Retromer Complex and PI3K Complex II-Related Genes Mediate the Yeast (Saccharomyces cerevisiae) Sodium Metabisulfite Resistance Response.
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ABSTRACT:
SUBMITTER: Jin X
PROVIDER: S-EPMC8699849 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Effect of either FLO8 or MSS11 deletion and -overexpression on yeast transcript profiles compared to wild type in laboratory yeast strains Σ1278b and S288c - also the effect of FLO11 (MUC1) overexpression in the Σ1278b genetic background The aim of this study was to (1) perform a repeat analysis (to improve statistical analysis of these data sets) similar to data submitted previously (GSE17716) and also (2) study the effect of FLO11 over-expression on the transcriptome. Background: The outer cell wall of the yeast Saccharomyces cerevisiae serves as the interface with the surrounding environment and defines cell-cell and cell-surface interactions. Many of these interactions are facilitated by specific adhesins that belong to the Flo protein family. This family of mannoproteins has been implicated in phenotypes such as flocculation and substrate adhesion as well as pseudohyphal growth. Genetic data strongly suggest that individual Flo proteins are responsible for many specific cellular adhesion phenotypes. However, it remains unclear whether such phenotypes are determined solely by the nature of the expressed FLO genes or rather the result of a combination of FLO gene expression and other cell wall properties and cell wall proteins. Mss11p has been shown to be a central element of FLO1 and FLO11 gene regulation and acts together with the cAMP-PKA-dependent transcription factor Flo8p. We use genome wide transcript analysis to identify genes that are direct ly or indirectly regulated by Mss11p in the genetic backgrounds: Sigma1278b and S288c. Sigma 1278b is the strain historically used for the study of pseudohyphae (FLO11 expression) but we also included S288c as this strain is widely used in the research community and was used to determine the first full genome sequence (Thus correspond with SGD information). We also compare this data with transcriptome data from Sigma 1278b yeast over-expressing FLO8 to compare similarities/differences between these two signalling factors. Finally the effect of FLO11 over-expression in Sigma1278b on global transcription is studied so that we can differentiate between "direct" gene targets of Flo8p or Mss11p, and those regulated as a result by the "indirect" effect caused by modified cell wall Flo11p levels.
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Project description:In eukaryotic cells, cohesin holds sister chromatids together until they separate into daughter cells during mitosis. We have used chromatin immunoprecipitation coupled with microarray analysis (ChIP chip) to produce a genome-wide description of cohesin binding to meiotic and mitotic chromosomes of Saccharomyces cerevisiae. A computer program, PeakFinder, enables flexible, automated identification and annotation of cohesin binding peaks in ChIP chip data. Cohesin sites are highly conserved in meiosis and mitosis, suggesting that chromosomes share a common underlying structure during different developmental programs. These sites occur with a semiperiodic spacing of 11 kb that correlates with AT content. The number of sites correlates with chromosome size; however, binding to neighboring sites does not appear to be cooperative. We observed a very strong correlation between cohesin sites and regions between convergent transcription units. The apparent incompatibility between transcription and cohesin binding exists in both meiosis and mitosis. Further experiments reveal that transcript elongation into a cohesin-binding site removes cohesin. A negative correlation between cohesin sites and meiotic recombination sites suggests meiotic exchange is sensitive to the chromosome structure provided by cohesin. The genome-wide view of mitotic and meiotic cohesin binding provides an important framework for the exploration of cohesins and cohesion in other genomes.
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Project description:Clusters of DNA damage, also called multiply damaged sites (MDS), are a signature of ionizing radiation exposure. They are defined as two or more lesions within one or two helix turns, which are created by the passage of a single radiation track. It has been shown that the clustering of DNA damage compromises their repair. Unresolved repair may lead to the formation of double-strand breaks (DSB) or the induction of mutation. We engineered three complex MDS, comprised of oxidatively damaged bases and a one-nucleotide (1 nt) gap (or not), in order to investigate the processing and the outcome of these MDS in yeast Saccharomyces cerevisiae. Such MDS could be caused by high linear energy transfer (LET) radiation. Using a whole-cell extract, deficient (or not) in base excision repair (BER), and a plasmid-based assay, we investigated in vitro excision/incision at the damaged bases and the mutations generated at MDS in wild-type, BER, and translesion synthesis-deficient cells. The processing of the studied MDS did not give rise to DSB (previously published). Our major finding is the extremely high mutation frequency that occurs at the MDS. The proposed processing of MDS is rather complex, and it largely depends on the nature and the distribution of the damaged bases relative to the 1 nt gap. Our results emphasize the deleterious consequences of MDS in eukaryotic cells.
