Interaction between DNA Polymerase ß and BRCA1.

Masaoka A, Gassman NR, Horton JK, Kedar PS, Witt KL, Hobbs CA, Kissling GE, Tano K, Asagoshi K, Wilson SH.
Source: PLoS ONE
Publication Date: (2013)
Issue: 8 (6): e66801
Research Area:
Molecular Biology
The breast cancer 1 (BRCA1) protein is a tumor suppressor playing roles in DNA repair and cell cycle regulation. Studies of DNA repair functions of BRCA1 have focused on double-strand break (DSB) repair pathways and have recently included base excision repair (BER). However, the function of BRCA1 in BER is not well defined. Here, we examined a BRCA1 role in BER, first in relation to alkylating agent (MMS) treatment of cells and the BER enzyme DNA polymerase ß (pol ß). MMS treatment of BRCA1 negative human ovarian and chicken DT40 cells revealed hypersensitivity, and the combined gene deletion of BRCA1 and pol ß in DT40 cells was consistent with these factors acting in the same repair pathway, possibly BER. Using cell extracts and purified proteins, BRCA1 and pol ß were found to interact in immunoprecipitation assays, yet in vivo and in vitro assays for a BER role of BRCA1 were negative. An alternate approach with the human cells of immunofluorescence imaging and laser-induced DNA damage revealed negligible BRCA1 recruitment during the first 60 s after irradiation, the period typical of recruitment of pol ß and other BER factors. Instead, 15 min after irradiation, BRCA1 recruitment was strong and there was ?-H2AX co-localization, consistent with DSBs and repair. The rapid recruitment of pol ß was similar in BRCA1 positive and negative cells. However, a fraction of pol ß initially recruited remained associated with damage sites much longer in BRCA1 positive than negative cells. Interestingly, pol ß expression was required for BRCA1 recruitment, suggesting a partnership between these repair factors in DSB repair.