P53 AND P21 WAF PROTEINS MODULATE JNK ACTIVITY

P53 和 P21 WAF 蛋白调节 JNK 活性

基本信息

批准号：
6350384
负责人：
JILL C. PELLING
金额：
$ 24.71万
依托单位：
UNIVERSITY OF KANSAS MEDICAL CENTER
依托单位国家：
美国
项目类别：
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/6350384
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

项目摘要

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 损伤制剂触发紫外线反应，其特点是诱导细胞周期调节蛋白，例如 p53 和细胞周期蛋白-细胞周期蛋白依赖性激酶抑制剂 p21WAF。细胞也对基因毒性有反应通过 Jun 氨基末端激酶激活产生应激/应激激活蛋白激酶途径（JNK/SAPK）。我们的初步结果表明野生型 p53 蛋白和 p2lWAF 蛋白都可以共在全细胞提取物中用 JNK1 酶进行免疫沉淀。这些据我们所知，结果代表了第一份报告 p53 和 p21WAF 蛋白均与 JNK1 酶相关细胞。此外，我们的研究表明，当细胞表达野生型p53蛋白和p2lWAF蛋白，基础JNK1活性低且 JNK1 可以被高于对照水平的紫外线诱导很多倍。相比之下当细胞表达突变型 p53val135 蛋白且低水平的 p2lWAF，基础 JNK1 活性升高，并且 JNK1 的诱导倍数上面控制的紫外线减少了。 JNK1蛋白的进一步分析表明提供了初步证据表明JNK1的磷酸化状态在表达野生型 P53 和 p21WAF1 蛋白的细胞中发生改变（与通常与 JNK1 激活相关的不同）。这些初步结果使我们推测野生型 p53 蛋白和 p21WAF 都与细胞中的 JNK1 结合，这种相互作用影响通过抑制 JNK1 基础活性和 JNK1 信号转导调节 JNK1 被应激刺激诱导的能力，例如紫外线照射。我们进一步假设磷酸化状态 JNK1 蛋白的功能受到 p53/p2lWAF 依赖性机制的调节。以下具体目标将检验这些假设：目标 1 将表征 p53 蛋白和 p21WAF 蛋白的相互作用培养细胞中的 JNK 蛋白。目标 2 将调查效果 p53/p21WAF 蛋白与 JNK1 酶的相互作用对 JNK1 功能的影响细胞。目标 3 将研究机制 JNK1 蛋白的磷酸化状态在表达的细胞中发生改变 p53/p21WAF。目标 4 将表征 p53 的相互作用体内蛋白、p2lWAF蛋白和JNK1蛋白。