Abstract
DNA double-strand breaks are repaired through two different pathways, homologous recombination (HR) and non-homologous end-joining (NHEJ). Yeast Xrs2, a homologue of human Nbs1, is a component of the Mre11-Rad50-Xrs2 (MRX) complex required for both HR and NHEJ. Xrs2/Nbs1 contains a conserved FHA domain in the N-terminal end, whose function is largely unknown. In this study, we showed that the FHA domain of Xrs2 plays a critical role in NHEJ, but not in HR. The FHA domain specifically interacts with Lif1, a component of the Ligase-IV complex, Dnl4-Nej1-Lif1 (DNL). Lif1 is phosphorylated in vivo, Serine 383 of Lif1 plays an important role for interaction with Xrs2. Our results suggest that the phosphorylation of Lif1 at Serine 383 is recognized by the FHA domain of Xrs2. The Serine 383 of Lif1 is present in a CK2 (Casein kinase-II) phosphorylation motif. We showed that human CK2 phosphorylates yeast Lif1 protein in vitro and this phosphorylation was largely decreased by the substitution of Serine 383 of Lif1. Interestingly, the interaction between Xrs2 and Lif1 through the FHA domain is conserved for human Nbs1 and Xrcc4, a Lif1 homologue of human. We propose that Nbs1, possibly MRN complex (Mre11-Rad50-Nbs1), is also required for NHEJ in mammalian cells as in yeast. Our results could explain the immuno-deficiency of Nijmigen breakage syndrome (NBS) patients and NBS-variant, which is caused by the dysfunction of Ligase-IV.
