Wang Y C, Maher V M, McCormick J J
Department of Microbiology, Michigan State University, East Lansing 48824-1316.
Proc Natl Acad Sci U S A. 1991 Sep 1;88(17):7810-4. doi: 10.1073/pnas.88.17.7810.
Xeroderma pigmentosum (XP) variant patients show the clinical characteristics of the disease, with increased frequencies of skin cancer, but their cells have a normal, or nearly normal, rate of nucleotide excision repair of UV-induced DNA damage and are only slightly more sensitive than normal cells to the cytotoxic effect of UV radiation. However, they are significantly more sensitive to its mutagenic effect. To examine the mechanisms responsible for this hypermutability, we transfected an XP variant cell line with a UV-irradiated (at 254 nm) shuttle vector carrying the supF gene as a target for mutations, allowed replication of the plasmid, determined the frequency and spectrum of mutations induced, and compared the results with those obtained previously when irradiated plasmids carrying the same target gene replicated in a normal cell line [Bredberg, A., Kraemer, K. H. & Seidman, M. M. (1986) Proc. Natl. Acad. Sci. USA 83, 8273-8277]. The frequency of mutants increased linearly with dose, but with a slope 5 times steeper than that seen with normal cells. Sequence analysis of the supF gene showed that 52 of 53 independent mutants generated in the XP variant cells contained base substitutions, with 62 of 64 of the substitutions involving a dipyrimidine. Twenty-eight percent of the mutations involved A.T base pairs, with the majority found at position 136, the middle of a run of three A.T base pairs. (In the normal cells, this value was only 11%.) If the rate of excision of lesions from supF in the two cell lines is equal, our data suggest that XP variant cells are less likely than normal cells to incorporate dAMP opposite bases involved in photo-products. If such incorporation also occurs during replication of chromosomal DNA, this could account for the hypermutability of XP variant cells with UV irradiation.
着色性干皮病（XP）变异型患者表现出该疾病的临床特征，皮肤癌发病率增加，但其细胞对紫外线诱导的DNA损伤的核苷酸切除修复率正常或接近正常，并且对紫外线辐射的细胞毒性作用仅比正常细胞稍敏感。然而，它们对紫外线的诱变作用明显更敏感。为了研究导致这种高突变性的机制，我们用携带supF基因作为突变靶点的紫外线（254nm）照射的穿梭载体转染了一个XP变异细胞系，让质粒复制，确定诱导的突变频率和谱，并将结果与之前在正常细胞系中携带相同靶点基因的照射质粒复制时获得的结果进行比较[布雷德伯格，A.，克雷默，K.H.和西德曼，M.M.（1986年）美国国家科学院院刊83，8273 - 8277]。突变体频率随剂量呈线性增加，但斜率比正常细胞陡5倍。对supF基因的序列分析表明，在XP变异细胞中产生的53个独立突变体中有52个包含碱基替换，其中64个替换中有62个涉及二嘧啶。28%的突变涉及A.T碱基对，大多数位于136位，即三个A.T碱基对的中间位置。（在正常细胞中，这个值仅为11%。）如果两个细胞系中从supF切除损伤的速率相等，我们的数据表明，XP变异细胞比正常细胞更不容易在光产物相关碱基对面掺入dAMP。如果这种掺入也发生在染色体DNA复制过程中，这可能解释了XP变异细胞在紫外线照射下的高突变性。
