BACKGROUND & AIMS:
Several recent studies have suggested that testicular germ cell tumors express high levels of wild-type p53 protein. To clarify and confirm this unexpected result, we have investigated seminomatous and nonseminomatous germ cell tumors at the genomic, mRNA, and protein levels. Thirty-five tumors were examined for p53 overexpression using antibodies directed against the p53 (PAb1801, PAb240, and CM1), mdm2 (IF2), and p21Waf1/Clp1 (EA10) proteins. Thirty-two tumors were screened for p53 mutations by single-strand conformation polymorphism analysis. Eighteen tumors were screened with a functional assay that tests the transcriptional competence of human p53 protein expressed in yeast. On frozen sections, 100, 65, 35, 73, and 0% of tumors reacted with the CM1, PAb240, PAb1801, IF2, and EA10 antibodies, respectively. No p53 mutations were detected by single-strand conformation polymorphism or by functional assay. The fact that many tumors overexpress wild-type p53 but not mdm2 rules out mdm2 overexpression as a general explanation for the presence of wild-type p53 in these tumors. The absence of p21 overexpression suggests that p53 may be unable to activate transcription of critical target genes, which may explain why the presence of wild-type p53 is tolerated in this tumor type, although the mechanism for this transcriptional inactivity remains to be established.
背景与目标:
最近的一些研究表明,睾丸生殖细胞肿瘤表达高水平的野生型p53蛋白。为了澄清和确认这一意外结果,我们在基因组,mRNA和蛋白质水平上研究了精原细胞和非精原细胞生殖细胞肿瘤。使用针对p53 (PAb1801,PAb240和CM1),mdm2 (IF2) 和p21Waf1/Clp1 (EA10) 蛋白的抗体检查了35个肿瘤的p53过表达。通过单链构象多态性分析筛选了32个肿瘤的p53突变。用功能测定法筛选了18个肿瘤,该测定法测试了酵母中表达的人p53蛋白的转录能力。在冷冻切片上,100、65、35、73和0% 的肿瘤分别与CM1、PAb240、PAb1801、IF2和EA10抗体反应。通过单链构象多态性或功能分析未检测到p53突变。许多肿瘤过表达野生型p53而非mdm2的事实排除了mdm2过表达作为这些肿瘤中野生型p53存在的一般解释。p21过表达的缺乏表明p53可能无法激活关键靶基因的转录,这可以解释为什么在这种肿瘤类型中可以耐受野生型p53的存在,尽管这种转录不活跃的机制仍有待建立。