Impact of RING and BRCT domain mutations on BRCA1 protein stability, localization and recruitment to DNA damage.
Mutations within the tumor suppressor BRCA1 cause the majority of hereditary breast and ovarian cancers. The BRCA1 protein is an important regulator of DNA double-strand break repair, and BRCA1-deficient cells are highly sensitive to ionizing radiation. Furthermore, BRCA1 function may contribute to enforcement of the G(2) cell cycle checkpoint. E3-ubiquitin ligase activity is the only known enzymatic activity of BRCA1, which is mediated by the N-terminal RING finger domain. The C-terminal BRCT repeat domain, which mediates protein-protein interactions, is the only other identified structural domain. By investigating cancer-linked mutations within each domain, we demonstrate that truncation of the BRCT domain greatly impairs the stability and nuclear localization of BRCA1 protein. A missense mutation within the RING domain does not affect these biochemical properties. However, both mutant forms of BRCA1 fail to colocalize in nuclear foci with the known BRCA1-interacting proteins BARD1 and BACH1, which are important for DNA repair. This failure occurs despite the continued ability of the RING mutant protein to interact with BACH1 and the ability of the BRCT mutant to interact with BARD1. Furthermore, neither mutant form of BRCA1 is recruited into DNA damage-associated foci marked by gamma-H2AX. Therefore, our data suggest that both the RING and BRCT domains of BRCA1 are required for an early step in the function of BRCA1 during DNA repair: recruitment to the sites of DNA damage.
Nelson
Andrew C
AC
Department of Pathology and Program in Cancer Biology, University of Colorado Denver, Aurora, Colorado 80045, USA.
Holt
Jeffrey T
JT
D000328:Adult; D019313:BRCA1 Protein / Q000235:genetics / Q000378:metabolism*; D001943:Breast Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D044584:Carcinoma, Ductal / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D004249:DNA Damage; D005260:Female; D006801:Humans; D009154:Mutation
D019313:BRCA1 Protein