The Regulation of the MAP Kinase Pathway and DNA Repair

描述（由申请人提供）：本提案的第一个目标是 研究MAP激酶信号通路对DNA修复的调控。使用 功能蛋白质组学，我们的实验室已经确定了hHR23B，一个蛋白质参与 在DNA损伤识别中，作为MAPK的新靶点。蛋白质印迹分析 用针对hHR23B的特异性抗体证明MAPK指导 hHR23B的蛋白水解加工。第一个具体目标的目标是 了解这个新靶点是如何被MAPK调控的。第一、 MAPK调控的翻译后修饰和加工位点 将确定，并将评估这些修改的重要性 在体外和体内修复测定中。初步观察表明， MAPK也被激活，以响应紫外线照射，因此MAPK可能发挥作用， 通过修饰hHR 23 B调节DNA修复的作用。这一目标的完成 将描述一个新的MAPK靶点的调控，并提供新的见解 通过MAPK途径调节DNA修复。在第二个具体目标中， 通过MAPK途径的信号传导的线性将使用 功能蛋白质组学尽管MAPK通路被经典地认为是一种 线性途径，（即Raf激活MKK，MKK激活ERK），存在 越来越多的证据表明，在这一途径中存在分叉，导致 不同靶蛋白的调节。我将首先确定分支点 靶向Raf和MKK水平。第二，我会识别 ERK下游激酶选择性调节的靶点。的 这一目标的完成将导致识别新的信号传导 靶向并解决MAPK途径调节特异性 细胞反应。

DESCRIPTION (provided by applicant): The first goal of this proposal is to study the regulation of DNA repair by the MAP kinase signaling pathway. Using functional proteomics, our laboratory has identified hHR23B, a protein involved in DNA damage recognition, as a novel target of MAPK. Western blot analysis with a specific antibody against hHR23B demonstrated that MAPK directs proteolytic processing of hHR23B. The goal of the first specific aim is to understand how this novel target is regulated by MAPK. First, post-translational modification(s) and site(s) of processing regulated by MAPK will be identified, and the importance of these modifications will be evaluated in in vitro and in vivo repair assays. Preliminary observations indicate that MAPK is also activated in response to UV exposure, therefore MAPK may play a role in modulating DNA repair by modifying hHR23B. The completion of this aim will characterize the regulation of a novel MAPK target and provide new insight into the regulation of DNA repair by MAPK pathways. In the second specific aim, the linearity of signaling through the MAPK pathway will be examined using functional proteomics. Although the MAPK pathway is classically thought of as a linear pathway, (i.e. Raf activates MKK which activates ERK), there is increasing evidence that bifurcations within this pathway exist that result in the regulation of distinct target proteins. I will first identify branch-point targets at the level of Raf and MKK using proteomics. Second, I will identify targets that are selectively regulated by kinases downstream of ERK. The completion of this aim will result in the identification of novel signaling targets and address the mechanism by which the MAPK pathway regulates specific cellular responses.