Regulation of DNA replication kinetics by BRCA2 after DNA damage

Germline mutations in BRCA2 predispose carriers to breast, ovarian, pancreatic, and other cancers. The gene encodes a very large protein that plays critical roles in genome integrity control by promoting homologous recombination (HR)-mediated repair of DNA double strand breaks (DSBs), DNA damage-induced cell cycle checkpoint, and stability of stalled DNA replication forks, etc. Besides, BRCA2 may play a direct role in DNA replication, as its mutant cells have long been known to have the so-called radio-resistant DNA synthesis (RDS) phenotype, which reflects a defect in the intra-S phase checkpoint, a mechanism that slows down DNA replication after DNA damage presumably to allow time for DNA repair and prevent replication of damage DNA. However, the potential role of BRCA2 in DNA replication initiation or elongation has not been defined. In our preliminary studies, we found that BRCA2 interacts with an essential DNA replication factor, MCM10, and that this interaction restrains replication fork progression, promotes fork stalling and sustains origin firing after DNA damage. We hypothesize that BRCA2 regulates DNA replication kinetics after DNA damage through its interaction with MCM10 and that a dysregulation of replication kinetics after DNA damage, or intra-S phase checkpoint defect, leads to increased mutation rate, cancer development and therapy resistance. We propose 3 specific aims to test the hypotheses. In Aim 1, we will define the mechanisms of BRCA2 function in DNA replication kinetics after DNA damage. In Aim 2, we will determine the role of the BRCA2-MCM10 interaction in DNA replication fidelity and cancer cell response to DNA damaging therapies. In Aim 3, we will determine the role of BRCA2-MCM10 interaction in spontaneous and radiation-induced tumor development. Our findings and proposed studies will elucidate key mechanisms of BRCA2 function in DNA replication, provide new insights into the regulation of replication kinetics after DNA damage, and define the impact of RDS and replication kinetics dysregulation on cancer development and tumor relapse after radiation and chemotherapy.