12 fatty acid desaturases of several eukaryotic organisms and 29-31 % identity with omega3 fatty acid desaturases of animals and plants . During Sk-FAD3 expression in Saccharomyces cerevisiae, alpha-linolenic acid accumulated only when linoleic acid was added to the culture medium . The disruption of Sk-FAD3 led to the disappearance of alpha-linolenic acid in S . kluyveri . These findings suggest that Sk-FAD3 is the only omega3 fatty acid desaturase gene in this yeast . Furthermore, transcriptional expression of Sk-FAD3 appears to be regulated by low-temperature stress in a manner different from the other fatty acid desaturase genes in S . kluyveri. Brief Bioinform, 2004 Mar, 5(1), 9 - 22 Saccharomyces genome database: underlying principles and organisation; Dwight SS et al.; A scientific database can be a powerful tool for biologists in an era where large-scale genomic analysis, combined with smaller-scale scientific results, provides new insights into the roles of genes and their products in the cell . However, the collection and assimilation of data is, in itself, not enough to make a database useful . The data must be incorporated into the database and presented to the user in an intuitive and biologically significant manner . Most importantly, this presentation must be driven by the user's point of view; that is, from a biological perspective . The success of a scientific database can therefore be measured by the response of its users - statistically, by usage numbers and, in a less quantifiable way, by its relationship with the community it serves and its ability to serve as a model for similar projects . Since its inception ten years ago, the Saccharomyces Genome Database (SGD) has seen a dramatic increase in its usage, has developed and maintained a positive working relationship with the yeast research community, and has served as a template for at least one other database . The success of SGD, as measured by these criteria, is due in large part to philosophies that have guided its mission and organisation since it was established in 1993 . This paper aims to detail these philosophies and how they shape the organisation and presentation of the database. Antonie Van Leeuwenhoek, 2004 May, 85(4), 297 - 304 Trichomonascus petasosporus sp . nov . and Sympodiomyces indianaensis sp . nov., two new members of the Saccharomycetales; Kurtzman CP; Two new yeasts are described that were recognized as novel from nucleotide divergence in domains D1/D2 of 26S rDNA . The new species and their type strains are the following: Trichomonascus petasosporus NRRL YB-2092T (CBS 9602T), mating type a, NRRL YB-2093 (CBS 9603), mating type alpha, and Sympodiomyces indianaensis NRRL YB-1950T (CBS 9600T) . Phylogenetic analysis placed the two new taxa, which are sister species, in the Sympodiomyces clade near Blastobotrys/Stephanoascus farinosus . Placement of Trichomonascus in the Saccharomycetales resolves the earlier uncertainties surrounding the classification of this morphologically unusual genus. Clin Exp Immunol, 2004 Mar, 135(3), 490 - 6 Anti-Saccharomyces cerevisiae antibodies (ASCA) in Crohn's disease are associated with disease severity but not NOD2/CARD15 mutations; Walker LJ et al.; Anti-Saccharomyces cerevisiae antibodies (ASCAs) have been proposed as serological markers, which may differentiate Crohn's disease (CD) from ulcerative colitis (UC) and predict disease phenotype . Their importance in pathogenesis is unproven . We investigated the relationship between ASCAs, disease phenotype and NOD2/CARD15 genotype in CD and whether ASCAs were related to antibodies to other fungal proteins . Serum from 228 patients {143 CD, 75 UC, 10 with indeterminate colitis (IC)} and 78 healthy controls (HC) were assayed for ASCA . Antibodies (IgA, IgG) to other fungal proteins (Fusarium species ATC20334, Mycoprotein) were measured in the same samples using an in-house enzyme-linked immunosorbent assay (ELISA) assay . ASCAs were present in 57% of CD, 19% of UC, 30% of IC and 8% of HCs . ASCA-positive status was a predictor for CD with sensitivity of 57%, specificity of 87%, positive predictive value of 78% and negative predictive value of 68% . ASCA was associated with proximal (gastroduodenal and small bowel involvement) rather than purely colonic disease (P < 0.001) and with a more severe disease phenotype and requirement for surgery over a median follow-up time of 9 years (P < 0.0001) . No associations with NOD2/CARD15 mutations were seen . There was no association between ASCA and antibodies to MP (IgA or IgG) . These data implicate ASCA as a specific marker of disease location and progression in CD, emphasizing the heterogeneity within IBD. Cell Biol Int, 2004, 28(3), 193 - 7 Farnesyl diphosphate synthase activity affects ergosterol level and proliferation of yeast Saccharomyces cerevisae; Karst F et al.; The yeast farnesyl diphosphate synthase (FPPS) gene was engineered so as to construct allelic forms giving various activities of the enzyme . One of the substitutions was F96W in the chain length determination region . The other, K197, conserved within a consensus sequence found in the majority of FPP and GGPP synthases, was substituted by R, E and V . An intricate correlation has been found between the FPPS activity, the amount of ergosterol synthesized and cell growth of a mutant strain defective in FPPS . About 40% of wt FPPS activity was sufficient to support normal growth of the mutant . With further decline of FPPS activity (20 down to 3%) the amount of ergosterol remained unchanged at approximately 0.16% (vs dry weight), whereas growth yield decreased and lag times increased . We postulate that, in addition to ergosterol initiating and maintaining growth of yeast cells, FPP and/or its derivatives participate in these processes. Cell Biol Int, 2004, 28(3), 171 - 8 Evolution of multicellularity in Metazoa: comparative analysis of the subcellular localization of proteins in Saccharomyces, Drosophila and Caenorhabditis; Hazkani-Covo E et al.; A comparison of the subcellular assignments of proteins between the unicellular Saccharomyces cerevisiae and the multicellular Drosophila melanogaster and Caenorhabditis elegans was performed using a computational tool for the prediction of subcellular localization . Nine subcellular compartments were studied: (1) extracellular domain, (2) cell membrane, (3) cytoplasm, (4) endoplasmic reticulum, (5) Golgi apparatus, (6) lysosome, (7) peroxisome, (8) mitochondria, and (9) nucleus . The transition to multicellularity was found to be characterized by an increase in the total number of proteins encoded by the genome . Interestingly, this increase is distributed unevenly among the subcellular compartments . That is, a disproportionate increase in the number of proteins in the extracellular domain, the cell membrane, and the cytoplasm is observed in multicellular organisms, while no such increase is seen in other subcellular compartments . A possible explanation involves signal transduction . In terms of protein numbers, signal transduction pathways may be roughly described as a pyramid with an expansive base in the extracellular domain (the numerous extracellular signal proteins), progressively narrowing at the cell membrane and cytoplasmic levels, and ending in a narrow tip consisting of only a handful of transcription modulators in the nucleus . Our observations suggest that extracellular signaling interactions among metazoan cells account for the uneven increase in the numbers of proteins among subcellular compartments during the transition to multicellularity. Int J Colorectal Dis, 2004 Jul, 19(4), 319 - 24 Epub 2004 Jan 27. Optimising the diagnostic value of anti-Saccharomyces cerevisiae-antibodies (ASCA) in Crohn's disease; Klebl FH et al.; BACKGROUND/AIMS: Anti-Saccharomyces cerevisiae-antibodies (ASCA) are used to discriminate Crohn's disease (CD) from ulcerative colitis (UC) . ASCA tests are not standardised and different methods for ASCA detection exist . This study was undertaken to compare ASCA tests and to clarify their diagnostic value . PATIENTS/METHODS: One hundred and two sera from CD patients, 53 from UC patients, and 50 sera from normal controls were examined for ASCA IgA and ASCA IgG using four different ELISA (Aesku.lab, Inova, Euroimmun, and Medipan) and indirect immunofluorescence (Euroimmun) . Sensitivity, specificity, and positive and negative predictive values (PPV and NPV) for CD in this population were determined . Agreement between tests was expressed by kappa statistics . RESULTS/FINDINGS: The presence of either ASCA IgA or ASCA IgG had a PPV between 77 and 88% . Only the combined presence of ASCA IgA and ASCA IgG had a specificity which was constantly above 90% . Sensitivity, specificity, PPV, and NPV were then: Inova: 53, 97, 95, and 68%; Euroimmun ELISA: 46, 97, 94, and 66%; Aesku.lab: 50, 97, 94, and 66%; Medipan: 30, 98, 94, and 59%; indirect immunofluorescence: 51, 97, 94, and 66% . Agreement between ELISA test results of the Aesku.lab, Euroimmun, Medipan IgA and Inova systems was good (kappa: 0.63-0.79); between the Medipan IgG ELISA or indirect immunofluorescence and the others, it was lower (kappa: 0.33-0.6) . If both ASCA IgA and ASCA IgG were detected by indirect immunofluorescence as well as ELISA, specificity for CD increased to >99% at a sensitivity of 23-38% . INTERPRETATION/CONCLUSIONS: The combined detection of ASCA IgA and ASCA IgG by indirect immunofluorescence as well as ELISA may optimise the discrimination of CD from UC. Nucleic Acids Res, 2004 Jan 23, 32(2), 511 - 21 Print 2004. Interaction of Saccharomyces Cdc13p with Pol1p, Imp4p, Sir4p and Zds2p is involved in telomere replication, telomere maintenance and cell growth control; Hsu CL et al.; Telomeres are the physical ends of eukaryotic chromosomes . They are important for maintaining the integrity of chromosomes and this function is mediated through a number of protein factors . In Saccharomyces cerevisiae, Cdc13p binds to telomeres and affects telomere maintenance, telomere position effects and cell cycle progression through G(2)/M phase . We identified four genes encoding Pol1p, Sir4p, Zds2p and Imp4p that interact with amino acids 1-252 of Cdc13p using a yeast two-hybrid screening system . Interactions of these four proteins with Cdc13p were through direct protein-protein interactions as judged by in vitro pull-down assays . Direct protein-protein interactions were also observed between Pol1p-Imp4p, Pol1p-Sir4p and Sir4p-Zds2p, whereas no interaction was detected between Imp4p-Sir4p and Zds2p-Imp4p, suggesting that protein interactions were specific in the complex . Pol1p was shown to interact with Cdc13p . Here we show that Zds2p and Imp4p also form a stable complex with Cdc13p in yeast cells, because Zds2p and Imp4p co-immunoprecipitate with Cdc13p, whereas Sir4p does not . The function of the N-terminal 1-252 region of Cdc13p was also analyzed . Expressing Cdc13(252-924)p, which lacks amino acids 1-252 of Cdc13p, causes defects in progressive cell growth and eventually arrested in the G(2)/M phase of the cell cycle . These growth defects were not caused by progressive shortening of telomeres because telomeres in these cells were long . Point mutants in the amino acids 1-252 region of Cdc13p that reduced the interaction between Cdc13p and its binding proteins resulted in varying level of defects in cell growth and telomeres . These results indicate that the interactions between Cdc13(1-252)p and its binding proteins are important for the function of Cdc13p in telomere regulation and cell growth . Together, our results provide evidence for the formation of a Cdc13p-mediated telosome complex through its N-terminal region that is involved in telomere maintenance, telomere length regulation and cell growth control. PLoS Biol . 