Barley R D, Enns L, Paterson M C, Mirzayans R
Department of Oncology, University of Alberta, Cross Cancer Institute, Edmonton, Canada.
Oncogene. 1998 Aug 6;17(5):533-43. doi: 10.1038/sj.onc.1202271.
The purpose of this study is to better understand the roles of the p53 tumor suppressor protein and the product of the p53-regulated gene p21WAF1 in the response of diploid human dermal fibroblast cultures to 254 nm ultraviolet (UV) light. We report that Li-Fraumeni syndrome (LFS) fibroblast strains heterozygous for TP53 mutation at either codon 245 or 234 exhibit markedly reduced or no expression of p21WAF1 following UV irradiation, respectively. These strains also exhibit defective nucleotide excision repair and pronounced inhibition of RNA synthesis following UV exposure, both of which are molecular hallmarks of cells derived from patients with the UV-sensitive syndrome xeroderma pigmentosum. In sharp contrast to xeroderma pigmentosum cells, however, the repair-deficient LFS cells show abnormal resistance, rather than hypersensitivity, to the killing effect of UV light. We further demonstrate that exposure of normal human fibroblasts to biologically relevant fluences (< or = 15 J/m2) of UV does not induce apoptotic cell death, indicating that UV resistant phenotype displayed by LFS strains is not associated with deregulated apoptosis. In normal fibroblasts, such treatment results in a moderate ( threefold) up-regulation of p53 protein, induction of the p21WAF1 gene, and a senescence-like growth arrest. On the other hand, exposure to > or = 20 J/m2 UV results in a striking up-regulation of p53, inhibition of p21WAF1 expression, and activation of an apoptotic pathway. We conclude that: (i) p21WAF1-mediated senescence is the principal mode of cell death induced by < or = 15 J/m2 UV light in normal human fibroblasts; (ii) there is a threshold effect for p53-dependent apoptosis and that, in normal human cells, this threshold level is induced upon expsoure to 20 J/m2 UV; (iii) the p53 signaling pathway is malfunctional in the TP53 heterozygous LFS strains examined; and (iv) the enhanced resistance to UV-induced cell killing displayed by these LFS strains is a consequence of diminished growth arrest, which is presumably mediated by p21WAF1 and not abnormalities in an apoptotic pathway.
本研究的目的是更深入地了解p53肿瘤抑制蛋白以及p53调控基因p21WAF1的产物在二倍体人皮肤成纤维细胞培养物对254nm紫外线（UV）照射的反应中所起的作用。我们报告，在密码子245或234处TP53发生突变的杂合型李-弗劳梅尼综合征（LFS）成纤维细胞系，在紫外线照射后分别表现出p21WAF1表达显著降低或无表达。这些细胞系在紫外线照射后还表现出核苷酸切除修复缺陷以及RNA合成受到明显抑制，这两者都是紫外线敏感综合征色素性干皮病患者来源细胞的分子特征。然而，与色素性干皮病细胞形成鲜明对比的是，修复缺陷的LFS细胞对紫外线的杀伤作用表现出异常抗性，而非超敏性。我们进一步证明，正常人成纤维细胞暴露于生物相关剂量（≤15 J/m2）的紫外线不会诱导凋亡性细胞死亡，这表明LFS细胞系所表现出的抗紫外线表型与凋亡失调无关。在正常成纤维细胞中，这种处理会导致p53蛋白适度（三倍）上调、p21WAF1基因诱导以及类似衰老的生长停滞。另一方面，暴露于≥20 J/m2的紫外线会导致p53显著上调、p21WAF1表达受到抑制以及凋亡途径激活。我们得出以下结论：（i）p21WAF1介导的衰老为正常人成纤维细胞中≤15 J/m2紫外线诱导的细胞死亡的主要模式；（ii）p53依赖性凋亡存在阈值效应，在正常人细胞中，该阈值水平在暴露于20 J/m2紫外线时被诱导；（iii）在所检测的TP53杂合型LFS细胞系中，p53信号通路功能失调；（iv）这些LFS细胞系对紫外线诱导的细胞杀伤增强的抗性是生长停滞减弱的结果，这可能是由p21WAF1介导的，而非凋亡途径异常所致。
