Rapid assessment of repair of ultraviolet DNA damage with a modified host-cell reactivation assay using a luciferase reporter gene and correlation with polymorphisms of DNA repair genes in normal human lymphocytes
Rapid assessment of repair of ultraviolet DNA damage with a modified host-cell reactivation assay using a luciferase reporter gene and correlation with polymorphisms of DNA repair genes in normal human lymphocytes
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DOI：
10.1016/s0027-5107(02)00219-1
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发表时间：
2002-11-30
影响因子：
2.3
通讯作者：
Wei, QY
中科院分区：
文献类型：
--
作者：
Qiao, YW;Spitz, MR;Wei, QY
由于 DNA 修复在癌症遗传易感性中发挥着重要作用，因此 DNA 修复表型的评估对于癌症的分子流行病学研究至关重要。在本报告中，我们比较了在修复 DNA 紫外线 (UV) 损伤的宿主细胞再激活 (HCR) (LUC) 测定中使用荧光素酶 (luc) 报告基因与我们之前用于病例对照研究的基于氯霉素 (cat) 基因的 HCR (CAT) 测定。我们对来自 102 名健康非西班牙裔白人受试者的冷冻淋巴细胞进行了这两项检测。 LUC 和 CAT 测定所测量的 DNA 修复能力 (DRC) 之间存在密切相关性。尽管这两种检测具有相似的变化，但 LUC 检测更快、更灵敏。我们还分析了 DRC 与受试者先前确定的基因型之间的关系，包括两个核苷酸切除修复 (NER) 基因（着色性干皮病 (XP) C 的内含子 9 和 XPD 的外显子 6、10 和 23）的四种多态性以及 X 射线互补组 1 (XRCC1) 的外显子 10 中的碱基切除修复基因的一种多态性。两个 NER 基因的一种或多种多态性纯合受试者的 DRC 显着低于具有其他基因型的受试者 (P = 0.010)。相比之下，多态性 XRCC1 等位基因对 DRC 没有显着影响。这些结果表明，UV 后 LUC 测定可测量 NER 表型，并且 XPC 和 XPD 基因的多态性调节 DRC。对于 DNA 修复表型的群体研究，需要评估许多样本，因此 LUC 测定比 CAT 测定有几个优点：LUC 测定更灵敏、变异更小、无放射性、更容易执行、需要的冻存细胞更少。这些特征使得基于 LUC 的 HCR 检测适用于分子流行病学研究。 (C) 2002 Elsevier Science B.V. 保留所有权利。
As DNA repair plays an important role in genetic susceptibility to cancer, assessment of the DNA repair phenotype is critical for molecular epidemiological studies of cancer. In this report, we compared use of the luciferase (luc) reporter gene in a host-cell reactivation (HCR) (LUC) assay of repair of ultraviolet (UV) damage to DNA to use of the chloramphenicol (cat) gene-based HCR (CAT) assay we used previously for case-control studies. We performed both the assays on cryopreserved lymphocytes from 102 healthy non-Hispanic white subjects. There was a close correlation between DNA repair capacity (DRC) as measured by the LUC and CAT assays. Although these two assays had similar variation, the LUC assay was faster and more sensitive. We also analyzed the relationship between DRC and the subjects' previously determined genotypes for four polymorphisms of two nucleotide-excision repair (NER) genes (in intron 9 of xeroderma pigmentosum (XP) C and exons 6, 10 and 23 of XPD) and one polymorphism of a base-excision repair gene in exon 10 of X-ray complementing group 1 (XRCC1). The DRC was significantly lower in subjects homozygous for one or more polymorphisms of the two NER genes than in subjects with other genotypes (P = 0.010). In contrast, the polymorphic XRCC1 allele had no significant effect on DRC. These results suggest that the post-UV LUC assay measures NER phenotype and that polymorphisms of XPC and XPD genes modulate DRC. For population studies of the DNA repair phenotype, many samples need to be evaluated, and so the LUC assay has several advantages over the CAT assay: the LUC assay was more sensitive, had less variation, was not radioactive, was easier to perform, and required fewer cryopreserved cells. These features make the LUC-based HCR assay suitable for molecular epidemiological studies. (C) 2002 Elsevier Science B.V. All rights reserved.