P53 AND P21 WAF PROTEINS MODULATE JNK ACTIVITY

P53 和 P21 WAF 蛋白调节 JNK 活性

• 批准号: 2842136
• 负责人: JILL C. PELLING
• 金额: $ 20.59万
• 依托单位: UNIVERSITY OF KANSAS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-04-15 至 2003-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2842136
• 关键词: biological signal transduction; calmodulin dependent protein kinase; cell growth regulation; cell line; enzyme activity; enzyme induction /repression; gene targeting; genetically modified animals; immunoprecipitation; laboratory mouse; oncoprotein p21; p53 gene /protein; phosphorylation; protein protein interaction; radiobiology; skin; ultraviolet radiation

Exposure of mammalian cells to ultraviolet light (UV) and DNA damaging agents triggers the UV response which is characterized by induction of cell cycle regulatory proteins such as p53 and the cyclin-cyclin dependent kinase inhibitor p21WAF. Cells also respond to genotoxic stress by activation of the Jun amino terminal kinase/stress-activated protein kinase pathway (JNK/SAPK). Our preliminary results demonstrate that both wildtype p53 protein and p2lWAF protein can be co- immuneprecipitated with JNK1 enzyme in whole cell extracts. These results represent, to the best of our knowledge, the first report that both p53 and p21WAF proteins are associated with JNK1 enzyme in the cell. In addition, our studies indicate that when cells are expressing wildtype p53 protein and p2lWAF protein, basal JNK1 activity is low and JNK1 can be induced many-fold by UV above control levels. In contrast when cells are expressing mutant p53val135 protein and low levels of p2lWAF, basal JNK1 activity is elevated and fold induction of JNK1 by UV above control is reduced. Further analysis of JNK1 protein has provided preliminary evidence that the phosphorylation status of JNK1 is altered in cells expressing wildtype P53 and p21WAF1 protein (distinct from that normally associated with JNK1 activation). These preliminary results have led us to hypothesize that wildtype p53 protein and p21WAF both bind to JNK1 in the cell and this interaction affects JNK1 signal transduction by inhibiting basal JNK1 activity and modulating the ability of JNK1 to be induced by stress stimuli such as UV irradiation. We further hypothesize that the phosphorylation status of the JNK1 protein is modulated by a p53/p2lWAF-dependent mechanism. The following specific aims will test these hypotheses: Aim number 1 will characterize the interaction of p53 protein and p21WAF protein with JNK protein in cultured cells. Aim number 2 will investigate the effect of p53/p21WAF protein interaction with JNK1 enzyme on JNK1 function in cells. Aim number 3 will investigate the mechanism by which the phosphorylation status of JNK1 protein is altered in cells expressing p53/p21WAF. Aim number 4 will characterize the interaction of p53 protein, p2lWAFprotein, and JNK1 protein in vivo.

哺乳动物细胞暴露于紫外线（UV）和DNA损伤 试剂触发UV响应，其特征在于诱导 细胞周期调节蛋白如p53和细胞周期蛋白-细胞周期蛋白 依赖性激酶抑制剂p21 WAF。 细胞也对遗传毒性 通过激活Jun氨基末端激酶的应激/应激激活的 蛋白激酶途径（JNK/SAPK）。 我们的初步结果表明 野生型p53蛋白和p21 WAF蛋白都可以共表达。 在全细胞提取物中用JNK 1酶免疫沉淀。这些 据我们所知，这些结果是第一份报告， p53和p21 WAF蛋白都与JNK 1酶相关， cell. 此外，我们的研究表明，当细胞表达 野生型p53蛋白和p21 WAF蛋白，基础JNK 1活性低， JNK 1可被UV诱导许多倍于对照水平。 相比之下 当细胞表达突变p53 val 135蛋白和低水平的 p21 WAF，基础JNK 1活性升高，并且通过p21 WAF，JNK 1的诱导倍数增加。 UV高于对照降低。 JNK 1蛋白的进一步分析 提供了JNK 1磷酸化状态的初步证据， 在表达野生型P53和p21 WAF 1蛋白的细胞中改变 （不同于通常与JNK 1活化相关的）。 这些 初步结果使我们假设野生型p53蛋白 和p21 WAF都与细胞中的JNK 1结合，这种相互作用影响 JNK 1信号转导通过抑制基础JNK 1活性， 调节JNK 1被应激刺激诱导的能力， 紫外线照射。 我们进一步假设， JNK 1蛋白的表达受p53/p21 WAF依赖性机制调节。 以下具体目标将检验这些假设： 将表征p53蛋白和p21 WAF蛋白与 培养细胞中的JNK蛋白。目标2将研究 p53/p21 WAF蛋白与JNK 1酶相互作用对JNK 1功能的影响 细胞第三个目标是研究 JNK 1蛋白的磷酸化状态在表达 p53/p21WAF。 目的4将描述p53与p53之间的相互作用。 蛋白、p21 WAF蛋白和JNK 1蛋白的表达。