Molecular Elucidation of Integrin Signaling

描述(由申请人提供)：我们建议的广泛的长期目标是获得对细胞黏附的详细的分子理解，并阐明它如何调节各种生理和病理过程。在过去的一段时间里，我们一直致力于研究整合素的机制，整合素是一类异二聚体(α/β)跨膜受体，在细胞黏附，特别是细胞与细胞外基质的黏附中发挥关键作用。通过进行强有力的结构研究，我们和其他人在了解整合素是如何激活以触发细胞-ECM黏附的分子方面取得了重大进展。在这个继续的提案中，我们希望开始研究整合素激活后的事件，即由外向内的信号转导，其中整合素胞外区与细胞外基质结合，并引发信号到细胞质表面，触发与细胞骨架相连的细胞内信号通路的级联。广泛的遗传学和生物化学研究表明，这种由外而内的信号转导过程对于调节细胞形态改变(扩散)、细胞迁移、增殖和存活等动态黏附事件是必不可少的，但其潜在的机制尚不清楚。作为我们研究的第一步，我们将通过解决以下直接问题来关注一些关键但具有代表性的途径：(I)在整合素激活及其随后的ECM参与时，小的整合素α/β细胞质尾巴(CT，主要是20-50个残基)究竟如何识别不同的效应器，导致不同的细胞内事件？(Ii)这些不同的整合素-效应器相互作用是如何被调节来调节动态细胞黏附事件的？在初步研究中，我们发现，虽然整合素效应器与各种整合素CTs特异地相互作用，但这些相互作用与其相应的下游CTs惊人地相互排斥。这导致我们假设了一种反馈机制，通过下游靶点来调节整合素-效应器相互作用的开/关开关。这种反馈调节可能促进细胞周期扩散和迁移过程中整合素-细胞骨架的高效组装/重组。因此，我们的目标有两个：(A)定义这些相互作用的分子特异性；(B)大力评估假设的反馈机制。我们将继续使用结构和职能相结合的办法来实现我们的目标。我们相信，这些结果不仅将为不同的整合素-效应器相互作用如何调节不同的自外向内信号通路提供关键的分子见解，而且还将揭开一个统一的邮政编码，即反馈控制，以指导高度协调和动态的细胞扩散和迁移过程。由于参与这些通路的组件的功能障碍与许多人类疾病相关，因此这里得出的具体结果也可能具有治疗价值。

DESCRIPTION (provided by applicant): The broad long-range objective of our proposal is to obtain a detailed molecular understanding of cell adhesion and to elucidate how it mediates various physiological and pathological processes. Over the past funding period, we have been focusing on studying the mechanism of integrins, a class of heterodimeric (alpha/beta) transmembrane receptors that play key roles in the cell adhesion, particularly the cell-ECM adhesion. By performing vigorous structural studies, we and others have made a major progress toward the molecular understanding of how integrins are activated to trigger the cell-ECM adhesion. In this continuation proposal, we wish to begin the investigation on the events following the integrin activation, i.e., outside-in signaling where integrin extracellular domain adheres to ECM and elicits signals to the cytoplasmic face triggering cascades of intracellular signaling pathways that link to cytoskeleton. Extensive genetic and biochemical studies have shown that such outside-in signaling process is essential for regulating dynamic adhesion events such as cell shape change (spreading), cell migration, proliferation, and survival, yet the underlying mechanism remains poorly understood. As a first step of our investigation, we will focus on some critical yet representative pathways by addressing the following immediate questions: (i) Upon integrin activation and its subsequent ECM engagement, how exactly do the small integrin alpha/beta cytoplasmic tails (CTs, mostly 20-50 residues) recognize different effectors leading to different intracellular events? (ii) how are these different integrin-effector interactions modulated to mediate the dynamic cell adhesion events? In preliminary studies, we have found that while integrin effectors specifically interact with various integrin CTs, these interactions are surprisingly mutually exclusive with their corresponding downstream ones. This has led us to hypothesize a feedback mechanism for regulating the on/off switch of the integrin-effector interactions by their downstream targets. Such feedback regulation may promote efficient integrin-cytoskeleton assembly/reassembly during the cyclic cell spreading and migration processes. Hence our goals are 2 fold: (a) to define the molecular specificity of these interactions; and (b) to vigorously evaluate the hypothesized feedback mechanism. We will continue to use the combined structural and functional approach to pursue our goals. We believe that the results will not only provide key molecular insights into how different integrin-effector interactions mediate distinct outside-in signaling pathways but also unravel a unifying zip code, i.e., the feedback control, for directing the highly coordinated and dynamic cell spreading and migration processes. Since dysfunctions of components involved in these pathways are correlated with numerous human disorders, the specific results derived here may be also of therapeutic value.