KSR, a Modifier of Ras Mediated Cell Transformation

描述（由申请人提供）：体内肿瘤形成和体外肿瘤形成 从原代组织培养的细胞的转化需要多个 致癌事件。这些事件是由于创造或引进 显性作用癌基因和肿瘤抑制因子的失活。 细胞对这些基因的转化特性的敏感性可能是 被其他细胞基因修饰，虽然不驱动致癌过程， 是转化事件转导所必需的。中央 该建议的假设是，基因激酶的蛋白质产物 Ras抑制因子（KSR）是一种有效的细胞转化调节剂， 致癌Ras这项提案的目的是解释分子机制 潜在的生物学行为的这种假定的修改器。KSR被鉴定为 作为一个功能丧失的等位基因，抑制活化Ras的表型， 果蝇和C.优雅这些数据表明，正常的KSR可能是一个 在低等真核生物中Ras调节途径的正调节因子。分析 KSR原代成纤维细胞中致癌Ras介导的转化， 小鼠表明KSR在哺乳动物中具有类似的功能。额外分析 证明KSR是Ras/Raf/MEK/ERK激酶的特异性效应子 级联。KSR是磷酸化的，这些突变的一个子集 磷酸化位点导致KSR从细胞质重新分布， 原子核这些观察导致了这样的假设， KSR的磷酸化决定了它的亚细胞定位， KSR的定位影响其调节Ras/Raf/MEK/ERK激酶的能力 级联和Ras介导的细胞转化。这些假设将得到检验 通过：1）鉴定KSR中调节其亚细胞的序列， 2）确定KSR磷酸化在调节细胞内的作用， KSR在细胞质和细胞核之间的分布; 3）表征 KSR与Raf、MEK和ERK物理相互作用及其生物学意义 结果;以及4）确定KSR对细胞的贡献 转化受其亚细胞分布和磷酸化的影响。

DESCRIPTION (provided by applicant): Tumor formation in vivo and the in vitro transformation of cells cultured from primary tissues require multiples oncogenic events. These events result from the creation or introduction of dominant acting oncogenes and the inactivation of tumor suppressors. Susceptibility of a cell to the transforming properties of these genes may be modified by other cellular genes that, while not driving the oncogenic process, are necessary for the transduction of the transforming event. The central hypothesis of this proposal is that the protein product of the gene Kinase Suppressor of Ras (KSR) is a potent modifier of cell transformation by oncogenic Ras. The goal of this proposal is to explain the molecular mechanisms underlying the biological actions of this putative modifier. KSR was identified as a loss-of-function allele that suppresses the phenotype of activated Ras in Drosophila and C. elegans. These data indicated that normal KSR may be a positive regulator of Ras-regulated pathways in lower eukaryotes. Analysis of oncogenic Ras-mediated transformation in primary fibroblasts from KSR about mice indicate that KSR functions similarly in mammals. Additional analyses demonstrate that KSR is a specific effector of the Ras/Raf/MEK/ERK kinase cascade. KSR is phosphorylated, and mutation of a subset of these phosphorylation sites causes the redistribution of KSR from the cytoplasm to the nucleus. These observations have lead to the hypotheses that the phosphorylation of KSR determines its subcellular location and that the localization of KSR affects its ability to regulate the Ras/Raf/MEK/ERK kinase cascade and Ras-mediated cell transformation. These hypotheses will be tested by: 1) identifying the sequences in KSR that regulate its subcellular distribution; 2) determining the role of KSR phosphorylation in regulating the distribution of KSR between the cytoplasm and the nucleus; 3) characterizing the physical interaction of KSR with Raf, MEK and ERK and its biological consequences; and 4) determining how the contribution of KSR to cell transformation is affected by its subcellular distribution and phosphorylation.