Rousset S, Nocentini S, Santella R M, Moustacchi E
UMR 218 CNRS, Laboratoire de Recherche Correspondant no. 1 du CEA, Institut Curie, Paris, France.
J Photochem Photobiol B. 1997 Apr;38(2-3):220-7. doi: 10.1016/s1011-1344(96)07461-1.
6,4,4'-Trimethylangelicin (TMA)-photoinduced monoadducts (MAs) were detected and quantified on DNA of normal human and Fanconi's anemia (FA) fibroblasts (complementation groups A and D) by immuno-electron microscopy. TMA-modified DNA was extracted from the cells just after photoreaction, or after a subsequent 24 h repair period, for analysis of the MA processing capabilities of the different cell lines. Unmodified DNA was extracted from the control cells in parallel. The immunoreaction with antibody 7E3 was performed on single-stranded DNA fragments obtained by heat-formamide denaturation. On single-stranded DNA fragments scanned in the electron microscope, IgG-labeled MA sites appeared as isolated or clustered IgG molecules, which were not homogeneously distributed. The isolated IgG and the different clusters (doublets, triplets or near-neighbors (within a distance of 250 nucleotides)) were measured separately for induction frequency and removal. Few interstrand cross-links (CLs) were present on X-shape DNA fragments. At time zero, the distribution patterns of TMA-photoinduced IgG-labeled MA sites and CLs, and their amount per 10(6) nucleotides, were similar in the three cell lines. After the 24 h repair period, FA cells from two different genetic complementation groups demonstrated impaired incision-excision repair capabilities for both MAs (singlets or clusters) and CLs when compared with normal cells. In each cell line, the relative proportions of TMA-induced lesions remaining at time 24 h were similar to those initially induced. This implies analogous processing kinetics towards the TMA-photoinduced clusters of MAs and CLs in a given cell line.
通过免疫电子显微镜检测并定量了6,4,4'-三甲基白芷素（TMA）光诱导的单加合物（MA）在正常人及范可尼贫血（FA）成纤维细胞（互补组A和D）DNA上的情况。在光反应刚结束后或随后24小时修复期后从细胞中提取TMA修饰的DNA，用于分析不同细胞系对MA的处理能力。同时从对照细胞中提取未修饰的DNA。对通过热甲酰胺变性获得的单链DNA片段进行与抗体7E3的免疫反应。在电子显微镜下扫描的单链DNA片段上，IgG标记的MA位点表现为孤立或成簇的IgG分子，分布不均匀。分别测量孤立的IgG和成簇的（双联体、三联体或近邻（在250个核苷酸距离内））的诱导频率和去除情况。在X形DNA片段上存在少量链间交联（CL）。在时间零点，三种细胞系中TMA光诱导的IgG标记的MA位点和CL的分布模式及其每10(6)个核苷酸的数量相似。在24小时修复期后，与正常细胞相比，来自两个不同遗传互补组的FA细胞对MA（单联体或簇）和CL的切口-切除修复能力受损。在每个细胞系中，24小时时残留的TMA诱导损伤的相对比例与最初诱导的比例相似。这意味着在给定细胞系中对TMA光诱导的MA和CL簇的处理动力学相似。