2004 Jan;2(1):E21 . Epub 2004 Jan 20. Role of saccharomyces single-stranded DNA-binding protein RPA in the strand invasion step of double-strand break repair; Wang X et al.; The single-stranded DNA (ssDNA)-binding protein replication protein A (RPA) is essential for both DNA replication and recombination . Chromatin immunoprecipitation techniques were used to visualize the kinetics and extent of RPA binding following induction of a double-strand break (DSB) and during its repair by homologous recombination in yeast . RPA assembles at the HO endonuclease-cut MAT locus simultaneously with the appearance of the DSB, and binding spreads away from the DSB as 5' to 3' exonuclease activity creates more ssDNA . RPA binding precedes binding of the Rad51 recombination protein . The extent of RPA binding is greater when Rad51 is absent, supporting the idea that Rad51 displaces RPA from ssDNA . RPA plays an important role during RAD51-mediated strand invasion of the MAT ssDNA into the donor sequence HML . The replication-proficient but recombination-defective rfa1-t11 (K45E) mutation in the large subunit of RPA is normal in facilitating Rad51 filament formation on ssDNA, but is unable to achieve synapsis between MAT and HML . Thus, RPA appears to play a role in strand invasion as well as in facilitating Rad51 binding to ssDNA, possibly by stabilizing the displaced ssDNA. Dig Liver Dis, 2003 Dec, 35(12), 862 - 8 Diagnostic role and clinical correlates of anti-Saccharomyces cerevisiae antibodies (ASCA) and anti-neutrophil cytoplasmic antibodies (p-ANCA) in Italian patients with inflammatory bowel diseases; Saibeni S et al.; BACKGROUND: Anti-Saccharomyces cerevisiae antibodies (ASCA) and perinuclear anti-neutrophil cytoplasmic antibodies (p-ANCA) are serological markers associated, respectively, with Crohn's disease and ulcerative colitis, whose clinical significance and possible diagnostic role are still poorly defined . AIMS: (a) To evaluate the sensitivity, specificity and predictive values of isolated and combined ASCA and p-ANCA assays in a large cohort of Italian patients with inflammatory bowel disease (IBD) and (b) to assess whether their presence is associated with particular clinical features of the disease . PATIENTS AND METHODS: Hundred and forty-six IBD patients (93 with Crohn's disease and 53 with ulcerative colitis) and 54 control patients were enrolled in the study . ASCA (IgA and IgG) and p-ANCA were determined by means of enzyme-linked immunosorbent assay and indirect immunofluorescence, respectively . RESULTS: The specificities were excellent for both tests (ASCA in Crohn's disease, 98.1% both for IgA and IgG, and p-ANCA in ulcerative colitis, 92.5%); however, the sensitivities of both tests were low (59.1% for ASCA IgA, 44.1% for ASCA IgG, 39.6% for p-ANCA) . ASCA specificity and positive predictive value reached 100% when positivity for both IgA and IgG was present . No significant association was found between the presence of a specific serological marker and patients' clinical features . CONCLUSIONS: This study confirms the low prevalence of p-ANCA observed in ulcerative colitis patients from the Mediterranean area . The low sensitivity of ASCA and p-ANCA, despite their rather high specificity, renders them of little value in the screening of the general population, where the prevalence of IBD is low . However, in our series, a double positivity for ASCA IgA and IgG identifies with certainty the presence of Crohn's disease. J Evol Biol, 2003 May, 16(3), 429 - 37 A role for the mismatch repair system during incipient speciation in Saccharomyces; Greig D et al.; The cause of reproductive isolation between biological species is a major issue in the field of biology . Most explanations of hybrid sterility require either genetic incompatibilities between nascent species or gross physical imbalances between their chromosomes, such as rearrangements or ploidy changes . An alternative possibility is that genomes become incompatible at a molecular level, dependent on interactions between primary DNA sequences . The mismatch repair system has previously been shown to contribute to sterility in a hybrid between established yeast species by preventing successful meiotic crossing-over leading to aneuploidy . This system could also promote or reinforce the formation of new species in a similar manner, by making diverging genomes incompatible in meiosis . To test this possibility we crossed yeast strains of the same species but from diverse historical or geographic sources . We show that these crosses are partially sterile and present evidence that the mismatch repair system is largely responsible for this sterility. Korean J Gastroenterol, 2003 Oct, 42(4), 297 - 302 {Diagnostic role of anti-Saccharomyces cerevisiae and antineutrophil cytoplasmic autoantibodies in pediatric inflammatory bowel disease}; Kim JE et al.; BACKGROUND/AIMS: Combined measurement of perinuclear antineutrophil cytoplasmic autoantibodies (pANCA) and anti-Saccharomyces cereviseae mannan antibodies (ASCA) has recently been suggested as a valuable diagnostic approach to inflammatory bowel disease (IBD) in the pediatric age group . The aim of this study was to test the accuracy of the assay using pANCA and ASCA in diagnosing pediatric ulcerative colitis (UC) and Crohn's disease (CD) . METHODS: Sera were collected from 25 patients with IBD (17 with CD, 8 with UC) and 32 healthy controls . The levels of pANCA and ASCA were determined by using a standard indirect immunofluorescence technique on ethanol-fixed granulocytes and an ELISA assay, respectively . RESULTS: In patients with UC, the sensitivity, specificity, and positive predictive value of the pANCA test were 38%, 88%, and 60%, respectively . Such values were not changed significantly in the case of positive pANCA and negative ASCA . The sensitivity, specificity, and positive predictive value of ASCA test in diagnosing CD were 71%, 88%, and 92%, respectively . The combination of pANCA negative and ASCA positive was not significant . CONCLUSIONS: ASCA and pANCA assays are highly disease specific for CD and UC, respectively . These serological tests can assist clinicians in diagnosing and categorizing patients with IBD and may be useful in making therapeutic decisions. Biotechnol Lett, 2003 Oct, 25(20), 1735 - 8 The first isolation of two types of trifluoroleucine resistant mutants of Saccharomyces servazzii; Tominaga T et al.; Saccharomyces servazzii plays a crucial role in the making of Japanese radish pickles . To make more flavorsome pickles, we sought to generate trifluoroleucine-resistant mutants of S . servazzii . The three resulting mutants could be classified into two types: one that produces more isoamyl alcohol than the parental strain, and one that produces less . The first type has been well documented in Saccharomyces cerevisiae but the latter appears to be novel and has been characterized as such. Eur J Gastroenterol Hepatol, 2003 Dec, 15(12), 1281 - 5 Do high serum levels of anti-Saccharomyces cerevisiae antibodies result from a leakiness of the gut barrier in Crohn's disease? Harrer M, Reinisch W, Dejaco C, Kratzer V, Gmeiner M, Miehsler W, Norman GL, Gangl A, Vogelsang H. OBJECTIVES: To examine the relationship between serum levels of anti-Saccharomyces cerevisiae antibodies (ASCAs) and intestinal permeability at a given time (hypothesis 1) and the probability of increased ASCA serum levels with increased intestinal permeability (hypothesis 2) in patients with Crohn's disease . METHODS: Each hypothesis was tested retrospectively with its own study population: group A for hypothesis 1 and group B for hypothesis 2 . Intestinal permeability was measured by lactulose/mannitol test and ASCAs were quantified by using ELISA . Patients received either no treatment or 5-aminosalicylates . The lactulose/mannitol test and sampling of sera for ASCA assessment had to be performed within 1 month in group A . In group B the highest intestinal permeability value obtained from among at least three measurements made during different stages of disease activity was chosen for evaluation . RESULTS: Both study populations consisted of 140 patients with Crohn's disease . Elevated IgG ASCAs were detected in 64% (90/140) in group A compared with 65% (91/140) in group B . In group A, 64% (90/140) and in group B 66% (92/140) were IgA ASCA positive . Correlation analysis showed a tendency for a positive relationship between IgG ASCAs and intestinal permeability in group A (tau = 0.16, P = 0.07) and in group B (tau = 0.16, P = 0.06) . A positive trend was seen for the combination of high intestinal permeability and high IgG ASCAs in group B (chi-squared test, P = 0.07) . CONCLUSION: Elevated serum levels of anti-S . cerevisiae antibodies do not seem to result primarily from a defect of the gut barrier . This observation points to an intrinsic pathomechanism in the development of increased ASCA serum levels. Eur J Pediatr Surg, 2003 Oct, 13(5), 319 - 23 Supplementation with Saccharomyces boulardii ameliorates hypoxia/reoxygenation-induced necrotizing enterocolitis in young mice; Akisu M et al.; Intestinal bacterial proliferation is an important aspect of gastrointestinal injury in neonatal necrotizing enterocolitis (NEC) . In the present investigation, we examined the protective action of oral supplementation with Saccharomyces boulardii (S . boulardii), non-pathogen probiotic yeast, against hypoxia-reoxygenation (H/R)-induced NEC in young mice . Young mice were divided into three groups; Group 1 mice (untreated) were subjected only to hypoxia-reoxygenation; Group 2 mice were subjected to hypoxia-reoxygenation and were then given lyophilized S . boulardii (10 mg suspended in 0.5 ml saline) twice a day by orogastric intubation for 10 days . Group 3 mice served as controls . Hypoxia was induced by placing young mice in a 100 % CO (2) chamber for 5 min . After hypoxia, the young mice were reoxygenated for 10 min with 100 % oxygen . We examined the intestinal lesions by light microscopy and measured intestinal generation of PAF and TNF-alpha in the H/R-induced model of NEC . In the probiotic group, NEC-induced intestinal tissue damage was greatly attenuated, with necrosis partially limited to the mucosa . Both intestinal tissue PAF and TNF-alpha concentrations were significantly higher in the untreated group than in controls (p < 0.001) . S . boulardii-supplemented young mice showed a significant decrease in intestinal generation of PAF compared with untreated young mice (p < 0.05) . On the other hand, no significant difference was observed in the intestinal concentration of TNF-alpha between untreated and probiotic groups ( p > 0.05) . The present study suggests that hypoxia/reoxygenation plays an important role in the pathogenesis of NEC and supports hypothesis that especially PAF and TNF-alpha are involved in the pathophysiological mechanism of H/R-induced NEC . This study also demonstrates that dietary supplementation with S . boulardii ameliorates the histologic evidence of H/R-induced intestinal injury . Based on these findings, the beneficial effects of probiotic S . boulardii in this model of NEC are mediated via mechanisms inhibiting intestinal proinflammatory mediator release. Am J Gastroenterol, 2003 Oct, 98(10), 2226 - 31 Anti-Saccharomyces cerevisiae antibodies: a stable marker for Crohn's disease during steroid and 5-aminosalicylic acid treatment; Teml A et al.; OBJECTIVES: An increased prevalence of elevated serum anti-Saccharomyces cerevisiae antibody (ASCA) levels in patients with Crohn's disease (CD) has been described . The aim of the present work was to investigate serum ASCA levels during the courses of prednisolone and mesalamine therapy in CD patients . METHODS: Serum samples of 25 patients with active CD were studied for ASCA levels before as well as 2 and 9 wk after initiation of a prednisolone tapering regimen . The influence of mesalamine (4 g o.d.) on serum ASCA levels compared to that of placebo was tested over 1 yr in 38 patients (20 mesalamine and 18 placebo) participating in a postoperative prophylaxis study . Serum IgG and IgA ASCA levels were measured by ELISA . Sera of 91 CD and 40 ulcerative colitis (UC) patients as well as 334 healthy donors were tested for ASCA to recalculate new cut-off values . RESULTS: For IgG ASCA cut-off values were determined to be 17.0 U and 25.0 U, and for IgA ASCA 9.3 U and 14.0 U . At baseline visit, 73.0% (46/63) of patients displayed serum ASCA positivity . During prednisolone therapy, a decrease in serum IgG and IgA ASCA levels from baseline to wk 2 (p < 0.0001 and p < 0.001, respectively) as well as to wk 9 (p < 0.001 and p = 0.01, respectively) was observed . A trend toward an association of ASCA positivity and steroid responsiveness was calculated (p = 0.07) . During mesalamine treatment, no differences in changes of ASCA levels were observed compared to placebo at any time point . CONCLUSIONS: ASCA are stable markers during steroid and mesalamine treatment, highlighting their reliability for use in diagnosis of CD. Inflamm Bowel Dis, 2003 Sep, 9(5), 302 - 7 Association of perinuclear antineutrophil cytoplasmic antibodies and anti-Saccharomyces cerevisiae antibodies with Vienna classification subtypes of Crohn's disease; Klebl FH et al.; BACKGROUND: The Vienna classification of Crohn's disease (CD) subdivides patients according to their age at diagnosis (A), disease location (L), and disease behavior (B).AIM The aim of this study was to test whether perinuclear antineutrophil cytoplasmic antibodies (pANCAs) or anti-Saccharomyces cerevisiae antibodies (ASCAs) correlate to subtypes of CD according to this classification . METHODS: pANCA, ASCA-immunoglobulin (Ig) A, and ASCA-IgG were detected by indirect immunofluorescence in 120 sera of patients with CD and compared with their Vienna classification . RESULTS: Patients with diagnosis of CD at an age of 40 years or older (A2) were more frequently pANCA-positive than those whose disease started at a younger age (A1) (19% vs . 5%, p < 0.05) . pANCA-positive patients almost exclusively belonged to the nonpenetrating disease groups (B1 and B2); only one patient had penetrating disease (B3) (B3 vs . non-B3: p = 0.02) . There was a cluster of pANCA-positive patients within the A1 B2 L3 subgroup (B2: stricturing disease; L3: ileocolonic involvement) . Only 7.5% of ASCA-IgA-positive patients presented with CD that was limited to the colon (L2) . ASCA-IgA and ASCA-IgG were more frequently found in patients with upper gastrointestinal disease (L4) (ASCA-IgA: 66.7% vs . 31.5% in non-L4, p < 0.05; ASCA-IgG: 44.4% vs . 27.9%, p < 0.05) . CONCLUSION: pANCA and ASCA may be useful in predicting subtypes of CD . They therefore may be helpful in developing subtype-specific disease management strategies. Prikl Biokhim Mikrobiol, 2003 Jul-Aug, 39(4), 438 - 40 {Level of vitamins B in yeasts of the Saccharomyces species depending on the composition of culture media}; Abramov ShA et al.; The qualitative and quantitative composition of water-soluble vitamins B in Saccharomyces yeasts cultivated on various nutrient media was studied by high-performance liquid chromatography . New strains of Saccharomyces oviformis Y-2635 and Saccharomyces vini F-5 yeasts grown in the nutrient medium with geothermal water differed in great biological value due to high intracellular concentrations of riboflavin, thiamine, nicotinic acid, and folic acid. J Biol Chem, 2003 Nov 28, 278(48), 47441 - 8 Epub 2003 Sep 18. The Hsp90 molecular chaperone complex regulates maltose induction and stability of the Saccharomyces MAL gene transcription activator Mal63p; Bali M et al.; Induction of the Saccharomyces MAL structural genes encoding maltose permease and maltase requires the MAL activator, a DNA-binding transcription activator . Genetic analysis of MAL activator mutations suggested that protein folding and stability play an important role in MAL activator regulation and led us to explore the role of the Hsp90 molecular chaperone complex in the regulation of the MAL activator . Strains carrying mutations in genes encoding components of the Hsp90 chaperone complex, hsc82 Delta hsp82-T101I and hsc82 Delta cpr7 Delta, are defective for maltase induction and exhibit significantly reduced growth rates on media containing a limiting concentration of maltose (0.05%) . This growth defect is suppressed by providing maltose in excess . Using epitope-tagged alleles of the MAL63 MAL activator, we showed that Mal63p levels are drastically reduced following depletion of cellular Hsp90 . Overexpression ( approximately 3-fold) of Mal63p in the hsc82 Delta hsp82-T101I and hsc82 Delta cpr7 Delta strains suppresses their Mal- growth phenotype, suggesting that Mal63p levels are limiting for maltose utilization in strains with abrogated Hsp90 activity . Consistent with this, the half-life of Mal63p is significantly shorter in the hsc82 Delta cpr7 Delta strain (reduced about 6-fold) and modestly affected in the Hsp90-ts strain (reduced about 2-fold) . Most importantly, triple hemagglutinin-tagged Mal63p protein is found in association with Hsp90 as demonstrated by co-immunoprecipitation . Taken together, these results identify the inducible MAL activator as a client protein of the Hsp90 molecular chaperone complex and point to a critical role for chaperone function in alternate carbon source utilization in Saccharomyces cerevisiae. Proc Natl Acad Sci U S A, 2003 Sep 16, 100(19), 10854 - 9 Epub 2003 Sep 08. Genetic regulation of telomere-telomere fusions in the yeast Saccharomyces cerevisae; Mieczkowski PA et al.; Yeast strains with mutations in both TEL1 and MEC1 have short telomeres and elevated rates of chromosome deletions . By using a PCR assay, we demonstrate that mec1 tel1 strains also have telomere-telomere fusions (T-TFs) . T-TFs require Lig4p (a ligase required for nonhomologous end-joining DNA repair) . The highest rates of T-TFs are found in strains with combination of mutations that affect telomere length and DNA damage checkpoints (mec1 tel1, mec3 tel1, mre11 mec1, and ddc1 tel1 strains) . Examining many mutant genotypes, we find good agreement between the level of T-TFs and the rate of chromosomal deletions . In addition, if telomeres are elongated in a mec1 tel1 strain, we eliminate T-TFs and reduce the deletion rate . The correlation between the level of T-TFs and the rate of deletions argues that many of these deletions reflect a cycle of T-TF formation (resulting in dicentric chromosomes), followed by chromosome breakage. Eur J Gastroenterol Hepatol, 2003 Jun, 15(6), 697 - 8 A pilot trial of Saccharomyces boulardii in ulcerative colitis; Guslandi M et al.; OBJECTIVES: Probiotics can be useful in the treatment of inflammatory bowel disease . In a previous report, the non-pathogenic yeast Saccharomyces boulardii was found to be beneficial in the maintenance treatment of Crohn's disease . The aim of this study was to assess the efficacy of S . boulardii in ulcerative colitis patients . METHODS: A group of 25 patients with a mild to moderate clinical flare-up of ulcerative colitis received additional treatment with S . boulardii 250 mg three times a day for 4 weeks during maintenance treatment with mesalazine . These patients were unsuitable for steroid therapy . Before and after treatment, Rachmilewitz's clinical activity index was calculated . The probiotic treatment was considered a therapeutic success only when the final score was lower than 6 . RESULTS: Of the 24 patients who completed the study, 17 attained clinical remission; this was confirmed endoscopically . CONCLUSIONS: Our preliminary results suggest that S . boulardii can be effective in the treatment of ulcerative colitis . Controlled studies with this probiotic agent are warranted. Acta Crystallogr D Biol Crystallogr, 2003 Jul, 59(Pt 7), 1267 - 9 Epub 2003 Jun 27. Crystallization and preliminary X-ray analysis of beta-alanine synthase from the yeast Saccharomyces kluyveri; Dobritzsch D et al.; In eukaryotes and some bacteria, the third step of reductive pyrimidine catabolism is catalyzed by beta-alanine synthase (EC 3.5.1.6) . Crystals of the recombinant enzyme from the yeast Saccharomyces kluyveri were obtained using sodium citrate as a precipitant . The crystals belong to space group P2(1) (unit-cell parameters a = 117.2, b = 77.1, c = 225.5 A, beta = 95.0 degrees ) and contain four homodimers per asymmetric unit . Data were collected to 2.7 A resolution . Introduction of heavy atoms into the crystal lattice induced a different set of unit-cell parameters (a = 