Characteristics of UV-induced mutation spectra in human XP-D/ERCC2 gene-mutated xeroderma pigmentosum and trichothiodystrophy cells.
To study the relationships between mutagenesis and carcinogenesis, we compared the mutations and their frequency induced by ultraviolet irradiation at 254 nm (UV-C) in XP-D (GM-08207B/XP6BE), TTD/XP-D (TTD1VI-LAS-KMT11) and wild-type (MRC-5V1) human cells. XP-D and TTD/XP-D cells, mutated in the same XP-D/ERCC2 gene, are deficient in nucleotide excision repair. Whereas XP-D patients develop early skin tumors, TTD patients do not exhibit abnormal levels of cancers. After verification of UV hypersensitivity and DNA repair defect of the immortalized cell lines XP-D and TTD compared with a wild-type cell line, UV-induced mutagenesis was studied with a new shuttle vector pR2, carrying the target lacZ' gene. The UV-mutation frequencies in XP-D and TTD cells were similar and significantly increased compared with normal cells. Sequence analysis of 312 independent mutant plasmids revealed that more rearrangements were induced in TTD cells (16%) than in XP-D (5%) and normal cells (1%), while XP-D cells exhibited a twofold higher rate of tandem mutations compared with TTD and normal cells. In the three cell lines, a predominance of G:C to A:T transitions was found, especially in chiefly on the cytosine at 5'-TC-3' sites. The types of UV-induced point mutations in TTD cells were, however, more similar to those in normal cells than those found in XP-D cells. XP-D mutations were preferentially located in 5'-TCPur-3' sites, while mutations in normal and TTD cells were mostly at 5'-TCC-3' sites. Analysis of mutation spectra revealed differences in the location of the mutational hotspots between the three lines. Although the mutation frequency of the UV-irradiated pR2 vector is much higher in TTD and XP-D cells than in normal cells, the mutation spectrum is closer between TTD and normal cells as compared with XP-D cells. These dissimilarities could contribute to an explanation of some of the differences between the two syndromes.
Marionnet
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Laboratory of Molecular Genetics, UPR 42 CNRS IFC1, Institut de Recherches sur le Cancer, Villejuif, France.
Benoit
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Benhamou
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Sarasin
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Stary
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D000595:Amino Acid Sequence; D001483:Base Sequence; D002460:Cell Line; D002461:Cell Line, Transformed; D002470:Cell Survival; D004260:DNA Repair / Q000235:genetics*; D005347:Fibroblasts; D005822:Genetic Vectors; D006201:Hair Diseases / Q000235:genetics*; D006801:Humans; D007763:Lac Operon / Q000235:genetics; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009154:Mutation; D017354:Point Mutation; D014466:Ultraviolet Rays; D014983:Xeroderma Pigmentosum / Q000235:genetics*