title: Modes of interaction among yeast Nej1, Lif1 and Dnl4 proteins and comparison to human XLF, XRCC4 and Lig4.
abstract: The nonhomologous end joining (NHEJ) pathway of double-strand break repair depends on DNA ligase IV and its interacting partner protein XRCC4 (Lif1 in yeast). A third yeast protein, Nej1, interacts with Lif1 and supports NHEJ, similar to the distantly related mammalian Nej1 orthologue XLF (also known as Cernunnos). XRCC4/Lif1 and XLF/Nej1 are themselves related and likely fold into similar coiled-coil structures, which suggests many possible modes of interaction between these proteins. Using yeast two-hybrid and co-precipitation methods we examined these interactions and the protein domains required to support them. Results suggest that stable coiled-coil homodimers are a predominant form of XLF/Nej1, just as for XRCC4/Lif1, but that similar heterodimers are not. XLF-XRCC4 and Nej1-Lif1 interactions were instead mediated independently of the coiled coil, and by different regions of XLF and Nej1. Specifically, the globular head of XRCC4/Lif1 interacted with N- and C-terminal domains of XLF and Nej1, respectively. Direct interactions between XLF/Nej1 and DNA ligase IV were also observed, but again appeared qualitatively different than the stable coiled-coil-mediated interaction between XRCC4/Lif1 and DNA ligase IV. The implications of these findings for DNA ligase IV function are considered in light of the evolutionary pattern in the XLF/XRCC4 and XLF/Nej1 family.
authors.affiliation: Department of Pathology, University of Michigan Medical School, Room 2065 BSRB, 109 Zina Pitcher Place, Ann Arbor, MI 48109-2200, USA.
authors.forename: Rajashree A
authors.initials: RA
authors.lastname: Deshpande
authors.forename: Thomas E
authors.initials: TE
authors.lastname: Wilson
mesh_terms: D000072481:DNA Ligase ATP; D011088:DNA Ligases / Q000378:metabolism*; D045643:DNA Repair Enzymes / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D019281:Dimerization; D006801:Humans; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*; D020798:Two-Hybrid System Techniques
chemical_list: D004268:DNA-Binding Proteins; C000607891:DNL4 protein, S cerevisiae; C114091:LIF1 protein, S cerevisiae; C000607893:LIG4 protein, human; C441270:NEJ1 protein, S cerevisiae; C508551:NHEJ1 protein, human; D029701:Saccharomyces cerevisiae Proteins; C101467:XRCC4 protein, human; D011088:DNA Ligases; D045643:DNA Repair Enzymes; D000072481:DNA Ligase ATP