The breast and ovarian predisposition protein BRCA1 is a required component of the mammalian response to double-stranded DNA damage. Its conserved BRCT domains are required for BRCA1 accumulation to sites of repair, while the conserved N-terminal RING domain is able to catalyse the conjugation of ubiquitin and act as an E3 ubiquitin ligase. Disruption of either of these domains by missense mutation is associated with disease development.
The SUMO conjugation pathway has been implicated in DNA damage response in model organisms, and in Caenorhabditis elegans the Brac1 binding partner Bard1 associates with the SUMO E2 conjugating enzyme Ubc9. In mammalian cells, BRCA1 has been found to be associated with free SUMO-1 resulting in altered transcription.
We undertook to examine the potential influence of the SUMO pathway on BRCA1 response to genotoxic stress.
Using a range of biochemical and cell-biology techniques, we have shown that BRCA1 is modified by SUMO in response to genotoxic stress, and co-localises at sites of DNA damage with SUMO1, SUMO2/3 and the SUMO conjugating enzyme Ubc9. PIAS SUMO E3 ligases co-localise with and modulate SUMO modification of BRCA1, and are required for BRCA1 ubiquitin ligase activity in cells. In vitro SUMO modification of the BRCA1:BARD1 heterodimer greatly increases its ligase activity, identifying it as a SUMO regulated ubiquitin ligase. Further, PIAS SUMO ligases are required for complete accumulation of double-strand DNA damage repair proteins subsequent to RNF8 accrual, and for proficient double-strand break repair. Because the two features of BRCA1 activity regulated by the SUMO pathway, ubiquitin ligase activity and accumulation at sites of DNA damage, are also inhibited by some BRCA1 mutations that predispose to breast cancer and ovarian cancer, it seems highly likely that the SUMO pathway will be of relevance to cancer predisposition and development.
Authors and Affiliations
King's College London, UK
JR Morris, M Keppler, R Densham, D Weekes, A Alamshah, L Butler, L Pangon, T Kiuchi, T Ng & E Solomon