Abstract 1850 The Association between Tumor Suppressor BRCA2 and DNA in Maintaining Genome Integrity

The key functions of the tumor suppressor BRCA2 include repairing DNA damages, such as DNA double-strand breaks (DSBs) or inter-strand crosslinks, and protecting stalled replication forks upon degradation. Thus, deleterious mutations in BRCA2 entail various cancers and Fanconi anemia. The BRCA2 protein possesses a composite of DNA-binding domains consisting of three Oligonucleotide Binding (OB) folds and a C-terminal region known as the C-terminal Recombinase Binding region (CTRB). The latter is encoded by the last gene exon (exon 27 in humans), also harboring the RAD51 recombinase binding region. The importance of these dual interactions in the functionality of the BRCA2 protein was examined in this study using biochemical and cell biological analyses. Firstly, we showed that the aberrations in both DNA and RAD51 binding in the CTRB lead to defects in DNA double-strand break (DSB) repair and replication fork preservation. Furthermore, the significance of DNA binding via OB folds and CTRB was explored through combinatory mutations that separately impair the DNA binding of OB folds and CTRB. The results reveal that the DNA binding of OB folds guides the recognition of ssDNA, while the CTRB facilitates interaction with dsDNA. We demonstrate that DNA binding through OB-folds is essential for RAD51-mediated homologous recombination, while the CTRB mainly plays a role in protecting DNA from nucleolytic degradation at the replication fork. These results highlight the distinctive roles of BRCA2 DNA-binding modules that contribute to the multifaceted actions of BRCA2 in DNA damage repair and replication fork protection. This study was supported by NIH grants, R50 CA265315 (Y.K.), R01 ES007061 (P.S.), and R35 CA241801 (P.S.)