Tyrosine Phosphorylation in Lens Cell Differentiation

描述（由申请人提供）：钙粘蛋白细胞-细胞粘附连接建立上皮细胞表型，阻止其上皮细胞向间质细胞转变，定义细胞结构和信号形态发生。我们试图了解这些细胞-细胞连接作为防止白内障诱导的监护人的功能。我们的研究显示了钙粘蛋白连接、细胞骨架连接和晶状体分化状态之间独特的结构/功能关系。动态的、高度可溶的N-cadherin/2-catenin粘附连接决定了晶状体上皮细胞的细胞结构，而分化的晶状体纤维细胞高度有序和细长的细胞结构的建立涉及到钙粘蛋白连接的稳定。我们发现晶状体纤维细胞中存在一种新的n -钙粘蛋白连接复合物，通过3-连环蛋白与中间丝细胞骨架相连。我们将研究这种新型钙粘蛋白复合物在为长晶体纤维细胞提供其分化和稳定所需的独特结构特性方面的作用。此外，我们将研究n-钙粘蛋白连接复合物在细胞骨架组织中的分化特异性作用，并研究细胞骨架在稳定钙粘蛋白分化特异性功能中的作用。有趣的是，Src家族酪氨酸激酶（SFKs），钙粘蛋白连接的调节因子，在晶状体分化中具有双重作用。SFK Src维持晶状体上皮细胞处于增殖、未分化状态，SFK Fyn信号晶状体纤维细胞分化。在本文中，我们继续研究SFK调控N-cadherin连接稳定性的机制，以及N-cadherin连接的SFK信号在涉及PI3K和Rac的晶体细胞分化途径中的作用。我们之前的研究表明，Src激酶的不受管制的组成性激活会破坏晶状体钙粘蛋白连接。我们现在在应激性白内障的培养模型中发现，晶状体混浊是由p38应激激酶/Src激酶信号通路激活引起的，该通路靶向钙粘蛋白连接。如果Src激酶激活被阻断，钙粘蛋白连接和正常的晶状体细胞结构都得到维持，晶状体混浊的发生被阻止。我们将研究白内障中n -钙粘蛋白连接不稳定的机制，重点关注蛋白酪氨酸磷酸酶（PTPs）作为Src激酶和钙粘蛋白连接的分子调节剂的作用。ptp被活性氧灭活，这是应力性白内障的常见原因。希望从长远来看，对这一途径的了解将为预防白内障提供合适的药物靶点。我们将从三个方面研究我们的假设：(1)检验n -钙粘蛋白调节细胞骨架以建立和维持晶状体结构和稳定性的假设；(2)验证Src激酶作为钙粘蛋白连接的上游调控因子和下游信号效应因子调控晶状体细胞分化状态的假设；(3)检验Src激酶的错误调控是否会取消钙粘蛋白连接和细胞骨架作为晶状体结构稳定的保护者和白内障守护者的作用。与公共卫生相关：白内障是晶状体混浊，影响其在视网膜上聚焦图像的能力。虽然白内障的许多特性已经被确定，但白内障诱发的机制和预防白内障的潜在方法仍然是难以捉摸的。这项建议的长期目标是了解钙粘蛋白细胞-细胞连接作为对抗这种晶状体疾病的守护者的作用。

DESCRIPTION (provided by applicant): Cadherin cell-cell adhesion junctions establish the epithelial cell phenotype, prevent their epithelial to mesenchymal transition, define cell cytoarchitecture and signal morphogenesis. We seek to understand the function of these cell-cell junctions as guardians against the induction of cataract. Our studies show a unique structure/function relationship between cadherin junctions, their cytoskeletal linkages and lens differentiation state. Dynamic, highly detergent-soluble N-cadherin/2-catenin adherens junctions define the cytoarchitecture of lens epithelial cells while establishment of the highly ordered and elongated cytoarchitecture of differentiating lens fiber cells involves stabilization of cadherin junctions. We discovered the presence of a novel N-cadherin junctional complex in lens fiber cells, linked to the intermediate filament cytoskeleton through 3-catenin. We will investigate this novel cadherin complex for its role in providing the elongating lens fiber cells with the unique structural properties required for their differentiation and stability. In addition, we will study the differentiation-specific role of N-cadherin junctional complexes in the organization of the cytoskeleton and examine the role of the cytoskeleton in stabilizing cadherin differentiation-specific function. Interestingly, Src family tyrosine kinases (SFKs), regulators of cadherin junctions, have a dual role in lens differentiation. The SFK Src maintains lens epithelial cells in a proliferative, undifferentiated state and the SFK Fyn signals lens fiber cell differentiation. In this proposal we continue our investigation of the mechanisms of SFK regulation of N-cadherin junctional stability, and N-cadherin-linked SFK signaling of lens cell differentiation, in a pathway involving PI3K and Rac. Our previous studies show that the unregulated, constitutive activation of Src kinases disassembles lens cadherin junctions. We now find in a culture model for stress-induced cataract that lens opacities result from activation of a p38 stress kinase/Src kinase signaling pathway that targets the cadherin junctions. If Src kinase activation is blocked, both cadherin junctions and normal lens cytoarchitecture are maintained, and the development of lens opacities is prevented. We will investigate the mechanism of N-cadherin junction destabilization in cataract, focusing on the role protein tyrosine phosphatases (PTPs) as molecular regulators of both Src kinases and cadherin junctions. PTPs are inactivated by reactive oxygen species, a common cause of stress-induced cataract. It is hoped that in the long-term, understanding of this pathway will provide suitable pharmaceutical targets for the prevention of cataract. Will investigate our hypotheses in three specific aims: (1) Examine the hypothesis that N-cadherin regulates the cytoskeleton to establish and maintain lens structure and stability; (2) Examine the hypothesis that the Src kinases regulate lens cell differentiation state by their action as both upstream regulators and downstream signaling effectors of cadherin junctions; (3) Examine the hypothesis that misregulation of Src kinases abrogates the role of cadherin junctions and the cytoskeleton as protectors of lens structural stability and guardians against cataract. PUBLIC HEALTH RELEVANCE: Cataracts are opacities of the lens that interfere with its ability to focus an image on the retina. While many of the properties of cataract have been determined, the mechanism of cataract induction and potential methods of cataract prevention still remain elusive. The long term goals of this proposal are to understand the role of cadherin cell-cell junctions as guardians against this lens disease.