Differential modulation of BRCA1 and BARD1 nuclear localisation and foci assembly by DNA damage.
The BRCA1/BARD1 heterodimer regulates genomic maintenance and contributes to the DNA damage checkpoint response. We previously reported that BRCA1 and BARD1 can shuttle between nucleus and cytoplasm. In this study, we evaluated the localisation patterns of BRCA1 and BARD1 in response to different types of DNA damaging agents and chemotherapeutic drugs. In MCF-7 cells, endogenous BRCA1 increased transiently in the nucleus at 2h after ionising radiation (IR), whereas BARD1 was unaffected. IR treatment did not induce nuclear export of either protein, in contrast to previous reports. DNA damage by UV radiation, etoposide or camptothecin caused a preferential down-regulation of nuclear BARD1 at 6h post-treatment. The UV-dependent loss of nuclear BARD1 was blocked by the proteasome inhibitor MG132, but not by leptomycin B, indicating a change in BARD1 nuclear degradation rather than nuclear export. MG132 also blocked the dispersal of BARD1/BRCA1 nuclear foci at 6h after UV, implicating the proteasome in repair foci disassembly. In the cytoplasm, BRCA1 and BARD1 were detected at centrosomes but their distribution was not altered by DNA damage. BARD1 displayed a stronger mitochondria accumulation than BRCA1, and became phosphorylated at mitochondria in response to DNA damage. The mitotic spindle poisons vincristine and paclitaxel had no effect on BRCA1 or BARD1 subcellular distribution. We conclude that BARD1 phosphorylation, expression and localisation patterns are regulated in the nucleus and at mitochondria in response to different forms of DNA damage, contributing to the role of BRCA1/BARD1 in DNA repair and apoptotic responses.
Westmead Institute for Cancer Research, University of Sydney, Westmead Millennium Institute at Westmead Hospital, NSW, Australia.
Kirsty M
KM
Brodie
Beric R
BR
Henderson
D021581:Active Transport, Cell Nucleus; D017209:Apoptosis; D019313:BRCA1 Protein / Q000032:analysis* / Q000378:metabolism; D002166:Camptothecin / Q000494:pharmacology; D045744:Cell Line, Tumor; D002466:Cell Nucleolus; D004249:DNA Damage; D004260:DNA Repair; D015536:Down-Regulation; D005047:Etoposide / Q000494:pharmacology; D006801:Humans; D007976:Leupeptins / Q000494:pharmacology; D010766:Phosphorylation; D034741:RNA, Small Interfering / Q000378:metabolism; D011839:Radiation, Ionizing; D025521:Tumor Suppressor Proteins / Q000032:analysis* / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases / Q000032:analysis* / Q000378:metabolism; D014466:Ultraviolet Rays
D019313:BRCA1 Protein; D007976:Leupeptins; D034741:RNA, Small Interfering; D025521:Tumor Suppressor Proteins; D005047:Etoposide; C102449:BARD1 protein, human; D044767:Ubiquitin-Protein Ligases; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde; D002166:Camptothecin