Mirzayans R, Paterson M C
Department of Medicine, Cross Cancer Institute, Edmonton, Alb., Canada.
Mutat Res. 1991 Jul;255(1):57-65. doi: 10.1016/0921-8777(91)90018-k.
The extent of DNA excision repair was determined in dermal fibroblast strains from clinically normal and xeroderma pigmentosum (XP; complementation group A) human donors after single or combined exposures to 254-nm ultraviolet light and 4-nitroquinoline 1-oxide (4NQO). The repair was monitored by incubation of the treated cultures in the presence of 1-beta-D-arabinofuranosylcytosine (araC), a potent inhibitor of long-patch excision repair, followed by quantitation of araC-accumulated DNA single-strand breaks (representing repair events) by velocity sedimentation analysis in alkaline sucrose gradients. The amount of repair in normal fibroblast strains increased as a function of UV fluence and reached a plateau at 15 J/m2; strand breaks were not detected when these same cultures were irradiated with as much as 60 J/m2 UV and incubated in the absence of araC, implying that an initial (incision) step is rate-limiting in the repair of UV damage. In normal fibroblasts (i) the incidence of araC-detectable lesions removed during fixed intervals following exposure to 4NQO (4 microM; 30 min) was approximately 2.5 times greater than that seen following irradiation with repair-saturating fluences (greater than or equal to 15 J/m2) of UV-rays; and (ii) the amount of repair in cultures treated simultaneously with 4NQO (0.5-6 microM; 30 min) and a repair-saturating fluence of UV (20 J/m2) was found to approach the sum of that arising from exposure to each separately. The XP cells (XP12BE) exhibited a deficiency in the removal of araC-detectable DNA lesions following exposure to either of the carcinogens. Since araC is known to inhibit the repair of alkali-stable 4NQO-DNA adducts (i.e., lesions assumed to be removed by the UV-like excision pathway) but not that of alkali-labile sites (i.e., DNA lesions operated on by the X-ray-like repair pathway), our results strongly imply that the multistep excision-repair pathway operative on UV photoproducts in human fibroblasts differs from that responsible for removing alkali-stable (araC-detectable) 4NQO adducts by at least one step, presumably the rate-limiting incision reaction mediated by a lesion-recognizing endonuclease.
在来自临床正常人和着色性干皮病（XP；互补组A）患者的皮肤成纤维细胞株中，测定了在单独或联合暴露于254纳米紫外线和4-硝基喹啉1-氧化物（4NQO）后DNA切除修复的程度。通过在强效长片段切除修复抑制剂1-β-D-阿拉伯呋喃糖基胞嘧啶（araC）存在的情况下孵育处理后的培养物来监测修复过程，随后通过碱性蔗糖梯度中的速度沉降分析对araC积累的DNA单链断裂（代表修复事件）进行定量。正常成纤维细胞株中的修复量随紫外线通量增加而增加，并在15 J/m²时达到平台期；当用高达60 J/m²的紫外线照射这些相同的培养物并在无araC的情况下孵育时，未检测到链断裂，这意味着在紫外线损伤修复中初始（切割）步骤是限速的。在正常成纤维细胞中：（i）暴露于4NQO（4 microM；30分钟）后在固定间隔内可被araC检测到的损伤去除率比用饱和修复通量（大于或等于15 J/m²）的紫外线照射后高约2.5倍；（ii）同时用4NQO（0.5 - 6 microM；30分钟）和饱和修复通量的紫外线（20 J/m²）处理的培养物中的修复量接近分别暴露于每种致癌物所产生的修复量之和。XP细胞（XP12BE）在暴露于任何一种致癌物后，在去除可被araC检测到的DNA损伤方面表现出缺陷。由于已知araC可抑制碱稳定的4NQO-DNA加合物（即假定通过类似紫外线的切除途径去除的损伤）的修复，但不抑制碱不稳定位点（即通过类似X射线的修复途径处理的DNA损伤）的修复，我们的结果强烈表明，在人成纤维细胞中作用于紫外线光产物的多步骤切除修复途径与负责去除碱稳定（可被araC检测到）的4NQO加合物的途径至少在一个步骤上不同，大概是由损伤识别内切酶介导的限速切割反应。
