Protein Tyrosine Phosphatase Alpha in Integrin Signaling and Cell Movement

细胞运动是一个基本的生理过程。细胞运动的主要决定因素是整合素，即细胞膜上的整合素受体，它协调粘连的细胞-底物接触和肌动蛋白细胞骨架的持续重组。整合素参与启动细胞内信号，驱动被称为焦点粘连(FA)的多蛋白复合体的组装。FA信号既控制细胞骨架的重塑，也控制细胞自身的动态组装和分解，从而调节细胞的形状和运动。*受体蛋白酪氨酸磷酸酶α(PTPA)是整合素信号的正调控因子，促进细胞运动。PTPA缺陷的小鼠胚胎成纤维细胞表现出整合素刺激的缺陷迁移，其特征是细胞骨架重排延迟和FA形成受损。通过对这些缺陷的研究，我们发现PTPA在整合素介导的信号转导中扮演两个角色，促进FA组装，最终促进细胞迁移：(I)作为Src-FAK激酶复合体的上游激活因子的催化作用，以及(Ii)作为Src-FAK复合体的下游靶标的非催化作用。在第二个作用中，PTPA被其C末端Tyr789的酪氨酸残基上的活性Src-FAK磷酸化。这对FA的形成和促进细胞迁移的信号转导是关键的，然而PTPA-phosphTyr789(PTPA-pTyr789)的信号作用在很大程度上还不清楚。*我们已经开始识别整合素刺激的调节PTPA磷酸化的分子机制和PTPA-和PTPA-pTyr789信号的新靶点。我们的证据表明，PTPA是FAs形成和功能所必需的分子相互作用的协调者。我们假设PTPA，特别是PTPA-phosphy Tyr789，是FA组装和重塑的关键调节因子，并作为协调分子相互作用的枢纽，这些分子相互作用指导整合素信号转导和决定细胞运动。我们将通过确定含有PTPA和PTPA-pTyr789的分子复合体的性质和调控形成，并阐明它们在FA信号转导和周转(组装和拆卸)中的功能作用来研究这一假说。为此，我们将使用整合的细胞和分子生物学方法来研究表达或缺失野生型和突变型PTPA的独特模型细胞系统。*目标1：研究PTPA和PTPA-pTyr789在调节FA动态中的作用：组装、周转和分子稳定性。*目标2：使用蛋白质组学定义PTPA的分子相互作用，调节其FA定位和信号。*我们的研究将阐明PTPA和PTPA-pTyr789在整合素调节的焦点黏附动力学和信号中的调节和作用。这将进一步加深我们对精确协调的分子网络的理解，该网络控制着细胞运动的基本过程。

Cell movement is a fundamental physiological process. Principal determinants of cell motility are the integrins, receptors on the cell membrane that co-ordinate adhesive cell-substrate contacts with continual re-organization of the actin cytoskeleton. Integrin engagement initiates intracellular signaling that drives the assembly of multiprotein complexes known as focal adhesions (FAs). FA signaling controls both cytoskeletal remodeling and also their own dynamic assembly and disassembly to regulate cell shape and movement.****Receptor protein tyrosine phosphatase alpha (PTPa) is a positive regulator of integrin signaling that promotes cell movement. PTPa-deficient mouse embryo fibroblasts exhibit defective integrin-stimulated migration that is characterized by delayed cytoskeletal rearrangement and impaired FA formation. Through investigation of these defects, we have discovered that PTPa plays two roles in integrin-mediated signaling to promote FA assembly and, ultimately, cell migration: (i) a catalytic role as an upstream activator of the Src-FAK kinase complex, and (ii) a non-catalytic role as a downstream target of the Src-FAK complex. In its second role, PTPa is phosphorylated by active Src-FAK on a tyrosine residue in its C-terminal tail, Tyr789. This is pivotal for FA formation and signaling that promotes cell migration, however the signaling actions of PTPa-phosphoTyr789 (PTPa-pTyr789) are largely unknown.****We have begun to identify integrin-stimulated molecular mechanisms that regulate PTPa phosphorylation and novel targets of PTPa- and PTPa-pTyr789 signaling. Our evidence indicates that PTPa is a coordinator of molecular interactions integral to the formation and function of FAs. We hypothesize that PTPa, specifically PTPa-phosphoTyr789, is a critical regulator of FA assembly and remodeling and acts as a hub to co-ordinate molecular interactions that direct integrin signaling and determine cell movement. We will investigate this hypothesis by determining the nature and regulated formation of PTPa- and PTPa-pTyr789-containing molecular complexes and elucidating their functional roles in FA signaling and turnover (assembly and disassembly). For this, we will use integrated cell and molecular biology approaches to investigate unique model cell systems that express or lack wild-type and mutant forms of PTPa.****Aim 1: Investigate the role of PTPa and PTPa-pTyr789 in regulating FA dynamics: assembly, turnover, and molecular stability.****Aim 2: Define the molecular interactions of PTPa, using proteomics, that regulate its FA localization and signaling.****Overall, our proposed research will elucidate the regulation and roles of PTPa and PTPa-pTyr789, in integrin-regulated focal adhesion dynamics and signaling. This will further our understanding of the precisely orchestrated molecular network that controls the essential process of cell movement.******