INDUCTION OF PHOSPHORYLATION ON BRCA1 DURING THE CELL-CYCLE AND AFTERDNA-DAMAGE

BRCA1, the familial breast cancer susceptibility gene product, is a 22 0-kDa phosphorylated protein, BRCA1 immunoprecipitated from MCF7 cells blocked in G(1)-S phase or progressing through S-phase of the cell cy cle migrated more slowly through SDS polyacrylamide gels than BRCA1 fr om cells maintained in serum-supplemented media, serum-free media for 24 h, or delayed in G(2)-M phase by treatment with colchicine. Restora tion of BRCA1 to the faster-migrating form, which occurred on release of cells from the G(1)-S-phase block, was prevented by the phosphatase inhibitor okadaic acid, Phosphatase treatment of immunoprecipitated B RCA1 resulted in the conversion of the slower-migrating form to the fa ster-migrating form. Although these results suggested that BRCA1 was p referentially hyperphosphorylated near the G(1)-S-phase boundary of th e cell cycle, exposure of cells to DNA-damaging agents including UV li ght or treatment with hydrogen peroxide (H2O2) also promoted BRCA1 hyp erphosphorylation. These same stimuli also eliminated the punctate nuc lear staining pattern normally observed for BRCA1 in control cells, Th ese results indicate that BRCA1 undergoes cyclic hyperphosphorylation during the cell cycle; however, this modification, as well as changes in BRCA1 nuclear staining, also occurs in response to DNA damage.