Genetic Analysis of E2F Function During Development

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DESCRIPTION (provided by applicant): The long-term goal of this research project is to understand the regulatory mechanisms used to control the G1-S cell cycle transition in a developing organism. The focus of the research is the E2F family of transcription factors, which both positively and negatively regulate the expression of genes required for the G1-S transition, including those involved in nucleotide biosynthesis, recognition of origins of replication, DNA replication, and control of cell cycle progression. The overexpression of E2F is sufficient to drive quiescent cells into S-phase, and genetic inhibition of E2F dependent transcription attenuates DNA synthesis and leads to the accumulation of cells in G1. Correspondingly, E2F genes behave as oncogenes when overexpressed, and mutations that deregulate E2F activity are found in many cancers. However, the full scope of E2F activity is undefined, and recent studies suggest that E2F may regulate the expression of many genes that are involved in diverse aspects of cellular function in addition to G1-S control. Moreover, the mechanisms by which E2F transcription factors are regulated by developmental signals in the context of a whole animal are not completely understood. The powerful molecular genetics, cell biology, and transgenic techniques of Drosophila melanogaster will be utilized to study E2F regulation and function. Drosophila contains two E2F transcription factors, termed E2F1 and E2F2. The current working model holds that E2F2 acts as a dedicated repressor while E2F1 functions as both an activator and a repressor. How these activities are regulated in order to control the cell cycle at different stages of development will be addressed in three aims: 1) Determine the mechanism of interplay between E2F1 and E2F2 during development using genetic and biochemical strategies, 2) Determine the contribution of E2F-dependent transcription to DNA replication through the use of gene expression microarrays to identify and characterize novel E2F target genes, and 3) Elucidate the developmental pathways that regulate E2F activity by employing a forward genetic screen to identify novel regulators of E2F during embryogenesis.