Two modules in the BRC repeats of BRCA2 mediate structural and functional interactions with the RAD51 recombinase.
The breast and ovarian cancer suppressor protein BRCA2 controls the RAD51 recombinase in reactions that lead to homologous DNA recombination (HDR). BRCA2 binds RAD51 via eight conserved BRC repeat motifs of approximately 35 amino acids, each with a varying capacity to bind RAD51. BRC repeats both promote and inhibit RAD51 assembly on different DNA substrates to regulate HDR, but the structural basis for these functions is unclear. Here, we demarcate two tetrameric clusters of hydrophobic residues in the BRC repeats, interacting with distinct pockets in RAD51, and show that the co-location of both modules within a single BRC repeat is necessary for BRC-RAD51 binding and function. The two modules comprise the sequence FxxA, known to inhibit RAD51 assembly by blocking the oligomerization interface, and a previously unrecognized tetramer with the consensus sequence LFDE, which binds to a RAD51 pocket distinct from this interface. The LFDE motif is essential in BRC repeats for modes of RAD51 binding both permissive and inhibitory to RAD51 oligomerization. Targeted insertion of point mutations in RAD51 that disrupt the LFDE-binding pocket impair its assembly at DNA damage sites in living cells. Our findings suggest a model for the modular architecture of BRC repeats that provides fresh insight into the mechanisms regulating homologous DNA recombination.
Rajendra
Eeson
E
The Medical Research Council Cancer Cell Unit, Hutchison/MRC Research Centre, Cambridge, UK.
Venkitaraman
Ashok R
AR
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D051017:Apoptosis Regulatory Proteins; D024682:BRCA2 Protein / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites; D002460:Cell Line; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D008958:Models, Molecular; D008969:Molecular Sequence Data; D051135:Rad51 Recombinase / Q000737:chemistry / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D020449:Repetitive Sequences, Amino Acid; D016415:Sequence Alignment
D051017:Apoptosis Regulatory Proteins; C118024:BLID protein, human; D024682:BRCA2 Protein; C551750:BRCA2 protein, human; D011993:Recombinant Fusion Proteins; C495153:RAD51 protein, human; D051135:Rad51 Recombinase