Mullenders L H, Hazekamp-van Dokkum A M, Kalle W H, Vrieling H, Zdzienicka M Z, van Zeeland A A
MGC-Department of Radiation Genetics and Chemical Mutagenesis, University of Leiden, The Netherlands.
Mutat Res. 1993 May;299(3-4):271-6. doi: 10.1016/0165-1218(93)90103-k.
UV-induced cyclobutane pyrimidine dimers (CPD) are selectively removed from the transcribed strand of transcriptionally active genes in V79 Chinese hamster cells. This strand specificity of repair corresponds well with the observation that UV-induced mutations in the HPRT gene are primarily generated by DNA photolesions in the non-transcribed strand. This strand bias for mutations is, however, much more pronounced at 2 J/m2 than at the higher dose of 12 J/m2. An alternative explanation for strand specificity of mutations would be that most of the mutations are caused by pyrimidone 6-4 pyrimidine photoproducts (6-4 PP). Indeed experiments with a V79-derived cell line capable of repairing 6-4 PP but not CPD have revealed direct evidence for 6-4 PP as the mutagenic lesions in UV-irradiated hamster cells. This implies that 6-4 PP are also preferentially repaired in the transcribed strand. We have investigated the repair of DNA photolesions in the HPRT gene by measuring the distribution of bromodeoxyuridine-labeled repair patches in the transcribed and non-transcribed strands of genes employing a newly developed immunoextraction procedure. Three cell lines with different capacities to remove CPD and 6-4 PP from the HPRT gene and from the genome overall were used. We found no evidence for preferential repair of 6-4 PP in the transcribed strand of the HPRT gene in cells exposed to 10 J/m2. These data are in favor of a lack of strand-specific repair of 6-4 PP underlying the much less pronounced strand bias for induced mutations at high UV dose. However, the conclusive test would be the demonstration of preferential repair of 6-4 PP in the transcribed strand of transcriptionally active genes in cells exposed to 2 J/m2.
紫外线诱导的环丁烷嘧啶二聚体（CPD）在V79中国仓鼠细胞中转录活跃基因的转录链上被选择性去除。这种修复的链特异性与以下观察结果非常吻合：次黄嘌呤-鸟嘌呤磷酸核糖转移酶（HPRT）基因中的紫外线诱导突变主要由非转录链中的DNA光损伤产生。然而，这种突变的链偏向性在2 J/m²时比在12 J/m²的更高剂量时更为明显。对突变链特异性的另一种解释是，大多数突变是由嘧啶酮6-4嘧啶光产物（6-4 PP）引起的。事实上，对一种能够修复6-4 PP但不能修复CPD的V79衍生细胞系进行的实验揭示了6-4 PP作为紫外线照射仓鼠细胞中诱变损伤的直接证据。这意味着6-4 PP也优先在转录链中得到修复。我们通过使用新开发的免疫提取程序测量基因转录链和非转录链中溴脱氧尿苷标记的修复片段的分布，研究了HPRT基因中DNA光损伤的修复情况。使用了三种对从HPRT基因和整个基因组中去除CPD和6-4 PP具有不同能力的细胞系。我们没有发现暴露于10 J/m²的细胞中HPRT基因转录链中6-4 PP优先修复的证据。这些数据支持在高紫外线剂量下诱导突变的链偏向性不太明显的情况下，6-4 PP缺乏链特异性修复。然而，决定性的测试将是证明暴露于2 J/m²的细胞中转录活跃基因转录链中6-4 PP的优先修复。
