Mutant yeast strains lacking the silencing proteins Sir2p, Sir3p, or Sir4p have a defect in a DNA double-strand break (DSB) repair pathway, called nonhomologous end joining (NHEJ). Mutations in sir genes also lead to the simultaneous expression of a and alpha mating type information, thus generating a nonmating haploid cell type with many properties shared with a/alpha diploids. We addressed whether cell type or Sir proteins per se regulate NHEJ by investigating the role of a novel haploid-specific gene in NHEJ. This gene, NEJ1, was required for efficient NHEJ, and transcription of NEJ1 was completely repressed in a/alpha diploid and sir haploid strains. The NEJ1 promoter contained a consensus binding site for the a1/alpha2 repressor, explaining the cell type-specific expression. Expression of Nej1p from a constitutive promoter in a/alpha diploid and sir mutant strains completely rescued the defect in NHEJ, thus showing that Sir proteins per se were dispensable for NHEJ. Nej1p and Lif1(P), the yeast XRCC4 homolog, interacted in two independent assays, and Nej1p localized to the nucleus, suggesting that Nej1p may have a direct role in NHEJ.