ABSTRACT
snm1 mutants of Saccharomyces cerevisiae have been shown to be specifically sensitive to DNA interstrand crosslinking agents but not sensitive to monofunctional alkylating agents, UV, or ionizing radiation. Five homologs of SNM1 have been identified in the mammalian genome and are termed SNM1, SNM1B, Artemis, ELAC2, and CPSF73. To explore the functional role of human Snm1 in response to DNA damage, we characterized the cellular distribution and dynamics of human Snm1 before and after exposure to DNA-damaging agents. Human Snm1 was found to localize to the cell nucleus in three distinct patterns. A particular cell showed diffuse nuclear staining, multiple nuclear foci, or one or two larger bodies confined to the nucleus. Upon exposure to ionizing radiation or an interstrand crosslinking agent, the number of cells exhibiting Snm1 bodies was reduced, while the population of cells with foci increased dramatically. Indirect immunofluorescence studies also indicated that the human Snm1 protein colocalized with 53BP1 before and after exposure to ionizing radiation, and a physical interaction was confirmed by coimmunoprecipitation assays. Furthermore, human Snm1 foci formed after ionizing radiation were largely coincident with foci formed by human Mre11 and to a lesser extent with those formed by BRCA1, but not with those formed by human Rad51. Finally, we mapped a region of human Snm1 of approximately 220 amino acids that was sufficient for focus formation when attached to a nuclear localization signal. Our results indicate a novel function for human Snm1 in the cellular response to double-strand breaks formed by ionizing radiation.
Hypersensitivity of Brca1-deficient MEF to the DNA interstrand crosslinking agent mitomycin C is associated with defect in homologous recombination repair and aberrant S-phase arrest