Role of homologous recombination in the replication stress response

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PROJECT SUMMARY/ABSTRACT Precise replication of DNA is required to maintain genome stability. Replication forks face many obstacles from both endogenous and exogenous sources that result in fork stalling or breakage threatening genome integrity. The double strand break repair pathway, homologous recombination, has critical roles at stalled replication forks independent of double strand break repair. The importance of this pathway is highlighted by patients with inherited chromosomal instability orders and cancer predisposition syndromes. Although an intense area of study, the mechanistic role of recombination proteins at replication forks is still poorly understood. A comprehensive understanding of the replication response is critical for the understanding the molecular mechanisms of human disease and to lead to development of novel therapeutics for patients harboring defects in replication response genes. The long-term goal of the PI to elucidate the roles of recombination proteins in the replication stress response. Here we will elucidate the role of recombination proteins at hydroxyurea-stalled replication forks in two distinct projects. In Project 1, we use a powerful separation-of-function allele of the central recombination enzyme, RAD51, to determine how RAD51 protects the integrity of the replication fork during unchallenged and stressed conditions using a combination of genetic, molecular biology and proteomic approaches. In Project 2, we propose a series of experiments to investigate the role of additional recombination accessory factors in the replication response. Preliminary data from our lab has uncovered a novel role for recombination proteins in protecting the integrity of stalled replication forks. We will test our current models through mechanistic dissection of the replication stress response pathway using genetic and molecular approaches. The work presented here will lay the foundation for future studies involving the elucidation of molecular mechanisms of recombination proteins involved in the replication stress response. The completion of this work will lead to not only a better of understanding of the role recombination proteins play at stalled replication forks, but will also provide molecular insight into how disruption of this pathway results in human disease. A complete understanding of this pathway is essential for development of therapeutics for patients with inherited genome instability disorders and cancer that target the cellular replication response.