Control of gene expression at the start of the eukaryotic cell division cycle is the initiating event for cell proliferation in most cell types. Mutations deregulating that crucial transcriptional switch lead to uncontrolled cell growth and division. In humans, defects in that regulatory pathway are associated with development disorders and cancer. We propose to study the analogous transcriptional regulatory network in the budding yeast in the interest of understanding the nature of that regulation. Greater than 300 genes are coordinately expressed during late G1 phase in preparation for entry into a new cell cycle. Those genes are under the control of two related transcription factors, SBF and MBF, functional analogs of the E2F gene family in humans. Those factors act coordinately to promote expression and are repressed as cells progress into S phase. However, SBF and MBF are regulated by distinct transcriptional repressors that govern both activation and repression. As a consequence, the two factors are subject to differential regulation by cellular and environmental signals. We propose to study the differences and similarities between the mechanisms governing gene expression controlled by SBF and MBF in the context of three Specific Aims. In the first Aim we will elucidate the specific regulatory factors governing the activation of MBF target genes, the more poorly understood of the two transcription factors. In the second Aim we will establish the mechanism of repression of MBF targets as cells exit G1 phase. Because transcriptional repression is central to the regulation of this gene network, we propose to apply high-resolution genome wide location analysis to study the influence of transcriptional repressors on histone modifying enzymes and their marks on nucleosomes. Finally, in the third Aim we will study the molecular basis for the differential regulation of SBF and MBF by their transcriptional repressors, Whi5 and Nrm1, respectively and establish the mechanism by which MBF evades one mechanism of transcriptional repression when the DNA replication checkpoint is induced by genotoxic stress. The comparative analysis of these two transcription factors acting on more than 200 genes will lead to a new understanding, not only of the mechanisms of transcriptional repression at cell cycle regulated genes, but also on the general role of transcriptional repression in eukaryotes.
This proposal outlines a plan to understand the transcriptional regulation of a large gene family that controls cell proliferation in yeast. This work has direct applications in understanding control of gene expression in eukaryotes. Mutations affecting the analogous regulatory network in humans are associated with developmental defects and cancer.
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