61.0, b = 77.9, c = 110.1 A, beta = 97.2 degrees ) in the same space group P2(1), with only one homodimer per asymmetric unit. Acta Gastroenterol Belg, 2003 Jan-Mar, 66(1), 1 - 6 Negative association between smoking and anti-saccharomyces cerevisiae antibodies in Crohn's disease; Van Kemseke C et al.; BACKGROUND: Crohn's disease (CD) is a polygenic multifactorial heterogeneous disease . Anti-Saccharomyces Cerevisiae antibodies (ASCA) correlate highly with CD and are present in 50-80% of patients . The reason for ASCA positivity or negativity in CD is unknown . The aim of our work was to analyse clinical, epidemiological and genetic characteristics in ASCA+ or ASCA- CD patients . METHODS: 113 patients with CD were tested for ASCA (IgA and IgG) by using a commercial kit (Medipan Diagnostica) . Age, gender, systemic manifestations, familial form of disease, age at diagnosis, location and behaviour of the disease, smoking habit as well as genotyping for -308 TNF gene polymorphisms were determined . RESULTS: 38.9% CD patients were negative for both IgA and IgG ASCA while 61.1% were ASCA positive (respectively IgA and IgG: 31.9%; IgA only: 9.7%; IgG only: 19.5%) . The only significant difference between ASCA+ and ASCA- patients was for smoking habit: there were 29% smokers in ASCA+ versus 50% in ASCA- CD patients (P = 0.03) . This low proportion of smokers was more prominent in ASCA IgA+ patients than in isolated ASCA IgG+ patients (25.6% versus 45.5%) and was minimal in patients with high titers of ASCA IgA (0/8) . Logistic regression showed smoking habit still borderline for significance (P = 0.057) . CONCLUSIONS: Our results suggest a negative association between smoking and ASCA positivity in CD . This association was more prominent for ASCA IgA+ . It indicates that smoking habit should be taken into account when analysing ASCA status in CD patients and may suggest an influence of smoking on immunization against intestinal material. FEMS Microbiol Lett, 2003 Jun 6, 223(1), 141 - 5 Severe reduction of superoxide dismutase activity in the yeast Saccharomyces cerevisae with the deletion or overexpression of GTS1; Abudugupur A et al.; We report herein that the level of reactive oxygen species (ROS) observed using dihydrorhodamine is much higher in either GTS1-deleted (gts1Delta) or GTS1-overexpressing (TMpGTS1) transformants than in the wild-type and that the levels of protein carbonyls are increased and the glutathione levels are decreased in both transformants . Consistently, the activities of superoxide dismutases (SODs) in both gts1Delta and TMpGTS1 were severely weakened, while the protein levels of both Cu/Zn-SOD and Mn-SOD were not so changed . As the intracellular copper levels were significantly increased in both transformants, we hypothesized that, in either gts1Delta or TMpGTS1 cells, the imbalanced homeostasis of copper induced an accumulation of ROS which caused inactivation of SODs further increasing ROS levels. Antonie Van Leeuwenhoek, 2003, 83(2), 155 - 66 Differentiation of six sibling species in the Saccharomyces sensu stricto complex by multilocus enzyme electrophoresis and UP-PCR analysis; Naumova ES et al.; UP-PCR analysis and multilocus enzyme electrophoresis were used to characterize 37 strains of the sibling species Saccharomyces cerevisiae, S . bayanus, S . cariocanus, S . kudriavzevii, S . mikatae and S . paradoxus . The results demonstrate that both molecular approaches are useful for discriminating between these phenotypically indistinguishable Saccharomyces species . The data obtained are in excellent agreement with previously reported genetic analyses, sequencing of the 18S rRNA and ITS regions, and DNA-DNA reassociation data. Science, 2003 Jul 4, 301(5629), 71 - 6 Epub 2003 May 29. Finding functional features in Saccharomyces genomes by phylogenetic footprinting; Cliften P et al.; The sifting and winnowing of DNA sequence that occur during evolution cause nonfunctional sequences to diverge, leaving phylogenetic footprints of functional sequence elements in comparisons of genome sequences . We searched for such footprints among the genome sequences of six Saccharomyces species and identified potentially functional sequences . Comparison of these sequences allowed us to revise the catalog of yeast genes and identify sequence motifs that may be targets of transcriptional regulatory proteins . Some of these conserved sequence motifs reside upstream of genes with similar functional annotations or similar expression patterns or those bound by the same transcription factor and are thus good candidates for functional regulatory sequences. FEMS Yeast Res, 2003 Jun, 3(4), 363 - 73 High-rate evolution of Saccharomyces sensu lato chromosomes; Spirek M et al.; Forty isolates belonging to the Saccharomyces sensu lato complex were analyzed for one nuclear and two mitochondrial sequences, and for their karyotypes . These data are useful for description and definition of yeast species based on the phylogenetic species concept . The deduced phylogenetic relationships among isolates based on the nuclear and mitochondrial sequences were usually similar, suggesting that horizontal transfer/introgression has not been frequent . The highest degree of polymorphism was observed at the chromosome level . Even isolates which had identical nuclear and mitochondrial sequences often exhibited variation in the number and size of their chromosomes . Apparently, yeast chromosomes have been frequently reshaped and therefore also the position of genes has been dynamic during the evolutionary history of yeasts. Dig Dis Sci, 2003 Apr, 48(4), 770 - 4 Dose-response relationship and mechanism of action of Saccharomyces boulardii in castor oil-induced diarrhea in rats; Girard P et al.; For biotherapeutic agents, there is a lack of information on dose-response relationships and mechanism of action . The present study was designed to address these issues for Saccharomyces boulardii using the rat model of castor oil-induced diarrhea . A single dose of Saccharomyces boulardii at 12 x 10(10) CFU/kg of viable cells given 1 hr before castor oil administration significantly reduced the onset of diarrhea . Repeated ingestion of the yeast, twice daily between 1.2 and 12 x 10(10) CFU/kg for 5 days before castor oil, showed a dose-response relationship . The percentage of rats with diarrhea decreased and a stronger protection was afforded by the repeated treatment . The mechanism of action of Saccharomyces boulardii in this model was investigated with two classes of antagonists, naloxone and L-arginine . The effect of Saccharomyces boulardii was not inhibited by naloxone but was significantly reduced by L-arginine . This last result suggests a novel mechanism of action for Saccharomyces boulardii involving a possible inhibition of nitric oxide production by the yeast. FEMS Yeast Res, 2002 Jan, 1(4), 323 - 31 Evidence for multiple interspecific hybridization in Saccharomyces sensu stricto species; de Barros Lopes M et al.; Fluorescent amplified fragment length polymorphism analysis demonstrates a high level of gene exchange between Saccharomyces sensu stricto species, with some strains having undergone multiple interspecific hybridization events with subsequent changes in genome complexity . Two lager strains were shown to be hybrids between Saccharomyces cerevisiae and the alloploid species Saccharomyces pastorianus . The genome structure of CBS 380(T), the type strain of Saccharomyces bayanus, is also consistent with S . pastorianus gene transfer . The results indicate that the cider yeast, CID1, possesses nuclear DNA from three separate species . Mating experiments show that there are no barriers to interspecific conjugation of haploid cells . Furthermore, the allopolyploid strains were able to undergo further hybridizations with other Saccharomyces sensu stricto yeasts . These results demonstrate that introgression between the Saccharomyces sensu stricto species is likely. FEMS Yeast Res, 2002 Dec, 2(4), 471 - 9 Concealed nuclei in Saccharomyces strains; Nevzglyadova OV et al.; We have found that cells derived from heterokaryons (HK) showing phenotypical traits, coded by the nucleus of one parental strain and by the cytoplasm of the other, may produce mitotic progeny in which the second nucleus is apparently present but not expressed . This 'concealed' nucleus can be forced to expression after growth of these cytoductants on proper selective media . Using a micromanipulator, the buds containing both parental nuclei were isolated in various crosses . Cloning these HK from a rich medium (YEPD) revealed that nearly all of them were composed of a mixture of hybrid cells and cells of one of the parents . Cells of the other parent were present in a very small proportion, if detectable at all . We showed that the percentage of concealed HK decreases when limiting the growth of the strain that serves as a donor of the concealed nucleus . Consequently, our explanation for the presence of concealed nuclei in HK is the low production rate of daughter cells containing both nuclei, which accounts for the lack of a visible phenotype in HK, together with the low replication rate (or fast nuclease degradation) of one of the nuclei . In homosexual crosses, selection allows us to switch the concealed nucleus to normal replication rate . Some abnormalities of meiosis due to hidden nuclei are shown. Ann Rheum Dis, 2003 May, 62(5), 455 - 9 Anti-saccharomyces cerevisiae IgA antibodies are raised in ankylosing spondylitis and undifferentiated spondyloarthropathy; Hoffman IE et al.; OBJECTIVES: To investigate whether anti-Saccharomyces cerevisiae antibodies (ASCA), a marker for Crohn's disease (CD), are present in spondyloarthropathies (SpA) and in the subgroups ankylosing spondylitis (AS), undifferentiated SpA (uSpA), and psoriatic arthritis (PsA), in comparison with healthy and inflammatory controls (patients with rheumatoid arthritis (RA)) . METHODS: ASCA IgA and IgG levels were measured with an enzyme linked immunosorbent assay (ELISA) kit (Medipan, Germany) in 26 patients with CD, 108 patients with SpA (43 patients with AS, 20 patients with uSpA, 45 patients with PsA), 56 patients with RA and 45 healthy controls . Gut biopsy samples were available in 18 AS and 10 patients with uSpA, these samples were screened for the presence of inflammation . RESULTS: Both ASCA IgG and IgA levels were raised in CD compared with healthy controls and patients with RA . ASCA IgA, but not IgG levels, were higher in SpA than in both healthy and RA controls . ASCA IgA levels were raised in AS and uSpA, but not in PsA . No significant differences in ASCA IgA levels were noted between patients with SpA with and without histological gut inflammation . CONCLUSION: ASCA IgA levels are significantly higher in SpA, and