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Project description:Effect of either FLO8 or MSS11 deletion and -overexpression on yeast transcript profiles compared to wild type in laboratory yeast strains Σ1278b and S288c - also the effect of FLO11 (MUC1) overexpression in the Σ1278b genetic background The aim of this study was to (1) perform a repeat analysis (to improve statistical analysis of these data sets) similar to data submitted previously (GSE17716) and also (2) study the effect of FLO11 over-expression on the transcriptome. Background: The outer cell wall of the yeast Saccharomyces cerevisiae serves as the interface with the surrounding environment and defines cell-cell and cell-surface interactions. Many of these interactions are facilitated by specific adhesins that belong to the Flo protein family. This family of mannoproteins has been implicated in phenotypes such as flocculation and substrate adhesion as well as pseudohyphal growth. Genetic data strongly suggest that individual Flo proteins are responsible for many specific cellular adhesion phenotypes. However, it remains unclear whether such phenotypes are determined solely by the nature of the expressed FLO genes or rather the result of a combination of FLO gene expression and other cell wall properties and cell wall proteins. Mss11p has been shown to be a central element of FLO1 and FLO11 gene regulation and acts together with the cAMP-PKA-dependent transcription factor Flo8p. We use genome wide transcript analysis to identify genes that are direct ly or indirectly regulated by Mss11p in the genetic backgrounds: Sigma1278b and S288c. Sigma 1278b is the strain historically used for the study of pseudohyphae (FLO11 expression) but we also included S288c as this strain is widely used in the research community and was used to determine the first full genome sequence (Thus correspond with SGD information). We also compare this data with transcriptome data from Sigma 1278b yeast over-expressing FLO8 to compare similarities/differences between these two signalling factors. Finally the effect of FLO11 over-expression in Sigma1278b on global transcription is studied so that we can differentiate between "direct" gene targets of Flo8p or Mss11p, and those regulated as a result by the "indirect" effect caused by modified cell wall Flo11p levels. We used two laboratory yeast strains that behave different with regard to adhesion phenotypes. By comparing yeast deleted in either FLO8 or MSS11 to wild type, or yeast overexpressing these genes, in both genetic backgrounds, we investigate the role of Flo8p and Mss11p on yeast transcription. In addition the effect of the over-expression of the adhesin gene FLO11 was studied in Sigma 1278b. By using similar growth conditions to what we use for adhesion phenotype determination we aim to correlate transcription profile changes to yeast behaviour (phenotypes).
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Project description:Genome analysis of the yeast Saccharomyces cerevisiae identified 68 genes encoding flavin-dependent proteins (1.1% of protein encoding genes) to which 47 distinct biochemical functions were assigned. The majority of flavoproteins operate in mitochondria where they participate in redox processes revolving around the transfer of electrons to the electron transport chain. In addition, we found that flavoenzymes play a central role in various aspects of iron metabolism, such as iron uptake, the biogenesis of iron-sulfur clusters and insertion of the heme cofactor into apocytochromes. Another important group of flavoenzymes is directly (Dus1-4p and Mto1p) or indirectly (Tyw1p) involved in reactions leading to tRNA-modifications. Despite the wealth of genetic information available for S. cerevisiae, we were surprised that many flavoproteins are poorly characterized biochemically. For example, the role of the yeast flavodoxins Pst2p, Rfs1p and Ycp4p with regard to their electron donor and acceptor is presently unknown. Similarly, the function of the heterodimeric Aim45p/Cir1p, which is homologous to the electron-transferring flavoproteins of higher eukaryotes, in electron transfer processes occurring in the mitochondrial matrix remains to be elucidated. This lack of information extends to the five membrane proteins involved in riboflavin or FAD transport as well as FMN and FAD homeostasis within the yeast cell. Nevertheless, several yeast flavoproteins, were identified as convenient model systems both in terms of their mechanism of action as well as structurally to improve our understanding of diseases caused by dysfunctional human flavoprotein orthologs.
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Project description:In the yeast Saccharomyces cerevisiae the TOR complex 1 (TORC1) controls many growth-related cellular processes and is essential for cell growth and proliferation. Macrolide antibiotic rapamycin, in complex with a cytosol protein named FKBP12, specifically inhibits TORC1, causing growth arrest. The FKBP12-rapamycin complex interferes with TORC1 function by binding to the FRB domain of the TOR proteins. In an attempt to understand the role of the FRB domain in TOR function, we identified a single point mutation (Tor2(W2041R) ) in the FRB domain of Tor2 that renders yeast cells rapamycin resistant and temperature sensitive. At the permissive temperature, the Tor2 mutant protein is partially defective for binding with Kog1 and TORC1 is impaired for membrane association. At the restrictive temperature, Kog1 but not the Tor2 mutant protein, is rapidly degraded. Overexpression of ubiquitin stabilizes Kog1 and suppresses the growth defect associated with the tor2 mutant at the nonpremissive temperature. We find that ubiquitin binds non-covalently to Kog1, prevents Kog1 from degradation and stabilizes TORC1. Our data reveal a unique role for ubiquitin in regulation of TORC1 and suggest that Kog1 requires association with the Tor proteins for stabilization.
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