Venkatachalam S, Denissenko M, Wani A A
Biochemistry Program, The Ohio State University, Columbus 43210, USA.
Oncogene. 1997 Feb 20;14(7):801-9. doi: 10.1038/sj.onc.1200890.
The rapid accumulation of the p53 gene product is considered to be an important component of the cellular response to a variety of genotoxins. In order to gain insights on the biochemical pathways leading to p53 stabilization, the effect of (+/-) 7,8-dihydroxy-anti-9, 10-epoxy-7,8,9,10-tetrahydrobenzo(a)-pyrene [(+/-)-anti-BPDE] induced DNA damage on p53 protein levels was investigated in various repair-proficient and repair-deficient human cells. Brief exposure of normal human fibroblasts to 0.05-1 microM (+/-)-anti-BPDE resulted in elevated p53 protein levels as compared to the constitutive levels of control cells. The rapid induction response, detectable within a few hours, was sustained up to a period of at least 24 h. Repair-proficient and repair-deficient (XPA) human lymphoblastoid cells showed a similar response. The poly(ADP-ribose) polymerase inhibitor, 3-aminobenzamide (3-AB), diminished the p53 induction response by concomitantly decreasing the extent of (+/-)-anti-BPDE induced DNA damage in cells pretreated with the inhibitor. However, the direct involvement of poly ADP-ribosylation was also apparent as 3-AB was able to attenuate (approximately 50%) the p53 response by post-damage inhibitor treatment of the cells. Inhibition of cellular DNA replication by hydroxyurea and AraC, in the presence or absence of DNA damage, also resulted in rapid p53 accumulation in repair-deficient cells. On the contrary, inhibition of protein kinase C (PKC) by calphostin-C led to an abrogation of (+/-)-anti-BPDE mediated p53 induction. Analysis of the downstream effects of carcinogen treatment showed that the lymphoblastoid cells undergo DNA fragmentation indicative of apoptosis while fibroblasts exhibit cell cycle arrest at the G1-S boundary.
p53基因产物的快速积累被认为是细胞对多种基因毒素反应的重要组成部分。为了深入了解导致p53稳定的生化途径，研究了（±）7,8-二羟基-反式-9,10-环氧-7,8,9,10-四氢苯并[a]芘[（±）-反式-BPDE]诱导的DNA损伤对各种修复功能正常和修复功能缺陷的人类细胞中p53蛋白水平的影响。正常人成纤维细胞短暂暴露于0.05 - 1 microM（±）-反式-BPDE后，与对照细胞的组成水平相比，p53蛋白水平升高。在数小时内即可检测到的快速诱导反应可持续至少24小时。修复功能正常和修复功能缺陷（XPA）的人类淋巴母细胞显示出类似的反应。聚（ADP-核糖）聚合酶抑制剂3-氨基苯甲酰胺（3-AB）通过同时降低用该抑制剂预处理的细胞中（±）-反式-BPDE诱导的DNA损伤程度，减弱了p53诱导反应。然而，聚ADP-核糖基化的直接参与也很明显，因为3-AB能够通过损伤后对细胞进行抑制剂处理来减弱（约50%）p53反应。在有或没有DNA损伤的情况下，羟基脲和阿糖胞苷对细胞DNA复制的抑制也导致修复功能缺陷细胞中p53的快速积累。相反，钙泊三醇-C对蛋白激酶C（PKC） 的抑制导致（±）-反式-BPDE介导的p53诱导被消除。致癌物处理的下游效应分析表明，淋巴母细胞经历了指示细胞凋亡的DNA片段化，而成纤维细胞在G1-S边界处表现出细胞周期停滞。
