DNA Cross-Link Repair Protein SNM1A Interacts with PIAS1 in Nuclear Focus Formation

Abstract

The yeast SNM1/PSO2 gene specifically functions in DNA interstrand cross-link (ICL) repair, and its role has been suggested to be separate from other DNA repair pathways. In vertebrates, there are three homologs of SNM1 (SNM1A, SNM1B, and SNM1C/Artemis; SNM1 family proteins) whose functions are largely unknown. We disrupted each of the SNM1 family genes in the chicken B-cell line DT40. Both SNM1A- and SNM1B-deficient cells were sensitive to cisplatin but not to X-rays, whereas SNM1C/Artemis-deficient cells exhibited sensitivity to X-rays but not to cisplatin. SNM1A was nonepistatic with XRCC3 (homologous recombination), RAD18 (translesion synthesis), FANCC (Fanconi anemia), and SNM1B in ICL repair. SNM1A protein formed punctate nuclear foci depending on the conserved SNM1 (metallo-β-lactamase) domain. PIAS1 was found to physically interact with SNM1A, and they colocalized at nuclear foci. Point mutations in the SNM1 domain, which disrupted the interaction with PIAS1, led to mislocalization of SNM1A in the nucleus and loss of complementation of snm1a cells. These results suggest that interaction between SNM1A and PIAS1 is required for ICL repair.

We thank Ryo Goitsuka (Science University of Tokyo) for λZAPII- and pJG4.5-DT40 cDNA libraries used in cDNA cloning and two-hybrid screening, respectively; Hideki Koyama (Yokohama City University) for LIG4-deficient DT40 cells; Tamotsu Nishida (Tokyo University of Pharmacy and Life Science) for the EGFP-hPIAS1 expression plasmid and helpful comments; Yoshinari Imajo and Jyuichi Kubota for irradiating cells with the linear accelerator; Taichi Shirao (Leica Microsystems), Kuniko Asahara, and Hiroyuki Kitao for advice with immunofluorescence and confocal microscopies; Sohsuke Seki for technical help; Mayu Fujii and Kazuko Hikasa for secretarial assistance; and Peter J. McHugh and Ian D. Hickson for critically reading the manuscript.

This work was supported in part by grants from the Ministry of Education, Culture, Sports, Science and Technology (M.I. and M.T.); by grants from the Uehara Memorial Foundation, the Okayama Medical Foundation, and the Kawasaki Medical and Medical Welfare Foundation (M.I.); and by grants from the Japan Space Forum (M.T.). Financial support also came from the Kawasaki Medical School (Project Research Grant 14-409).