Lif1p targets the DNA ligase Lig4p to sites of DNA double-strand breaks.
DNA ligases catalyse the joining of DNA single- and double-strand breaks. Saccharomyces cerevisiae Cdc9p is a homologue of mammalian DNA ligase I and is required for DNA replication, recombination and single-strand break repair. The other yeast ligase, Lig4p/Dnl4p, is a homologue of mammalian DNA ligase IV, and functions in the non-homologous end-joining (NHEJ) pathway of DNA double-strand break repair [1] [2] [3] [4]. Lig4p interacts with Lif1p, the yeast homologue of the human ligase IV-associated protein, XRCC4 [5]. This interaction takes place through the carboxy-terminal domain of Lig4p and is required for Lig4p stability. We show that the carboxy-terminal interaction region of Lig4p is necessary for NHEJ but, when fused to Cdc9p, is insufficient to confer NHEJ function to Cdc9p. Also, Lif1p stimulates the in vitro catalytic activity of Lig4p in adenylation and DNA ligation. Nevertheless, Lig4p is inactive in NHEJ in the absence of Lif1p in vivo, even when Lig4p is stably expressed. We show that Lif1p binds DNA in vitro and, through in vivo cross-linking and chromatin immuno precipitation assays, demonstrate that it targets Lig4p to chromosomal DNA double-strand breaks. Furthermore, this targeting requires another key NHEJ protein, Ku.
Teo
S H
SH
The Wellcome Trust and Cancer Research Campaign, Institute of Cancer and Developmental Biology, University of Cambridge, Cambridge, CB2 1QR, UK.
Jackson
S P
SP
D034961:Antigens, Nuclear; D004249:DNA Damage; D004265:DNA Helicases; D000072481:DNA Ligase ATP; D011088:DNA Ligases / Q000378:metabolism*; D004260:DNA Repair; D004271:DNA, Fungal / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D000072200:Ku Autoantigen; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins
D034961:Antigens, Nuclear; D004271:DNA, Fungal; D004268:DNA-Binding Proteins; C000607891:DNL4 protein, S cerevisiae; C114091:LIF1 protein, S cerevisiae; C000607892:LIG1 protein, human; C000607893:LIG4 protein, human; D009687:Nuclear Proteins; D029701:Saccharomyces cerevisiae Proteins; C081647:high affinity DNA-binding factor, S cerevisiae; D004265:DNA Helicases; C106893:XRCC5 protein, human; C000606509:Xrcc6 protein, human; D000072200:Ku Autoantigen; D011088:DNA Ligases; D000072481:DNA Ligase ATP