Generation and characterization of phosphosite mutants in S. cerevisiae
Ontology highlight
ABSTRACT:
ORGANISM(S): Saccharomyces cerevisiae
PROVIDER: GSE123846 | GEO | 2018/12/15
REPOSITORIES: GEO
Similar Datasets
Project description:Genomic profiling of (1) histone H3K56 acetylation and (2) meiotic DSBs flanked by replication protein A (RPA). The major goal was to reveal the genome-wide distribution DSBs when the H3K56 residue was mutated to alanine (H3K56A).
2019-12-25 | GSE37487 | GEO
Project description:Cell cycle sensing of oxidative stress in Saccharomyces cerevisiae by oxidation of a specific cysteine residue in the transcription factor Swi6p. Yeast cells begin to bud and enter S phase when growth conditions are favourable during G1 phase. When subjected to oxidative stress, cells arrest at G1 delaying entry into the cell cycle allowing repair of cellular damage. Hence, oxidative stress sensing is coordinated with the regulation of cell cycle. We identified a redox sensing cysteine residue in the cell-cycle regulator of Saccharomyces cerevisiae, Swi6p, at position 404. Mutation of Cys404 to alanine abolished the ability of the cells to arrest at G1 upon treatment by lipid hydroperoxide. By constructing a truncated form of Swi6p, the Cys404 residue was found to be oxidised when cells were subjected to the oxidant. Furthermore, microarray analysis revealed that mutation of Cys404 to alanine led to loss of suppression of G1-cyclins CLN1 and PCL1 when the cells were exposed to lipid hydroperoxide. In conclusion, oxidation of Cys404 serves as a molecular sensor of oxidative stress and inhibits entry into the cell cycle by suppression of G1-cyclin expression. We used a gene expression approach to assess the involvement of Cys404 in oxidative stress by mutating this residue to alanine in order to study whether it contributes to Swi6p, a transcriptional factor, function for redox regulation of the cell cycle. Wild type, swi6-deletant, and swi6 C404A-mutated yeast cells were treated with either linoleic acid hydroperoxide (LoaOOH) or control. Three replicates per group/treatment.
2011-01-31 | E-GEOD-18334 | biostudies-arrayexpress
Project description:Generation and characterization of phosphosite mutants in S. cerevisiae
| PRJNA510038 | ENA
Project description:Investigation of whole genome gene expression level changes in HT1080 fibrosarcoma cell line after transfection CRABP1 gene and R131A CRABP1 mutant (arginine-alanine substitution in a protein active site, protein lacks the ability to interact with retinoic acid), compared to HT1080 line transfected with empty pLXSN vector.
2013-09-01 | GSE48673 | GEO
Project description:To explore the role of individual amino acids in the binding of GATAD1 peptide (position 95-109) to 14-3-3 proteins, we conducted an alanine scanning experiment. Each amino acid surrounding the phosphorylation site within the peptide was systematically mutated to alanine, except for the original alanine residues, which were mutated to glycine. These mutant peptides were immobilized on a cellulose membrane and employed to capture proteins from Hek293T lysate through pulldown assays.
2024-02-15 | PXD043788 | Pride
Project description:A systematic approach allowing the identification of the molecular way-of-action of novel potential drugs represents the golden-tool for drug-discovery. While high-throughput screening technologies of large libraries is now well established, the assessment of the drug targets and mechanism of action is still under development. Taking advantage of the yeast model Saccharomyces cerevisiae, we herein applied BarSeq, a Next Generation Sequencing-based method to the analysis of both haploinsufficiency and homozygous fitness effects of a novel antifungal drug ('089') compared to the well-known antifungal ketoconazole. '089' was a novel compound identified in during a screen for antifungal drugs, as it was showing fungicidal effects, and able to affect the yeast fitness at the mitochondrial level (Stefanini et al., 2010. (Dissection of the Effects of Small Bicyclic Peptidomimetics on a Panel of Saccharomyces cerevisiae Mutants;.J Biol Chem, 285: 23477-23485.) Integrative bioinformatic analysis of BarSeq, whole genome expression analysis and classical biological assays identified the target and cell pathways affected by the novel antifungal. Confirmation of the effects observed in the yeast model and in pathogenic fungi further demonstrated the reliability of the multi-sided approach and the novelty of the targets and way-of-action of the new class of molecules studied representing a valuable source of novel antifungals.
