Uncovering novel transcriptional regulatory pathways in the cell cycle

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DESCRIPTION (provided by applicant): Misregulated cell cycle progression is a defining characteristic of cancer, thus a clear understanding of normal cell cycle progression is critical to an informed picture of carcinogenesis. Recent studies in yeast and mammals suggest that cell cycle-dependent transcription is more extensive than was previously apreciated. Nearly 800 genes (14% of the genome) in yeast exhibit cell cycle-dependent patterns of transcription, yet fewer than 40% of these genes are directly regulated by known cell cycle transcription factors. This indicates that we are missing several regulatory pathways involved in cell cycle progression. This proposal aims to address two fundamental questions in cell cycle biology: What are the DNA-protein interactions that control cell cycle-specific transcription, and how are these interactions regulated by the core cell cycle machinery? Specifi Aim 1: Idenitfy novel DNA-protein interactions that regulate the cell cycle. I will use flow cytometry of transcription factor deletion yeast strains to find transcription factors required for normal cell cycle progression. I will identify the target genes regulated by these transcription factors using genome-wide expression profiling and chromatin immunoprecipitation, and determine the associated cis-regulatory sites using an array of computational and experimental tools. The results of this aim will reveal novel cell cycle transcription factors, their targets, and associated cis-regulatory sites. Specific Aim 2: Understand how the cyclin-dependent kinase Cdc28 regulates cell cycle progression through interactions with transcription factors. Using flow cytometry and mutations at conserved phosphorylation sites, I will identify transcription factors whose Cdc28 phosphorylation sites are required for normal cell cycle progression. I will verify that these transcription factors are true Cdc28 substrates through in vitro and in vivo kinase assays. I will use genome-wide expression profiling to identify target genes that are misexpressed when Cdc28 regulation of their cognate transcription factors is abolished. Successful completion of this work will lead to a greater knowledge of the DNA-protein interactions that make up the cell cycle transcriptional regulatory network, as well as lead to a better understanding of how this network interacts with the core cell cycle machinery. Relevance: Many human diseases, notably cancer, involve the loss of control of cell division. Understanding how cell division is regulated is essential to our ability to treat and prevent such diseases. This project aims to study the control of cell division by using genomic tools to examine how genes are regulated during the cell division cycle.