more specifically in AS, than in healthy controls and patients with RA . This is the first serum marker associated with SpA . No correlation between the presence of subclinical bowel inflammation and ASCA IgA levels was noted . However, it remains to be evaluated whether patients with SpA with ASCA have an increased risk of developing CD. J Gastroenterol, 2003, 38(2), 121 - 6 Anti-Saccharomyces cerevisiae antibodies in Japanese patients with inflammatory bowel disease: diagnostic accuracy and clinical value; Hisabe T et al.; BACKGROUND: The diagnostic accuracy of the determination of anti- Saccharomyces cerevisiae antibodies (ASCA) and its clinical significance in Crohn's disease (CD) have been reported in Western countries, but scarcely reported in Japan, where the dietary habits of people differ markedly from those of Western countries . Th present study was undertaken to examine the diagnostic accuracy and clinical significance of ASCA determination in Japanese patients with CD or ulcerative colitis (UC) . METHODS: Seventy-five serum samples obtained from 68 patients with CD, 34 serum samples obtained from 30 patients with UC, 35 serum samples from 35 patients with liver cirrhosis, and 31 serum samples from 31 healthy controls were examined . The optical density of each sample was measured by an enzyme-linked immunosorbent assay (ELISA) method to quantify ASCA IgA and IgG . RESULTS: The cutoff level, as determined by discriminant analysis of the data, was 0.1502 for ASCA IgA and 0.156 for ASCA IgG . Of the 68 patients with CD, 31 (45.6%) were ASCA-positive, and of the 30 patients with UC, 4 (13.3%) were ASCA-positive according to these cutoff levels . The sensitivity and specificity of ASCA determination for the differential diagnosis of CD from UC were 45.6% and 86.7%, respectively . When the relationship between ASCA and the clinical features of CD was analyzed, ASCA positivity was found to be correlated with duration of illness . CONCLUSIONS: The specificity of ASCA was high; however, the sensitivity was not . ASCA titers were generally low in the Japanese population examined . It would be desirable to determine cutoff levels for ASCA tailored to the Japanese people for the diagnosis of inflammatory bowel disease. Infect Immun, 2003 Feb, 71(2), 766 - 73 Saccharomyces boulardii interferes with enterohemorrhagic Escherichia coli-induced signaling pathways in T84 cells; Dahan S et al.; Enterohemorrhagic Escherichia coli (EHEC) infections are associated with the modification of tight-junction permeability and synthesis of proinflammatory cytokine interleukin-8 (IL-8) . In a previous study, it was demonstrated that EHEC-induced IL-8 secretion is due to the involvement of the mitogen-activated protein kinase (MAPK), AP-1, and NF-kappaB pathways . In this study, we investigated the effect of the yeast Saccharomyces boulardii on EHEC infection in T84 cells . For this purpose, cells were (i) incubated with bacteria and yeast at the same time or (ii) incubated overnight with yeast cells that were maintained during infection or eliminated by several washes before infection . Coincubation is sufficient to maintain the transmonolayer electrical resistance (TER) of EHEC-infected cells, whereas the preincubation of cells with the yeast without elimination of the yeast during infection is necessary to significantly decrease IL-8 secretion . We thus analyzed the mechanisms of S . boulardii action . We showed that S . boulardii has no effect on EHEC growth or on EHEC adhesion . Kinetics studies revealed that EHEC-induced myosin light chain (MLC) phosphorylation precedes the decrease of TER . ML-7, an MLC kinase inhibitor, abolishes the EHEC-induced MLC phosphorylation and decrease of TER . Studies show that S . boulardii also abolishes EHEC-induced MLC phosphorylation . We demonstrated that the preincubation of cells with S . boulardii without washes before EHEC infection inhibits NF-kappaB DNA binding activity, phosphorylation and degradation of IkappaB-alpha, and activation of the three members of a MAPK group (extracellular signal-regulated protein kinases 1 and 2, p38, and c-jun N-terminal kinase) . These findings demonstrate that S . boulardii exerts a preventive effect on EHEC infection by (i) interfering with one of the transduction pathways implicated in the control of tight-junction structure and (ii) decreasing IL-8 proinflammatory secretion via inhibition of the NF-kappaB and MAPK signaling pathways in infected T84 cells. Nucleic Acids Res, 2003 Jan 1, 31(1), 216 - 8 Saccharomyces Genome Database (SGD) provides biochemical and structural information for budding yeast proteins; Weng S et al.; The Saccharomyces Genome Database (SGD: has recently developed new resources to provide more complete information about proteins from the budding yeast Saccharomyces cerevisiae . The PDB Homologs page provides structural information from the Protein Data Bank (PDB) about yeast proteins and/or their homologs . SGD has also created a resource that utilizes the eMOTIF database for motif information about a given protein . A third new resource is the Protein Information page, which contains protein physical and chemical properties, such as molecular weight and hydropathicity scores, predicted from the translated ORF sequence. Tohoku J Exp Med, 2002 Sep, 198(1), 1 - 9 Effects of octreotide acetate and Saccharomyces boulardii on bacterial translocation in an experimental intestinal loop obstruction model of rats; Aldemir M et al.; Intestinal obstruction (IO) induces bacterial translocation (BT) due to mucosal disruption, motility dysfunction, and increased intestinal volume, leading to bacterial overgrowth . This study was conducted to investigate the effects of octreotide acetate (OA) and Saccharomyces boulardii (SB) on the BT and intestinal integrity in an animal model of intestinal loop obstruction (LO) . Forty adult male Sprague-Dawley rats (250-300 g) were randomized into 4 groups containing 10 rats each . Complete IO was created in the distal ileum of rats by a single 3-0 silk suture (LO) . Group Sham: Sham (Laparotomy only was performed in this group); group LO: LO; group OA: LO plus OA (100 microg/kg, at 0, 12 hours of obstruction); group (SB): LO plus SB (800 mg/kg/day, via orogastric and preoperative for 3 days) . After 24 hours, samples of mesenteric lymph nodes (MLN), liver, spleen and blood were obtained and cultured . The terminal ileum specimens were examined histopathologically . There were no BT in group Sham, but BT was noticed totally in 31 (77.5%) cultures in group LO . This rate was reduced to 30% (n = 12), 10% (n = 4) in the groups OA and SB respectively . Bacterial translocations of MLN and the liver in group LO were significantly higher than those of groups OA and SB . Bacterial translocations of the both spleen and blood in group LO were significantly higher than those of groups OA and SB . The mean bacterial counts, colony-forming units per gram tissue (cfu/g), in the MLN, liver and spleen of group LO were found significantly higher than those of groups OA and SB . The mean villus height in group OA was significantly higher than that of group LO and it in the group SB significantly higher than those of groups LO and OA . The present experimental study has demonstrated that OA and SB may have protective effects against BT in mechanical bowel obstruction and additionally SB preserves intestinal mucosal integrity. Proc Natl Acad Sci U S A, 2002 Dec 24, 99(26), 16893 - 8 Epub 2002 Dec 13. A systematic approach to reconstructing transcription networks in Saccharomycescerevisiae; Wang W et al.; Decomposing regulatory networks into functional modules is a first step toward deciphering the logical structure of complex networks . We propose a systematic approach to reconstructing transcription modules (defined by a transcription factor and its target genes) and identifying conditionsperturbations under which a particular transcription module is activateddeactivated . Our approach integrates information from regulatory sequences, genome-wide mRNA expression data, and functional annotation . We systematically analyzed gene expression profiling experiments in which the yeast cell was subjected to various environmental or genetic perturbations . We were able to construct transcription modules with high specificity and sensitivity for many transcription factors, and predict the activation of these modules under anticipated as well as unexpected conditions . These findings generate testable hypotheses when combined with existing knowledge on signaling pathways and protein-protein interactions . Correlating the activation of a module to a specific perturbation predicts links in the cell's regulatory networks, and examining coactivated modules suggests specific instances of crosstalk between regulatory pathways. Digestion, 2002, 66(3), 173 - 7 Anti-saccharomyces cerevisiae antibodies in patients with inflammatory bowel disease and their first-degree relatives: potential clinical value; Glas J et al.; Anti-Saccharomyces cerevisiae antibodies (ASCA) have been described as specific markers in Crohn's disease and their healthy first-degree relatives . 171 patients with Crohn's disease, their 105 first-degree relatives, 145 patients with ulcerative colitis and 101 first-degree relatives of patients with ulcerative colitis, 50 patients with infectious enterocolitis and 100 healthy controls were tested for ASCA employing the ELISA technique . When compared with the healthy controls (p < 0.0001) and patients with infectious enterocolitis (p < 0.0001) the prevalence of ASCA was significantly increased in patients with Crohn's disease and their first-degree relatives (p < 0.01) . Further significant differences concerning the frequency of ASCA within the different groups of our study population were not observed . In particular, ASCA were not found in increased prevalence in infectious enterocolitis . These observations are compatible with a role of ASCA as a marker of genetic predisposition to Crohn's disease . Acta Crystallogr D Biol Crystallogr, 2002 Dec, 58(Pt 12), 2135 - 7 Epub 2002 Nov 23. Purification, crystallization and preliminary X-ray diffraction analysis of the carbohydrate-binding domain of flocculin, a cell-adhesion molecule from Saccharomyces carlsbergensis; Groes M et al.; The recombinant carbohydrate-binding domain of the cell-surface lectin flocculin from brewer's yeast has been identified, purified and crystallized . The expression of the protein is associated with the nutritional state of the yeast . P2(1)2(1)2(1) crystals with unit-cell parameters a = 36.5, b = 59.7, c = 83.1 A were obtained in hanging drops at 295 K using 25%(w/v) PEG 4000, 0.05 M KH(2)PO(4) as precipitant . X-ray diffraction data have been obtained to 2.6 A . The asymmetric