title: The C-terminus of Nej1 is critical for nuclear localization and non-homologous end-joining.
abstract: Nej1 is an essential factor in the non-homologous end-joining (NHEJ) pathway and interacts with the DNA ligase complex, Lif1-Dnl4, through interactions with Lif1. We have mapped K331-V338 in the C-terminal region of Nej1 to be critical for its functionality during repair. Truncation and alanine scanning mutagenesis have been used to identify a motif in Nej1, KKRK (331-334), which is important for both nuclear targeting and NHEJ repair after localization. We have identified F335-V338 to be important for proper interaction with Lif1, however this region is not required for Nej1 recruitment to HO endonuclease-induced DNA double-strand breaks in vivo. Phenylalanine at position 335 is particularly important for the role of Nej1 in repair and the loss of association between Nej1 and Lif1 correlates with a decrease in cell survival upon either transient or continuous HO expression in nej1 mutants.
authors.affiliation: Department of Biochemistry and Molecular Biology, Southern Alberta Cancer Research Institute, University of Calgary, 3330 Hospital Drive NW, Calgary, AB, Canada T2N 4N1.
authors.forename: Brandi L
authors.initials: BL
authors.lastname: Mahaney
authors.affiliation: Department of Biochemistry and Molecular Biology, Southern Alberta Cancer Research Institute, University of Calgary, 3330 Hospital Drive NW, Calgary, AB, Canada T2N 4N1.
authors.forename: Susan P
authors.initials: SP
authors.lastname: Lees-Miller
authors.affiliation: Department of Biochemistry and Molecular Biology, Southern Alberta Cancer Research Institute, University of Calgary, 3330 Hospital Drive NW, Calgary, AB, Canada T2N 4N1. Electronic address: jcobb@ucalgary.ca.
authors.forename: Jennifer A
authors.initials: JA
authors.lastname: Cobb
mesh_terms: D021581:Active Transport, Cell Nucleus; D002467:Cell Nucleus / Q000378:metabolism*; D004249:DNA Damage; D059766:DNA End-Joining Repair; D000072481:DNA Ligase ATP; D011088:DNA Ligases / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D019913:Nuclear Localization Signals / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
chemical_list: D004268:DNA-Binding Proteins; C114091:LIF1 protein, S cerevisiae; C441270:NEJ1 protein, S cerevisiae; D019913:Nuclear Localization Signals; D029701:Saccharomyces cerevisiae Proteins; D011088:DNA Ligases; D000072481:DNA Ligase ATP
keywords: 4′,6-diamidino-2-phenylindole; C-terminal region; CTR; DAPI; DNA repair; DSBs; Double-strand breaks; Lif1p; NHEJ; Nej1p; Non-homologous end-joining; PBS; UTR; double-strand breaks; non-homologous end-joining; phosphate buffered saline; untranslated region