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PATHWAYS OF INSULIN AND IGFI RECEPTOR SIGNALING

胰岛素和 IGFI 受体信号传导途径

基本信息

批准号：
2331471
负责人：
MORDECAI P BLAUSTEIN
金额：
$ 17.9万
依托单位：
UNIVERSITY OF MARYLAND BALTIMORE
依托单位国家：
美国
项目类别：
财政年份：
1996
资助国家：
美国
起止时间：
1996-02-17 至 1998-01-31
项目状态：
已结题

项目摘要

Much progress has been made in recent years toward an understanding of the molecular mechanisms by which receptors with intrinsic tyrosine kinase activity mediate their cellular effects. These advances have come predominantly from the identification and cloning of cytoplasmic proteins which interact with the activated receptor tyrosine kinase (RTK) and transduce signals from the receptor to the interior of the cell. The insulin receptor (IR) and insulin-like growth factor I receptor (IGFIR) represent a subclass of RTKs which mediate a variety of effects in responsive tissues including regulation of cellular mitosis, metabolism and differentiation. Although a number of well-characterized mitogenic signaling pathways have been shown to emanate from these receptors, the nature of the metabolic and differentiation-related signaling is less clear. It is of great importance that we have a full understanding of the pathways which mediate signaling by these receptors. Such information will allow insight into diseases such as Type II diabetes which is characterized by insulin resistance and certain neoplastic diseases in which IGFI has been implicated. In this proposal we will ask the following fundamental questions regarding insulin and IGFI signaling, (1) What are the intracellular targets and signaling pathways responsible for mediating the diverse effects of the IR and IGFIR in skeletal muscle, an insulin- and IGFI- responsive tissue?, (2) How do signaling proteins acting through the structurally similar IR and IGFIR regulate such distinct cellular effects as mitogenesis, metabolism and differentiation?, and (3) What is the molecular nature of the novel (non-SH2) phosphotyrosine-dependent interaction of SHC and IRS-1 with the IR and IGFIR which we have identified? To begin to address these questions, we have developed a novel approach with which to rapidly identify and clone cDNAs whose protein products interact directly with the cytoplasmic domains of these receptors. We have successfully adapted the "two-hybrid" assay of protein:protein interaction in the yeast Saccharomyces cerevisiae to demonstrate and characterize the interaction of the IR and IGFIR with several proteins known to be involved in insulin and IGFI signaling including IRS-1, p85, and SHC. This assay has allowed the identification of a novel homologous motif within SHC and IRS-1 which mediates the phosphotyrosine-dependent interaction with the NPEY motif of the IR and IGFIR. In addition, we have demonstrated the utility of the two-hybrid assay in the identification and cloning of known and novel interacting proteins from a cDNA library. This new approach should allow a more complete understanding of the fundamental molecular mechanisms by which these important receptors function.

近几年来，在理解受体与内源性酪氨酸作用的分子机制激酶活性调节它们的细胞效应。这些进步已经到来主要来自细胞质蛋白的鉴定和克隆它们与激活的受体酪氨酸激酶(RTK)相互作用，将信号从受体传递到细胞内部。这个胰岛素受体和胰岛素样生长因子I受体表示RTK的一个子类，它们在反应组织，包括对细胞有丝分裂、代谢的调节和差异化。尽管一些特性良好的有丝分裂原信号通路已被证明来自这些受体，即本质上与代谢和分化相关的信号较少安全。重要的是，我们要充分了解通过这些受体传递信号的通路。这样的信息将使人们能够深入了解II型糖尿病等疾病以胰岛素抵抗和某些肿瘤疾病为特征其中IGFI已被牵连。在这项提案中，我们将提出以下基本问题关于胰岛素和IGFI信号，(1)什么是细胞内靶点和信号通路负责调节不同的胰岛素和胰岛素样生长因子对骨骼肌IR和IGFIR的影响响应性组织？(2)信号蛋白如何通过结构相似的IR和IGFIR调节这种不同的细胞效应作为有丝分裂、新陈代谢和分化？以及(3)什么是新型(非SH2)磷酸酪氨酸依赖的分子性质 SHC和IRS-1与已有的IR和IGFIR的相互作用有身份吗？为了开始解决这些问题，我们开发了一种新的方法用于快速鉴定和克隆其蛋白质产物的cDNA 直接与这些受体的细胞质区域相互作用。我们成功地采用了蛋白质的“双杂交”分析方法：蛋白质在酵母中的相互作用展示和酿酒酵母 IR和IGFIR与几种蛋白质相互作用的表征已知参与胰岛素和IGFI信号转导，包括IRS-1，p85，和SHC。这项检测已经能够鉴定出一种新的同源物 SHC和IRS-1中介导磷酸酪氨酸依赖的基序与IR和IGFIR的NPEY基序相互作用。此外，我们还有证明了双杂交技术在鉴定中的实用性。以及从cDNA文库中克隆已知的和新的相互作用蛋白。这种新的方法应该可以更全面地理解这些重要受体的基本分子机制功能。