Repair of chromosomal double-strand breaks by precise ligation in human cells

Abstract

Double-strand breaks (DSBs), a common type of DNA lesion, occur daily in human cells as a result of both endogenous and exogenous damaging agents. DSBs are repaired in two general ways: by the homology-dependent, error-free pathways of homologous recombination (HR) and by the homology-independent, error-prone pathways of nonhomologous end-joining (NHEJ), with NHEJ predominating in most cells. DSBs with compatible ends can be re-joined in vitro with DNA ligase alone, which raises the question of whether such DSBs require the more elaborate machinery of NHEJ to be repaired in cells. Here we report that chromosomal DSBs with compatible ends introduced by the rare-cutting endonuclease, ISceI, are repaired by precise ligation nearly 100% of the time in human cells. Precise ligation depends on the classical NHEJ components Ku70, XRCC4, and DNA ligase IV, since siRNA knockdowns of these factors significantly reduced the efficiency of precise ligation. Interestingly, knockdown of the tumor suppressors p53 or BRCA1 showed similar effects as the knockdowns of NHEJ factors. In contrast, knockdown of components involved in alternative NHEJ, mismatch repair, nucleotide excision repair, and single-strand break repair did not reduce precise ligation. In summary, our results demonstrate that DSBs in human cells are efficiently repaired by precise ligation, which requires classical NHEJ components and is enhanced by p53 and BRCA1.