unit contains one molecule and has a solvent content of 32% . An isomorphous PtCl(4)(2-) derivative has been obtained. Genetika, 2002 Oct, 38(10), 1428 - 33 {The SRM12/ADA1 gene acts as a cis-active factor when inserted into recombinant plasmids and makes them more stable in Saccharomyces}; Chekhuta IA et al.; A DNA fragment containing the SRM12/ADA1 gene sequence inserted into a recombinant circular plasmid improves its maintenance in budding yeast (Saccharomyces cerevisiae) cells . Plasmid stabilization caused by the integrated SRM12 sequence does not require the SRM12 function complementing the srm12 mutation and depends on the orientation of the inserted fragment in the vector . This stabilization is mainly due to a decrease in spontaneous plasmid underreplication/copy loss rather than an increase in the fidelity of mitotic plasmid segregation. Proc Natl Acad Sci U S A, 2002 Nov 26, 99(24), 15530 - 5 Epub 2002 Nov 13. Parallel phenotypic analysis of sporulation and postgermination growth in Saccharomycescerevisiae; Deutschbauer AM et al.; We have quantitatively monitored the sporulation and germination efficiencies of approximately 4,200 yeast deletion strains in parallel by using a molecular bar coding strategy . In a single study, we doubled the number of genes functionally implicated in sporulation to approximately 400, identifying both positive and negative regulators . Our set of 261 sporulation-deficient genes illustrates the importance of autophagy, carbon utilization, and transcriptional machinery during sporulation . These general cellular factors are more likely to exhibit fitness defects when deleted and less likely to be transcriptionally regulated than sporulation-specific genes . Our postgermination screening assay identified recombinationchromosome segregation genes, aneuploid strains, and possible germination-specific factors . Finally, our results facilitate a genome-wide comparison of expression pattern and mutant phenotype for a developmental process and suggest that 16% of genes differentially expressed during sporulation confer altered efficiency of spore production or defective postgermination growth when disrupted. J Clin Immunol, 2002 Sep, 22(5), 281 - 8 Diagnostic value of anti-Saccharomyces cerevisiae and antineutrophil cytoplasmic antibodies for inflammatory bowel disease: high prevalence in patients with celiac disease; Damoiseaux JG et al.; Both celiac disease and inflammatory bowel disease (IBD) are characterized by chronic diarrhea and the presence of distinct (auto)antibodies . In the present study we wanted to determine the prevalence of serological markers for inflammatory bowel disease, i.e., perinuclear antineutrophil cytoplasmic antibodies (pANCA) and/or anti-Saccharomyces cerevisiae antibodies (ASCA), in 37 patients with biopsy-confirmed celiac disease (Marsh IIIb/c) . The majority of the patients was positive for IgA (auto)antibodies typically associated with celiac disease, i.e., antiendomysium antibodies (EMA) (86.5%), antigliadin antibodies (AGA) (73%), and antirecombinant human tissue transglutaminase antibodies (rh-tTGA) (86.5%) . Four patients with selective IgA deficiency could be identified by analyzing EMA, AGA, and rh-tTGA for the IgG isotype . The prevalence of pANCA and ASCA, markers that are used for IBD, was unexpectedly high in our cohort of patients with celiac disease: 8 patients were positive for pANCA (IgG) and 16 patients were positive for ASCA (IgG and/or IgA) . These results indicate that the presence of pANCA or ASCA in the serum of patients with chronic diarrhea does not exclude celiac disease . A prospective study is required to determine whether pANCA and/or ASCA identify patients at risk for developing secondary autoimmune disease. Clin Exp Rheumatol, 2002 Jul-Aug, 20(4 Suppl 26), S21 - 4 Anti-Saccharomyces cerevisiae antibodies--a novel serologic marker for Behçet's disease; Krause I et al.; OBJECTIVE: To evaluate the prevalence and clinical correlations of antibodies against Saccharomyces cerevisiae (ASCA) among patients with BD . METHODS: Twenty-seven BD patients were studied . Data from medical files and from patients' interviews was collected, regarding the entire spectrum of disease manifestations, and a severity score was calculated for each patient . IgA- and IgG-ASCA levels, determined by ELISA, were studied in all BD patients and in three control groups: patients with recurrent aphthous stomatitis (RAS), systemic lupus erythematosus (SLE) and healthy volunteers . RESULTS: Thirteen BD patients (48.1%) were ASCA-positive, compared to one patient in each control group (10%, p = 0.01) . The mean value of IgG-ASCA in the BD patients was 20.7 +/- 12.3 units, significantly higher than in patients with RAS (10.0 +/- 5.5, p < 0.001), SLE (11.8 +/- 9.3, p < 0.03) or healthy volunteers (10.8 +/- 9.8, p < 0.02) . Mean IgA-ASCA level was 16.8 +/- 8.8 units in the BD patients, significantly higher compared to healthy volunteers (11.0 +/- 5.0, p = 0.02) but similar to patients with RAS (17.0 +/- 5.3) . No correlation was found between ASCA and any BD-associated clinical manifestation nor the presence of HLA-B5 . No difference was found in the rate of major oral ulcers nor in the systemic disease severity score between positive- and negative-ASCA patients (27.3% vs . 30.8%, and 7.31 +/- 1.80 vs . 7.28 +/- 2.27 respectively, NS) . CONCLUSION: The results of our study associate, for the first time, the presence of a distinct antibody, i.e . ASCA, with BD . ASCA were not linked to a specific clinical manifestation of the disease and probably do not pose an increased risk for a more severe disease course. Appl Microbiol Biotechnol, 2002 Sep, 59(6), 727 - 30 Epub 2002 Jul 20. Saccharomyces bayanus var . uvarum in Tokaj wine-making of Slovakia and Hungary; Naumov GI et al.; Using genetic hybridisation analysis and molecular karyotyping we revealed an association of Saccharomyces bayanus var . uvarum species with Tokaj wine-making . Along with identification of Saccharomyces strains isolated by E . Minarik in Slovakia, the composition of Tokaj populations in Hungary was studied . Twenty-eight Hungarian Saccharomyces strains were analysed in terms of karyotype . The majority of strains belong to S . bayanus var . uvarum . Two non-identified Saccharomyces strains were found to be polyploid according to their complex karyotype patterns. Dis Colon Rectum, 2002 Aug, 45(8), 1062 - 9 Diagnostic role of anti-Saccharomyces cerevisiae mannan antibodies combined with antineutrophil cytoplasmic antibodies in patients with inflammatory bowel disease; Kim BG et al.; PURPOSE: Perinuclear antineutrophil cytoplasmic autoantibody is known to be a marker for ulcerative colitis, and anti-Saccharomyces cerevisiae Mannan antibody is a serologic marker associated with Crohn's disease . The aim of this study was to assess the value of detecting perinuclear antineutrophil cytoplasmic autoantibody and/or anti-Saccharomyces cerevisiae Mannan antibody for the diagnosis of ulcerative colitis, Crohn's disease, Behcet's colitis, and tuberculous colitis . METHODS: Serum samples were obtained from 85 patients with Crohn's disease, 77 with ulcerative colitis, 36 with Behcet's colitis, 14 with tuberculous colitis, 20 healthy controls, and 21 first-degree relatives of patients with Crohn's disease . Determination of perinuclear antineutrophil cytoplasmic autoantibody and anti-Saccharomyces cerevisiae Mannan antibody was performed with the standardized indirect immunofluorescence technique and an enzyme-linked immunosorbent assay, respectively . RESULTS: A relatively high percentage of patients with Crohn's disease (49.4 percent), relatives of Crohn's disease patients (61.9 percent), and patients with Behcet's disease (41.7 percent) tested seropositive for anti-Saccharomyces cerevisiae Mannan antibody compared with normal controls (10 percent) . In cases of ulcerative colitis, 44.2 percent tested seropositive for perinuclear antineutrophil cytoplasmic autoantibody, whereas the controls showed 0 percent positivity . The combination of a positive anti-Saccharomyces cerevisiae Mannan antibody test and a negative perinuclear antineutrophil cytoplasmic autoantibody yielded a sensitivity and specificity of 48.2 and 87 percent, respectively, for Crohn's disease . The combination of a positive perinuclear antineutrophil cytoplasmic autoantibody test and a negative anti-Saccharomyces cerevisiae Mannan antibody test yielded a sensitivity and specificity of 36.4 and 97.6 percent, respectively, for ulcerative colitis . CONCLUSION: Anti-Saccharomyces cerevisiae Mannan antibody may be associated with Crohn's disease and Behcet's disease and perinuclear antineutrophil cytoplasmic autoantibody with ulcerative colitis . A combination of both tests may aid the differential diagnosis of inflammatory bowel disease. Comput Chem, 2002 Jul, 26(5), 437 - 45 Genome compartimentation by a hybrid chromosome model (HXM) . Application to Saccharomyces cerevisae subtelomeres; de Brevern AG et al.; The aim of this paper is to present a new approach, called 'Hybrid Chromosome Model' (HXM), which allows both the extraction of regions of similarity between two sequences, and the compartimentation of a set of DNA sequences . The principle of the method consists in compacting a set of sequences (split into fragments of fixed length) into a 'hybrid chromosome', which results from the stacking of the whole sequence fragments . We have illustrated our approach on the 32 subtelomeres of Saccharomyces cerevisae . The compartimentation of these chromosome extremities into common regions of similarity has been carried out . The approach HXM is a fast and efficient tool for mapping entire genomes and for extracting ancient duplications within or between genomes. Intensive Care Med, 2002 Jun, 28(6), 797 - 801 Epub 2002 May 10. Seven cases of fungemia with Saccharomyces boulardii in critically ill patients; Lherm T et al.; Saccharomyces boulardii (Sb) is a particular strain of Saccharomyces cerevisiae (Sc) . This viable yeast is used in intensive care adult patients, delivered in packets of 500 mg, for preventing diarrhea associated with antibiotics or enteral feeding at a regimen of 1-2 g/day . Between June 1996 and October 1998, seven cases of fungemia with Sb occurred in a 12-bed intensive care unit (ICU) . All the patients concerned were severely ill patients, mechanically ventilated, treated by broad spectrum antibiotics with central venous catheter and were pretreated with Sb, except for one patient . In this study, Sb was identified by specific mycologic methods and confirmed the genomic identity between isolates of blood culture and yeasts from the treatment packets, contrary to a few other reports concerning Saccharomyces species published in international literature . The hypothesis discussed for explaining these cases of Sb fungemia are: (1) an intestinal translocation of Sb administered at a high dosage in severely ill patients, (2) a contamination of the central venous catheter, especially in the patient not pretreated with Sb and (3) a massive colonization of critically ill patients by the yeast as has been reported for Candida species . We note that cases of fungemia with Sc and Sb have become more and more frequent in the international literature during the last 10 years and we do not recommend administering Sb treatment in critically ill patients. Microbes Infect, 2002 Jun, 4(7), 733 - 9 Experimental effects of Saccharomyces boulardii on diarrheal pathogens; Czerucka D et al.; Saccharomyces boulardii is a selected strain of yeast that may have applications in the prevention and treatment of intestinal infections . The animal models and in vitro studies developed to elucidate the mechanisms of this protection are reviewed and discussed. Proc R Soc Lond B Biol Sci, 2002 Jun 7, 269(1496), 1167 - 71 Epistasis and hybrid sterility in Saccharomyces; Greig D et al.; Hybrid sterility is thought to be due to deleterious epistatic interactions between genes from different species . Here we demonstrate that dominant genic incompatibility does not contribute to sterility in hybrids between Saccharomyces cerevisiae and five closely related species . Sterile diploids were made fertile by genome doubling to produce hybrid tetraploids . Based on these and previous results, we conclude that neither genic incompatibility nor classical chromosomal speciation models apply. Genes Dev, 2002 Jun 1, 16(11), 1383 - 96 Saccharomyces Rrm3p, a 5' to 3' DNA helicase that promotes replication fork progression through telomeric and subtelomeric DNA; Ivessa AS et al.; In wild-type Saccharomyces cerevisiae, replication forks slowed during their passage through telomeric C(1-3)A/TG(1-3) tracts . This slowing was greatly exacerbated in the absence of RRM3, shown here to encode a 5' to 3' DNA helicase . Rrm3p-dependent fork progression was seen at a modified Chromosome VII-L telomere, at the natural X-bearing Chromosome III-L telomere, and at Y'-bearing telomeres . Loss of Rrm3p also resulted in replication fork pausing at specific sites in subtelomeric DNA, such as at inactive replication origins, and at internal tracts of C(1-3)A/TG(1-3) DNA . The ATPase/helicase activity of Rrm3p was required for its role in telomeric and subtelomeric DNA replication . Because Rrm3p was telomere-associated in vivo, it likely has a direct role in telomere replication. EMBO J, 2002 May 1, 21(9), 2282 - 91 Directional bias during mating type switching in Saccharomyces is independent of chromosomal architecture; Simon P et al.; Haploid Saccharomyces cells have the remarkable potential to change mating type as often as every generation, a process accomplished by an intrachromosomal gene conversion between an expressor locus MAT and one of two repositories of mating type information, HML or HMR . The particular locus selected as donor is dictated by the mating type of the cell, a bias that ensures productive mating type interconversion . Here we use green fluorescent protein tagging of the expressor and donor loci on chromosome III to show that this preference for donor locus does not result from a predetermined organization of chromosome III: HML and MAT as well as HMR and MAT remain separated in cells of both mating types . In fact, cells in which the inappropriate donor locus is artificially tethered to MAT still predominantly select the correct donor . We find, though, that initiation of switching leads to a rapid association of the correct donor locus with MAT . Thus, in mating type switching in Saccharomyces, donor preference is imposed at commitment to recombination rather than at physical contact of interacting DNA strands. Proc Natl Acad Sci U S A, 2002 Apr 16, 99(8), 5412 - 7 A Los1p-independent pathway for nuclear export of intronless tRNAs in Saccharomycescerevisiae; Feng W et al.; Los1p, the Saccharomyces cerevisiae exportin-t homologue, binds tRNA and functions in pre-tRNA splicing and export of mature tRNA from the nucleus to the cytosol . Because LOS1 is unessential in yeast, other pathways for tRNA nuclear export must exist . We report that Cca1p, which adds nucleotides C, C, and A to the 3' end of tRNAs, is a multicopy suppressor of the defect in tRNA nuclear export caused by los1 null mutations . Mes1p, methionyl-tRNA synthetase, also suppresses the defect in nuclear export of tRNA(Met) in los1 cells . Thus, Cca1p and Mes1p seem to function in a Los1p-independent tRNA nuclear export pathway . Heterokaryon analysis indicates that Cca1p is a nucleus/cytosol-shuttling protein, providing the potential for Cca1p to function as an exporter or an adapter in this tRNA nuclear export pathway . In yeast, most mutations that affect tRNA nuclear export also cause defects in pre-tRNA splicing leading to tight coupling of the splicing and export processes . In contrast, we show that overexpressed Cca1p corrects the nuclear export, but not the pre-tRNA-splicing defects of los1Kan(r) cells, thereby uncoupling pre-tRNA splicing and tRNA nuclear export. Genes Genet Syst, 2001 Dec, 76(6), 393 - 410 Defects in glycosylphosphatidylinositol (GPI) anchor synthesis activate Hog1 kinase and confer copper-resistance in Saccharomyces cerevisisae; Toh-e A et al.; Las21/Gpi7 contains a heavy-metal-associated motif at its N-terminus . When this motif was disrupted by amino acid substitution, the cells acquired weak copper-resistance . We found that the previously isolated las21 mutants were strongly resistant to copper . Metallothionein is necessary for the expression of the copper-resistance of the las21 mutants . However, hyper-production of metallothionein is unlikely to be the cause of copper-resistance of the las21 mutants . Copper-sensitive mutants (collectively called Cus mutants) were isolated from the las21delta and characterized . One of the Cus genes was found to be PBS2, which encodes Hog1 MAP kinase kinase, indicating that the Hog1 MAP kinase pathway is needed for the expression of copper-resistance of the las21 mutants . As expected, the las21delta hog1delta strain was no longer copper-resistant . We found that Hog1 was constitutively activated in las21delta cells and in ssk1delta las21delta cells but not in sho1delta las21delta cells . Inactivation of either FSR2/MCD4 or MPC1/GPI13, both of which are involved in GPI anchor synthesis, like LAS21, caused a similar level of constitutive activation of Hog1 kinase and copper-resistance as found in the las21delta strain . The constitutive activation was canceled by introducing the sskl mutation, but not the sho1 mutation, in each GPI anchor mutant tested, suggesting that the defect in GPI anchor synthesis specifically affects the Slnl branch of the MAP kinase pathway . Since the wild-type cells grown in YPD containing 0.5 M NaCl do not show copper-resistance, mere activation of Hog1 is not sufficient for expression of copper-resistance . We propose that a defect in GPI anchor synthesis has multiple consequences, including activation of the Hog1 MAP kinase cascade and conferring copper-resistance. Pediatr Res, 2002 Apr, 51(4), 528 - 34 Saccharomyces boulardii enhances N-terminal peptide hydrolysis in suckling rat small intestine by endoluminal release of a zinc-binding metalloprotease; Buts JP et al.; Saccharomyces boulardii (S . boulardii), a biotherapeutic agent effective in acute and chronic enterocolopathies, produces trophic intestinal effects at least in part mediated by the endoluminal release of polyamines . However, the effects of the yeast on peptide hydrolysis have not yet been studied . The objectives of this study were to assess in suckling rats the endoluminal and mucosal aminopeptidase activities in response to S . boulardii treatment and to analyze their related mechanisms . Peptidase activities were assayed on yeast cells by using several L-amino acid-p-nitroanilide substrates in the pH range of 2 to 10 . A marked hydrolytic activity was found for L-leucine-p-nitroanilide that peaked at pH = 8 (K(m) = 0.334 mM, V(max) = 44.7 micromol.min(-1).g(-1) protein) . N-terminal peptide hydrolysis was confirmed using as substrate L-Leu-Gly-Gly (K(m) = 4.71 mM, V(max) = 18.08 micromol.min(-1).g(-1) protein) . Enzyme reactions were inhibited in the presence of 1 mM Zn(2+) . Oral treatment of sucklings with S . boulardii significantly enhanced jejunal and ileal mucosal leucine-aminopeptidase activities by 24 and 34%, respectively, over controls . In concordance, aminopeptidase activity was enhanced in jejunal and ileal endoluminal fluid samples by 47 and 105%, respectively . By use of an IgG-purified antibody raised against the zinc-binding domain common to metalloproteases, the yeast aminopeptidase was immunoprecipitated and detected as an heteromeric enzyme of 108 and 87-kD subunits . S . boulardii, when given orally to suckling rats, is able to significantly enhance hydrolysis of N-terminal oligopeptides in both endoluminal fluid and intestinal mucosa by the endoluminal release of a leucine aminopeptidase that appears to be a zinc-binding metalloprotease belonging to the M1 family of peptidases. Oncogene, 2002 Jan 21, 21(4), 512 - 21 Transcriptional silencing at Saccharomyces telomeres: implications for other organisms; Tham WH et al.; Telomeres are the natural ends of eukaryotic chromosomes . In most organisms, telomeres consist of simple, repeated DNA with the strand running 5' to 3' towards the end of the chromosome being rich in G residues . In cases where the very end of the chromosome has been examined, the G-strand is extended to form a short, single stranded tail . The chromatin structure of telomeric regions often has features that distinguish them from other parts of the genome . Because telomeres protect chromosome ends from degradation and end-to-end fusions and prevent the loss of terminal DNA by serving as a substrate for telomerase, they are essential for the stable maintenance of eukaryotic chromosomes . In addition to their essential functions, telomeres in diverse organisms are specialized sites for gene expression . Transcription of genes located next to telomeres is repressed, a phenomenon termed telomere position effect (TPE) . TPE is best characterized in the yeast Saccharomyces cerevisiae . This article will focus on the silencing properties of Saccharomyces telomeres and end with speculation on the role of TPE in yeasts and other organisms. Clin Immunol, 2002 Feb, 102(2), 162 - 8 Antineutrophil cytoplasmic antibodies, anti-Saccharomyces cerevisiae antibodies, and specific IgE to food allergens in children with inflammatory bowel diseases; Bartunkova J et al.; Differential