Rnd Effector Molecules in Epithelial Cell Transformation

描述(由申请人提供)： 小的GTP酶Ras将激活的细胞表面受体的信号传递给细胞内的信号通路。在起源于极化上皮细胞的腺癌中，RAS信号经常由于激活突变或基因扩增而被解除调控。致癌RAS诱导极化上皮细胞转化的机制尚不清楚。RAS信号的一个主要效应通路是Raf-MEK-ERK通路，它模拟了RAS对细胞的众多作用。极化上皮细胞中活化的RAS或Raf的表达导致肌动蛋白细胞骨架的深刻变化以及与之相关的细胞结构的改变。肌动蛋白细胞骨架受Rho家族GTP结合蛋白的控制。在极化的上皮细胞中，RAS或Raf的激活导致Rho样蛋白RND3的诱导表达，RND3似乎是Rho蛋白的内源性拮抗剂。因此，RND蛋白对细胞的影响被激活形式的Rho蛋白的表达所抵消。对RND信号的效应分子几乎一无所知。识别RND蛋白的效应物很重要，因为它们在细胞中的表达引起了一种表型，表明它们在细胞转化和发育过程中发挥关键作用。在新的研究中，我们发现Rho GTP酶激活蛋白p190可能是RND信号的效应分子。目的1是分析RND和p190蛋白之间的相互作用，并产生这种相互作用的抑制分子。目的二是利用p190被敲除的细胞，验证RND和p190分子之间的相互作用影响p190分子功能域的假设，以及p190是RND蛋白的效应分子的假说。目的3探讨RND-p190相互作用是否通过Ras激活的Raf-MEK-ERK途径，通过Aim 1中产生的抑制分子，对极化上皮细胞的致癌转化起关键作用。

DESCRIPTION (provided by applicant): The small GTPase Ras relays signals from activated cell surface receptors to intracellular signaling pathways. In adenocarcinomas, which originate from polarized epithelial cells, Ras signaling is often deregulated as a result of activating mutations or gene amplification. The mechanisms by which oncogenic Ras elicits transformation of polarized epithelial cells are poorly understood. A major effector pathway of Ras signaling is the Raf-MEK-ERK pathway, which mimics numerous effects of Ras on cells. Expression of activated Ras or Raf in polarized epithelial cells leads to profound alterations in the actin cytoskeleton and associated changes in cellular architecture. The actin eytoskeleton is subject to control by Rho family GTP-binding proteins. Ras or Raf activation in polarized epithelial cells leads to induced expression of the Rho-like protein Rnd3, which appears to function as an endogenous antagonist to Rho proteins. Accordingly, effects of Rnd proteins on cells are counteracted by expression of activated forms of Rho proteins. Virtually nothing is known about effector molecules of Rnd signaling. It is important to identify effectors of Rnd proteins as their expression in cells elicits a phenotype that suggests that they play key roles in cell transformation and developmental processes. In new studies we have identified the Rho GTPase activating protein p190 as a putative effector molecule of Rnd signaling. Aim 1 is to dissect the interaction between Rnd and p190 proteins and to generate an inhibitory molecule of this interaction. Aim 2 is to test the hypothesis that the interaction between Rnd and p190 molecules affects functional domains of the p190 molecule, and that p190 is an effector molecule of Rnd proteins in vivo using cells in which p190 has been knocked out. Aim 3 addresses whether the Rnd-p190 interaction is critical to oncogenic transformation of polarized epithelial by the Ras activated Raf-MEK-ERK pathway using the inhibitory molecule generated in Aim 1. Together, the proposed studies will elucidate mechanisms whereby Ras transforms epithelial cells leading to invasion and metastasis.