Strand specificity of mutagenic bypass replication of DNA containing psoralen monoadducts in a human cell extract.
Strand specificity of mutagenic bypass replication of DNA containing psoralen monoadducts in a human cell extract.
复制标题
人类细胞提取物中含有补骨脂素单加合物的 DNA 的诱变旁路复制的链特异性。
DOI：
10.1128/mcb.16.5.2537
复制
发表时间：
1996
影响因子：
5.3
通讯作者：
Kunkel,TA
中科院分区：
文献类型：
--
作者：
Thomas,DC;Svoboda,DL;Vos,JM;Kunkel,TA
补骨脂素是一种植物来源的致突变化合物，用于治疗各种皮肤病、血细胞癌和自身免疫性疾病的光敏剂。为了研究补骨脂素对人的致突变机制，我们检测了补骨脂素衍生物4‘-羟甲基-4，5’，8-三甲基补骨脂素加UVA照射后，人细胞M13mp2 DNA提取液中猴病毒40的依赖于来源的复制的效率和保真度。不同剂量的4‘-羟甲基-4，5’，8-三甲基补骨脂素和固定的UVA剂量可抑制DNA的复制，且呈浓度依赖关系。然而，观察到共价闭合的单体长度的环状复制产物。用250~280 nm紫外光交联后的复性琼脂糖凝胶电泳法分析表明，在体外复制过程中，大约9个补骨脂素单加合物中有1个被绕过。将产物DNA导入Escherichia Colito Score复制错误的Zareporter基因中，表明受损DNA的复制比未损伤DNA的复制更具诱变性。对突变株的序列分析表明，与损伤相关的复制错误主要是T·A→C·G转换、C·G碱基对的颠倒和单个A·T碱基对的缺失，最后一种错误最常发生在均聚体中。与复制起点不对称地放置在突变靶的相对侧的两种底物的错误特异性的比较表明，滞后链复制设备对于补骨脂素单加合物依赖的缺失错误的准确性低于领先链复制设备。提出了一种基于单加成碱基从链上优先环出的模型，该链是逆行不连续合成的模板。
Psoralens are mutagenic compounds of vegetable origin that are used as photosensitizing agents in the treatment of various skin diseases, blood cell cancer, and autoimmune disorders. To study the mechanism of mutagenicity of psoralens in humans, we examined the efficiency and fidelity of simian virus 40 origin-dependent replication in a human cell extract of M13mp2 DNA randomly treated with the psoralen derivative 4′-hydroxymethyl-4,5′,8-trimethyl psoralen plus UVA irradiation. Replication of DNA treated with variable amounts of 4′-hydroxymethyl-4,5′,8-trimethyl psoralen and a fixed UVA fluence was inhibited in a concentration-dependent manner. However, covalently closed monomer-length circular replication products were observed. Product analysis by renaturing agarose gel electrophoresis after cross-linking with 250- to 280-nm UV light indicated that approximately 1 of 9 psoralen monoadducts was bypassed during in vitro replication. Introduction of product DNA intoEscherichia colito score replication errors in thelacZareporter gene demonstrated that replication of the damaged DNA was more mutagenic than was replication of undamaged DNA. Sequence analysis oflacZmutants revealed that damage-dependent replication errors were predominantly T · A→C · G transitions, transversions at C · G base pairs, and deletions of single A · T base pairs, the last occurring most frequently in homopolymeric runs. A comparison of error specificities with two substrates having the replication origin asymmetrically placed on opposite sides of the mutational target suggests that the lagging-strand replication apparatus is less accurate than the leading-strand replication apparatus for psoralen monoadduct-dependent deletion errors. A model is proposed based on the preferential loopout of the monoadducted base from the strand that templates retrograde discontinuous synthesis.