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Layers of regulation on cell-cycle gene expression in the budding yeast Saccharomyces cerevisiae

ABSTRACT: Transcript profiling from a population of cell-cycle-arrested B-type cyclin mutant budding yeast. We compared transcript to protein expression levels in mutant cells.

ORGANISM(S): Saccharomyces cerevisiae

PROVIDER: GSE104904 | GEO | 2018/10/10

REPOSITORIES: GEO

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Project description:In the budding yeast Saccharomyces cerevisiae, transcription factors (TFs) regulate the periodic expression of many genes during the cell cycle, including gene products required for progression through cell-cycle events. Experimental evidence coupled with quantitative models suggest that a network of interconnected TFs is capable of regulating periodic genes over the cell cycle. Importantly, these dynamical models were built on transcriptomics data and assumed that TF protein levels and activity are directly correlated with mRNA abundance. To ask whether TF transcripts match protein expression levels as cells progress through the cell cycle, we applied a multiplexed targeted mass spectrometry approach (parallel reaction monitoring) on synchronized populations of cells. We found that protein expression of many TFs and cell-cycle regulators closely followed their respective mRNA transcript dynamics in cycling wild-type cells. Discordant mRNA/protein expression dynamics were also observed for a subset of cell-cycle TFs and for proteins targeted for degradation by E3 ubiquitin ligase complexes such as SCF (Skp1/Cul1/F-box) and APC/C (anaphase-promoting complex/cyclosome). We further profiled mutant cells lacking B-type cyclin/CDK activity (clb1-6), where oscillations in ubiquitin ligase activity, cyclin/CDKs, and cell-cycle progression are halted. We found that a number of proteins were no longer periodically degraded in clb1-6 mutants compared to wild type, highlighting the importance of post-transcriptional regulation. Finally, the TF complexes responsible for activating G1/S transcription (SBF and MBF) were more constitutively expressed at the protein level than their periodic mRNA expression levels in both wild-type and mutant cells. This comprehensive investigation of cell-cycle regulators reveals that multiple layers of regulation (transcription, protein stability, and proteasome targeting) affect protein expression dynamics during the cell cycle.

Project description:The Saccharomyces cerevisiae MYO1 gene encodes the myosin type II heavy chain (Myo1p), a protein required for normal cytokinesis in budding yeast. Deletion of the MYO1 gene prevents actomyosin-driven cytokinesis thereby activating an alternative mechanism that involves the synthesis of a remedial septum. Myo1p deficiency in yeast (myo1) also causes the formation of attached cells, abnormal budding patterns, formation of enlarged and elongated cells, increased osmotic sensitivity, delocalized chitin deposition, and increased chitin synthesis. To determine how the differential expression of genes is related to these diverse phenotypes, we analyzed the global mRNA expression profile of myo1 strains. Global mRNA expression profiles of myo1 strains and their corresponding wild type controls were obtained by hybridization to yeast oligonucleotide microarrays. Results for selected genes were confirmed by real time RT-PCR. A total of 547 differentially expressed genes were identified with 263 up-regulated and 284 down regulated genes in the myo1 strains. Gene set enrichment analysis revealed the significant over-representation of genes in the protein biosynthesis and stress response categories. Genes involved in cell wall assembly (GAS1, PSA1, CIS3, FIT1, WSC2), MAP kinase activity (SLT2), Rho1p Guanine Exchange Factor (ROM1), and regulation of cell proliferation (RAS1) were also differentially expressed in myo1 strains. Conclusions: We have presented a global mRNA expression analysis of yeast myo1 strains and hypothesized about possible correlations with morphological and biochemical phenotypes observed in these strains. We report 547 differentially regulated genes in the myo1 mutant strains. Genes involved in the control of cell proliferation, protein synthesis and maturation, DNA replication, and cell division processes were largely down regulated, suggesting a mechanism for delayed cell cycle progression and growth that involves coordinated regulation of these processes in myo1 strains. Other genes involved in the cellular response to cell wall stress and cell wall organization were largely up-regulated suggesting that cell wall biogenesis is important for the completion of cytokinesis and cell wall morphogenetic processes that may also be affected by myosin II deficiency. Gene set enrichment analysis indicates that stress response and protein biosynthesis gene categories are inversely correlated in this mutant. Keywords: Comparative genomic hybridization

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Layers of regulation on cell-cycle gene expression in the budding yeast Saccharomyces cerevisiae

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