MOLECULAR ANALYSIS OF CXPD MUTATIONS IN THE REPAIR-DEFICIENT HAMSTER MUTANTS UV5 AND UVL-13

The cDNA sequence of the Chinese hamster xeroderma pigmentosum group D (CXPD) nucleotide excision repair gene was analyzed from three Chines e hamster ovary (CHO) cell lines: repair proficient strain AA8 and rep air deficient, UV complementation group 2 strains UV5 and UVL-13. CXPD encodes a presumed ATP-dependent DNA helicase and is single copy in C HO lines due to the hemizygosity of chromosome 9. Comparison of the de duced wild-type AA8 CXPD protein sequence with that of the Chinese ham ster V79 lung-derived cell line revealed two amino acid polymorphisms. Position 285 is glutamine in AA8 and arginine in V79, and position 29 8 is alanine in AA8 and threonine in V79. Comparison with the human XP D, Saccharomyces cerevisiae RAD3, and Schizosaccharomyces pombe rad15 homologs shows variability at these positions. Analysis of the CXPD se quence in the repair deficient CHO lines UV5 and UVL-13 revealed, in e ach case, a single base substitution resulting in an amino acid substi tution. Position 116 is tyrosine in UV5 and cysteine in AA8, and the c orresponding positions of XPD, RADS, and rad15 are cysteine. Position 615 is glutamic acid in UVL-13 and glycine in AA8, and the correspondi ng positions of XPD, RAD3, and rad15 are glycine. In both UV5 and UVL- 13, positions 285 and 298 are glutamine and alanine, respectively, as seen in AA8. These results suggest that cysteine 116 and glycine 615 a re critical to the repair function of CXPD.