diagnosis between ulcerative colitis (UC) and Crohn's disease (CD) is difficult in the initial phases in pediatric patients with inflammatory bowel diseases (IBD) . This study was performed to determine the significance of anti-neutrophil cytoplasmic antibodies (ANCA) and anti-Saccharomyces cerevisiae antibodies (ASCA) in IBD . ANCA were specified with regard to their antigenic specifity, significance to the diagnosis, and correlation of titer with the disease activity . The occurrence of food allergy was questioned, too . Serum samples from 44 children with UC (n = 23) or CD (n = 21) and from disease-control children (coeliac disease, n = 21) were analyzed for IgG ANCA, ANCA target antigens, IgA and IgG ASCA, and IgE to food allergens . Results show that ANCA occur more frequently in UC than in CD and disease-control (74, 24, and 10%, respectively) . The presence of ANCA does not reflect disease activity . Antigenic specificity does not differ in any group . IgA-ASCA are found more often in patients with CD (76% versus 17% in UC) . The testing for both ANCA and ASCA enabled clear-cut differential diagnosis between UC and CD based on the high specificity (ANCA+ ASCA- 92.5% for UC, ANCA- ASCA+ 93.2% for CD) . Specific IgE to food allergens were found in 8.7, 14.3, and 23.8% of patients with UC, CD, and coeliac disease, respectively . We conclude that combined testing of ANCA and ASCA represents a valuable tool in the differential diagnosis between UC and CD in pediatric patients, minimizing invasive diagnostic procedures . Monitoring of ANCA, its specificity, and titer determination does not bring more information . Testing for specific IgE to food allergens may be considered in individual patients . Dig Liver Dis, 2001 Dec, 33(9), 755 - 61 Detection of anti-Saccharomyces cerevisiae antibodies in Crohn's disease: is it a reliable diagnostic and prognostic marker? Sostegni R, Daperno M, Ercole E, Rigazio C, Bresso F, Masoero G, Castellino F, Zaffino C, Rocca R, Molinaro GC, Rocca G, Astegiano M, Pera A. BACKGROUND: In the past few years, serologic markers have been proposed in inflammatory bowel disease . Anti-Saccharomyces cerevisiae antibodies showed high specificity for Crohn's disease . A prognostic role for serology has also been hypothesised . AIMS: To evaluate anti-Saccharomyces cerevisiae antibody distribution in an unselected Italian inflammatory bowel disease population . To analyse whether anti-Saccharomyces cerevisiae antibody status (positive/negative) and/or anti-Saccharomyces cerevisiae antibody titres are associated with clinical variables and outcome measures in Crohn's disease patients . PATIENTS AND METHODS: A series of 299 inflammatory bowel disease patients were evaluated; serum samples were taken and a short clinical history was recorded . anti-Saccharomyces cerevisiae antibodies IgG enzyme-linked immunosorbent assay Medilab (Milan, Italy) kit was used in order to determine anti-Saccharomyces cerevisiae antibody status . RESULTS: Sensitivity, specificity and likelihood ratio for positive test in the differential diagnosis of inflammatory bowel disease was 59%, 89%, 8.1, respectively . Clinical variables significantly associated with anti-Saccharomyces cerevisiae antibody status in logistic regression were found to be ileal location (p=0.01) and earlier age at diagnosis (p<0.01) . Among ileal Crohn's disease patients, there was a trend in concordance between anti-Saccharomyces cerevisiae antibody titres and higher number of surgical procedures which was not statistically significant applying more complex statistics . CONCLUSIONS: In an Italian inflammatory bowel disease population, anti-Saccharomyces cerevisiae antibodies status showed characteristics similar to those previously reported . Anti-Saccharomyces cerevisiae antibody positivity is associated with ileal involvement and with earlier onset of Crohn's disease. Nucleic Acids Res, 2001 Dec 15, 29(24), 5156 - 62 Effect of chromosomal locus, GC content and length of homology on PCR-mediated targeted gene replacement in Saccharomyces; Gray M et al.; Targeted gene replacement (TGR) using fragments generated by PCR is a widely-used technique for deleting genes in Saccharomyces cerevisiae . We found that the efficiency of this procedure, defined as the fraction of transformants that delete the targeted gene, varied by >10-fold depending on the sequence being targeted . We examined the effect of chromosomal position, length of homology and GC content on TGR efficiency . When URA3 was positioned at five different chromosomal locations, the efficiency of replacing this gene with LEU2 remained the same . Similarly, varying the length of homology from 35 to 60 bp had only a small effect on the efficiency of targeting (<50%), though an increase in the length of homology to 200 bp on one end of the disruption fragment did increase TGR efficiency . Strikingly, as GC content in the target sequence increased, the efficiency of targeting also increased . When TGR efficiency was high, the frequency of untargeted integration events was low . These results suggest two strategies for designing TGR primers: (i) use 40 bp targeting sequences containing 40-50% GC, and (ii) if necessary, increase TGR efficiency by extending the length of homology on one end of the disruption fragment. Mol Biol Cell, 2001 Dec, 12(12), 4078 - 89 Repair of chromosome ends after telomere loss in Saccharomyces; Mangahas JL et al.; Removal of a telomere from yeast chromosome VII in a strain having two copies of this chromosome often results in its loss . Here we show that there are three pathways that can stabilize this broken chromosome: homologous recombination, nonhomologous end joining, and de novo telomere addition . Both in a wild-type and a recombination deficient rad52 strain, most stabilization events were due to homologous recombination, whereas nonhomologous end joining was exceptionally rare . De novo telomere addition was relatively rare, stabilizing <0.1% of broken chromosomes . Telomere addition took place at a very limited number of sites on chromosome VII, most occurring close to a 35-base pair stretch of telomere-like DNA that is normally approximately 50 kb from the left telomere of chromosome VII . In the absence of the Pif1p DNA helicase, telomere addition events were much more frequent and were not concentrated near the 35-base pair tract of telomere-like DNA . We propose that internal tracts of telomere-like sequence recruit telomerase by binding its anchor site and that Pif1p inhibits telomerase by dissociating DNA primer-telomerase RNA interactions . These data also show that telomeric DNA is essential for the stable maintenance of linear chromosomes in yeast. Proc Natl Acad Sci U S A, 2001 Dec 4, 98(25), 14524 - 9 Epub 2001 Nov 27. Meiotic recombination frequencies are affected by nutritional states in Saccharomycescerevisiae; Abdullah MF et al.; Meiotic recombination in the yeast Saccharomyces cerevisiae is initiated by programmed double-strand breaks at selected sites throughout the genome (hotspots) . alpha-Hotspots are binding sites for transcription factors . Double-strand breaks at alpha-hotspots require binding of transcription factor but not high levels of transcription per se . We show that modulating the production of the transcription factor Gcn4p by deletion or constitutive transcription alters the rate of gene conversion and crossing-over at HIS4 . In addition, we show that alterations in the metabolic state of the cell change the frequency of gene conversion at HIS4 in a Gcn4p-dependent manner . We suggest that recombination data obtained from experiments using amino acid and other biosynthetic genes for gene disruptions and/or as genetic markers should be treated cautiously . The demonstration that Gcn4p affects transcription of more than 500 genes and that the recombinationally "hottest" ORFs tend to be Gcn4p-regulated suggest that the metabolic state of a cell, especially with respect to nitrogen metabolism, is a determinant of recombination rates . This observation suggests that the effects of metabolic state may be global and may account for some as yet unexplained features of recombination in higher organisms, such as the differences in map length between the sexes. Mol Biol Cell, 2001 Nov, 12(11), 3417 - 27 Lag1p and Lac1p are essential for the Acyl-CoA-dependent ceramide synthase reaction in Saccharomyces cerevisae; Schorling S et al.; Lag1p and Lac1p are two homologous transmembrane proteins of the endoplasmic reticulum in Saccharomyces cerevisiae . Homologous genes have been found in a wide variety of eukaryotes . In yeast, both genes, LAC1 and LAG1, are required for efficient endoplasmic reticulum-to-Golgi transport of glycosylphosphatidylinositol-anchored proteins . In this study, we show that lag1 Delta lac1 Delta cells have reduced sphingolipid levels due to a block of the fumonisin B1-sensitive and acyl-CoA-dependent ceramide synthase reaction . The sphingolipid synthesis defect in lag1 Delta lac1 Delta cells can be partially corrected by overexpression of YPC1 or YDC1, encoding ceramidases that have been reported to have acyl-CoA-independent ceramide synthesis activity . Quadruple mutant cells (lag1 Delta lac1 Delta ypc1 Delta ydc1 Delta) do not make any sphingolipids, but are still viable probably because they produce novel lipids . Moreover, lag1 Delta lac1 Delta cells are resistant to aureobasidin A, an inhibitor of the inositolphosphorylceramide synthase, suggesting that aureobasidin A may be toxic because it leads to increased ceramide levels . Based on these data, LAG1 and LAC1 are the first genes to be identified that are required for the fumonisin B1-sensitive and acyl-CoA-dependent ceramide synthase reaction. Mol Genet Genomics, 2001 Aug, 265(6), 1120 - 8 Genetic selection in Saccharomyces of mutant mammalian adenylyl cyclases with elevated basal activities; Haney SA et al.; We show that co-expression of rat Galphas together with type I, II, IV, or VI mammalian adenylyl cyclase (AC) can suppress the growth defect of cyr1 strains of Saccharomyces cerevisiae, which lack a functional endogenous AC . Complemention of cvr1 is not observed in the absence of Galphas, indicating that the mammalian ACs retain their normal regulatory behavior in yeast . Selection for Galphas-independent growth of (cyr1 strains expressing type IV AC yielded several ACIV mutants with enhanced basal activity, each of which had a single amino acid substitution in the conserved C1a or C2a region of the protein . Expression of two of the mutant ACs in HEK293 cells resulted in increased levels of cAMP and elevated adenylyl cyclase activity . Further selection for reverting mutations in one of these constitutively active AC mutants yielded three independent intragenic suppressor mutations . The d