2016-04-30 | E-SYBR-6 | biostudies-arrayexpress
Project description:We have previously shown that in Arabidopsis the three enzymes of lower glycolysis phosphoglycerate mutase (PGAM), enolase and pyruvate kinase form a complex which play an important role in co-sub-cellar localization the mitochondria to the chloroplast. Given that their metabolites of these mutants, the complementation of pgam mutant and overexpression of PGAM are still unclear, here, we present the gas chromatography mass spectrometry-based metabolomics data and plant growth phenotypes of them. Compared with wild type, both sugar and amino acid are significantly changed at mutants of phosphoglycerate mutase, enolase, pyruvate kinase. Overexpression the PGAM could decrease the content of 3PGA, sugar and several amino acids and increase the content of alanine and pyruvate. In addition, pgam mutant could not be fully complemented by nuclear sub-localized, the side-directed-mutated and the E.coli PGAM at plant phenotype and metabolites profiling, especially the content of 3PGA, serine and intermediates of TCA cycle. These data suggested that the low glycolysis complex play an important role at both mitochondria-chloroplast co-subcellar localization and the metabolites exchange.
2022-10-10 | MTBLS4066 | MetaboLights
Project description:The fidelity of signal transmission requires the binding of regulatory molecules to their cognate targets. However, the crowded cell interior risks off-target interactions between proteins that are functionally unrelated. Understanding the constraints this imposes on cell systems evolution requires the fitness cost of spurious interactions to be quantified. Towards this end, we express human tyrosine kinases in the budding yeast S. cerevisiae. Yeast lacks bona fide tyrosine kinases and so the majority of resulting pY sites are functionless and artificial. We express 24 unique tyrosine kinases in total and perform phosphoproteomics in each case, resulting in ~30,000 phosphosites sites mapping to 3500 phosphoproteins. Examination of the fitness costs in each strain reveals a strong correlation between the number of spurious pY sites generated and negative effects on growth. Moreover, the prediction of pY effects on protein structure and on protein function (conservation-based) reveals potential for the widespread perturbation of the yeast proteome. Comparing the spurious pY sites (pre-selection) with native pY sites in human (post-selection) also demonstrates the recurrent modification of proteins and sites with no homology to native substrates. However, examination of these data together (fitness and phosphoproteomics) strongly suggests that a large number of the pY sites generated have a negligible effect on fitness. Finally, we test the hypothesis of pY counter-selection following the emergence of tyrosine kinases in metazoan species, but find no strong evidence for proteome-wide selection against spurious Y phosphorylation.
2024-09-12 | PXD045466 | Pride
Project description:Transfer RNAs (tRNAs) maintain translational fidelity through strict charging by their cognate aminoacyl-tRNA synthetase and codon:anticodon base pairing with the mRNA at the ribosome. Mistranslation occurs when an amino acid not specified by the genetic code is incorporated into a protein. Since alanyl-tRNA synthetase uniquely recognizes a G3:U70 base pair in alanine tRNAs and the anticodon plays no role in charging, alanine tRNA variants with anticodon mutations have the potential to mistranslate alanine. Our goal was to quantify mis-incorporation of alanine into proteins in Saccharomyces cerevisiae strains expressing one of 57 different alanine tRNA anticodon variants. Using mass spectrometry, we observed mistranslation for 45 of the variants when expressed on single-copy plasmids.
2023-10-02 | PXD038242 | Pride
Project description:Spore germination in Saccharomyces cerevisiae is a process in which a quiescent cell begins to divide. During germination, the cell undergoes dramatic changes in cell wall and membrane composition, as well as in gene expression. To understand germination in greater detail, we screened the S. cerevisiae deletion set for germination mutants. Our results identified two genes, TRF4 and ERG6, that are required for normal germination on solid media. TRF4 is a member of the TRAMP complex that, together with the exosome, degrades RNA polymerase II transcripts. ERG6 encodes a key step in ergosterol biosynthesis. Taken together, these results demonstrate the complex nature of germination and two genes important in the process.
| S-EPMC3276131 | biostudies-literature