Purified human BRCA2 stimulates RAD51-mediated recombination.
Mutation of the breast cancer susceptibility gene, BRCA2, leads to breast and ovarian cancers. Mechanistic insight into the functions of human BRCA2 has been limited by the difficulty of isolating this large protein (3,418 amino acids). Here we report the purification of full-length BRCA2 and show that it both binds RAD51 and potentiates recombinational DNA repair by promoting assembly of RAD51 onto single-stranded DNA (ssDNA). BRCA2 acts by targeting RAD51 to ssDNA over double-stranded DNA, enabling RAD51 to displace replication protein-A (RPA) from ssDNA and stabilizing RAD51-ssDNA filaments by blocking ATP hydrolysis. BRCA2 does not anneal ssDNA complexed with RPA, implying it does not directly function in repair processes that involve ssDNA annealing. Our findings show that BRCA2 is a key mediator of homologous recombination, and they provide a molecular basis for understanding how this DNA repair process is disrupted by BRCA2 mutations, which lead to chromosomal instability and cancer.
Jensen
Ryan B
RB
Department of Microbiology, University of California, Davis, California 95616, USA.
Carreira
Aura
A
Kowalczykowski
Stephen C
SC
D020816:Amino Acid Motifs; D051017:Apoptosis Regulatory Proteins; D024682:BRCA2 Protein / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D018797:Cell Cycle Proteins / Q000378:metabolism; D002460:Cell Line; D043171:Chromosomal Instability; D004247:DNA / Q000737:chemistry / Q000378:metabolism; D004260:DNA Repair; D004277:DNA, Single-Stranded / Q000737:chemistry / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D006801:Humans; D009154:Mutation; D011485:Protein Binding; D051135:Rad51 Recombinase / Q000378:metabolism*; D011995:Recombination, Genetic; D051716:Replication Protein A / Q000378:metabolism; D012689:Sequence Homology, Nucleic Acid; D013379:Substrate Specificity
D051017:Apoptosis Regulatory Proteins; C118024:BLID protein, human; D024682:BRCA2 Protein; C551750:BRCA2 protein, human; D018797:Cell Cycle Proteins; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D051716:Replication Protein A; D004247:DNA; C495153:RAD51 protein, human; D051135:Rad51 Recombinase; C074556:DMC1 protein, human