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.

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.