MECHANISM OF RAF1 ACTIVATION

描述：Raf-1是MEK 1/2上游的生理激酶，其 下游靶点MAPK/Erk首先被鉴定为胰岛素应答性的 激酶。Raf-1是原癌基因c-Ras的主要下游效应子 并且其活化对于增殖的调节是至关重要的， 分化和其他细胞功能。尽管如此， Raf激活的详细机制尚不完全。研究从 一些实验室已经提出，细胞外信号，如 如胰岛素、IGF和EGF最初与其氨基末端相互作用， 监管领域。其他人和这些研究人员的发现表明， 这种结合打开了无活性Raf-的紧密结构， 可访问到尚未识别的激活器，将Raf锁定在一个活跃的 通过磷酸化和可能的其他机制来调节。我们最近 显示14-3-3是这个激活过程所需的，或者通过 介导Raf-1的二聚化或使其与其它分子相互作用， 帮助可活化Raf装配的蛋白质因子 activation.拟议中的研究将使用生物化学方法来检验这一模型。 和分子方法。具体目标如下：1.到 阐明低聚/二聚作用的机制 Raf-1的激活。2.探讨14-3-3在 Raf激酶活性的调节。3.为了确定Raf上的序列， 对它的激活至关重要重点将是确定一个新的 当EGF激活时，Raf-1的肽图上出现的磷酸肽 了4.为了鉴定磷酸化所述序列的激酶， 在目标3中。这些研究应该提供新的见解的机制， 其中Raf-1响应于其细胞外的变化而被激活， 细胞内环境他们还应该扩大我们对 作用14-3-3和细胞信号转导中的二聚化。

DESCRIPTION: Raf-1 is a physiological kinase upstream of MEK 1/2, whose downstream target, MAPK/Erk was first identified as an insulin-responsive kinase. Raf-1 is a major downstream effector of the proto-oncogene c-Ras and its activation is crucial for the regulation of proliferation, differentiation and other cellular functions. Despite this, knowledge of the detailed mechanism of Raf activation is still incomplete. Studies from a number of laboratories have suggested that extracellular signals, such as insulin, IGF and EGF initially interacting with its amino-terminal regulatory domain. Findings by others and these investigators suggest that this binding opens the tight structure of inactive Raf- and makes it accessible to as yet unidentified activators, which lock Raf in an active state by phosphorylation and possibly other mechanisms. We have recently shown that 14-3-3 is required for this activation process, either by mediating dimerization of Raf-1 or causing it to interact with other protein factors that assist in the assembly of an activatable Raf activation. The proposed studies will examine this model using biochemical and molecular approaches. The specific aims are as follows: 1. To elucidate the mechanism by which oligomerization/dimerization contributes to the activation of Raf-1. 2. To explore the role of 14-3-3 in the regulation of Raf kinase activity. 3. To identify sequences on Raf that are critical for its activation. The focus will be on identifying a new phospho-peptide that appear on the peptide map of Raf-1 when EGF activates it. 4. To identify the kinase that phosphorylates the sequences described in aim 3. These studies should provide new insights into the mechanism by which Raf-1 is activated in responses to changes in its extracellular and intracellular milieu. They should also expend our understanding of the role 14-3-3 and of dimerization in cellular signal transduction.