CDK-dependent phosphorylation of BRCA2 as a regulatory mechanism for recombinational repair

Abstract

Inherited mutations in BRCA2 are associated with a predisposition to early-onset breast cancers. The underlying basis of tumorigenesis is thought to be linked to defects in DNA double-strand break repair by homologous recombination. Here we show that the carboxy-terminal region of BRCA2, which interacts directly with the essential recombination protein RAD51, contains a site (serine 3291; S3291) that is phosphorylated by cyclin-dependent kinases. Phosphorylation of S3291 is low in S phase when recombination is active, but increases as cells progress towards mitosis. This modification blocks C-terminal interactions between BRCA2 and RAD51. However, DNA damage overcomes cell cycle regulation by decreasing S3291 phosphorylation and stimulating interactions with RAD51. These results indicate that S3291 phosphorylation might provide a molecular switch to regulate RAD51 recombination activity, providing new insight into why BRCA2 C-terminal deletions lead to radiation sensitivity and cancer predisposition.

XRCC4 in G1 suppresses homologous recombination in S/G2, in G1 checkpoint-defective cells

“…As HR does not act in G1, and NHEJ has been proposed to act throughout the cell cycle (Saintigny et al, 2001;Rothkamm et al, 2003;Saleh-Gohari and Helleday, 2004;Esashi et al, 2005), the simplest explanation was that the HR over-stimulation reflected the defect in NHEJ in S and G2 phases. Surprisingly, we show here that, (1) an NHEJ defect in G1, but not in G2, stimulated IR-induced HR; (2) the cell cycle restriction for HR, that is in S/G2 phase, was not affected by the NHEJ defect; (3) the absence of delay at the G1/S transition was essential for the overstimulation of IR-induced HR in XRCC4 À cells; this allowed the presence in S/G2 of unrepaired DSB generated in G1.…”

“…HR appears to be active from mid-S to G2 as attested by the fact that: (i) HR-defective vertebrate cells are more sensitive when irradiated in the late S/G2 phase (Cheong et al, 1994;Takata et al, 1998;Rothkamm et al, 2003;Hinz et al, 2005); (ii) Rad51 foci do not occur in G1 (Yuan et al, 2003;Aten et al, 2004); (iii) phosphorylation by cyclin-dependent kinase of BRCA2, an essential factor for Rad51 foci assembly, consistently disrupts BRCA2-Rad51 interaction only in G1 phase (Esashi et al, 2005); (iv) conservative HR has been proposed to occur preferentially in S phase (SalehGohari and Helleday, 2004). However, one study challenged such interpretations as it found Rad51 foci in G1 (Kim et al, 2005).…”

Minding the gap: The underground functions of BRCA1 and BRCA2 at stalled replication forks

“…Our current knowledge of the Rad51 loading mechanism in multicellular organisms suggests that it is mediated by a complex set of protein-protein and protein-DNA interactions, orchestrated by Atm and Atr signaling and perhaps by more diverse cell signaling pathways [147]. By analogy with yeast, there may also be negative regulators, operating to inhibit Rad51 loading or disassemble Rad51 nucleoprotein filaments and suppress inappropriate recombination events [148].…”

The checkpoint kinases Chk1 and Chk2 regulate the functional associations between hBRCA2 and Rad51 in response to DNA damage

“…Phosphorylation of S3291 is low in S phase when recombination is active, but increases as cells progress toward mitosis. Phosphorylation of this residue impedes the interactions between Rad51 and the carboxyterminal domain of hBRCA2 (Esashi et al, 2005).…”