Roles of Chromatin Modification in BRCA1 Dependent DNA Repair

Contact PD/PI: Greenberg, Roger A. Summary Encounters between DNA replication proteins (replisome) and specific genomic lesions invoke requirements for BRCA1 and BRCA2 (BRCA) dependent repair by homologous recombination (HR). Replisome conflicts with endogenous DNA damage also elicit error prone homology directed repair (HDR) mechanisms and damage tolerance pathways that allow replication through persistent genomic lesions. Such noncanonical HDR pathways become prominent in BRCA mutated cells. They are also responsible for telomere maintenance in ~15% of human cancers that utilize the alternative lengthening of telomeres (ALT) mechanism. We have developed unique systems to investigate the endogenous lesions that provoke BRCA dependent and independent HDR and have used them to understand how each repair mechanism is initiated and resolved. In the current R01 funding period, we have demonstrated that (1) loss of the PAR dependent chromatin remodeler, ALC1, potentiates PARP inhibitor responses by several orders of magnitude specifically in HR deficient cells, (2) Break induced replication utilizes PCNA-Ub interaction with SNM1A for resection dependent template switch recombination bypass of polymerase blocking lesions, and (3) BLM helicase activity generates single-strand DNA at replicating telomeres to promote a damage response that executes ALT. The proposed research investigates the biochemical basis for these phenomena, and how they process specific endogenous DNA lesions to provoke either classical or alternative forms of recombination. We utilize approaches to identify the full spectrum of responses that occur at damaged telomeres and interrogate specific DNA replication associated structures that ALC1, SNM1A, and BLM act upon to affect repair mechanism. Overlapping genetic vulnerabilities in ALT and BRCA mutant cells suggest commonality in the repair processes that ensue in each scenario. Our overarching goals are to delineate molecular events necessary for each process and understand how classical and alternative mechanisms of homologous recombination intersect in the settings of (1) BRCA mutation, and (2) during ALT. These objectives will be performed in parallel and with equal emphasis. Our studies will yield fundamental advances to the understanding cancer genome integrity control.