Ohnishi T, Ohnishi K, Wang X, Takahashi A, Okaichi K
Department of Biology, Nara Medical University, Kashihara, Japan.
Radiat Res. 1999 Apr;151(4):498-500.
We have previously reported that heat stress induces expression of wild-type TP53 (formerly known as p53) activated factor 1 (CDKN1A, formerly known as WAF1) only when TP53 protein is wild-type using cells of a human glioblastoma cell line (A-172) and cells of its transformant (A-172/mp53/ 143) with a mutant TP53 (point mutation at codon 143 from Val to Ala) vector. Transfection of A-172 cells with the mutant TP53 vector abolished the heat-induced expression of CDKN1A, demonstrating the dominant negative nature of this TP53 mutant over the endogenous wild-type TP53. This kind of dominant negative TP53 mutant occurs frequently in various types of cancer. Overcoming this dominance or restoring the normal functions to these TP53 mutants is a new strategy for TP53-targeted cancer therapies. We examined whether glycerol can act as a chemical chaperone to correct the mutant TP53 conformation. No CDKN1A expression was induced after heating or treatment with glycerol at concentrations of 0.6 and 1.2 M in these transformants. In contrast, A-172/mp53/ 143 cells showed CDKN1A expression when they were heated in the presence of glycerol at 0.6 or 1.2 M, which was similar to the response of the parental and neo vector-transfected control cells. To test the generality of the effects of glycerol on mutant TP53, we used human osteosarcoma Saos-2 cells (lacking TP53) transfected with mutant TP53 and cells of two other human glioblastoma cell lines carrying mutant TP53. These cells showed similar CDKN1A expression when heated in the presence of glycerol at 0.6 or 1.2 M. These results suggest that glycerol is effective in restoring several TP53 mutants to normal TP53 function, leading to normal CDKN1A expression after heat stress. This observation provides a novel tool for correction of mutant TP53 conformation and may be applicable for TP53-targeted cancer therapy.
我们之前报道过，热应激仅在TP53蛋白为野生型时，才能诱导野生型TP53（以前称为p53）激活因子1（CDKN1A，以前称为WAF1）的表达，我们使用了人胶质母细胞瘤细胞系（A-172）的细胞及其携带突变型TP53（第143位密码子由缬氨酸突变为丙氨酸的点突变）载体的转化细胞（A-172/mp53/143）。用突变型TP53载体转染A-172细胞消除了热诱导的CDKN1A表达，证明了这种TP53突变体相对于内源性野生型TP53的显性负性作用。这种显性负性TP53突变体在各种类型的癌症中频繁出现。克服这种显性作用或恢复这些TP53突变体的正常功能是针对TP53的癌症治疗的新策略。我们研究了甘油是否可以作为化学伴侣来纠正突变型TP53的构象。在这些转化细胞中，加热或用0.6和1.2 M浓度的甘油处理后，未诱导出CDKN1A表达。相比之下，A-172/mp53/143细胞在0.6或1.2 M甘油存在下加热时显示出CDKN1A表达，这与亲本细胞和新载体转染的对照细胞的反应相似。为了测试甘油对突变型TP53作用的普遍性，我们使用了转染了突变型TP53的人骨肉瘤Saos-2细胞（缺乏TP53）以及另外两种携带突变型TP53的人胶质母细胞瘤细胞系的细胞。当在0.6或1.2 M甘油存在下加热时，这些细胞显示出相似的CDKN1A表达。这些结果表明，甘油可有效将几种TP53突变体恢复为正常TP53功能，导致热应激后CDKN1A正常表达。这一观察结果为纠正突变型TP53构象提供了一种新工具，可能适用于针对TP53的癌症治疗。
