Tyrosine Phosphorylation in Lens Cell Differentiation

描述(由申请人提供)：Src酪氨酸激酶的激活是对氧化、热和紫外线应激的反应，所有这些应激刺激都会促进晶状体白内障的形成。我们发现，抑制Src家族酪氨酸激酶(SFK)可以阻止晶状体混浊的形成。在这项建议中，我们将确定SFK激活诱导白内障形成的分子机制。我们的长期目标是确定导致晶状体疾病的信号通路的变化。这些网络中的信号中间体可能是药物干预以抑制晶状体混浊形成的候选对象。了解SFK的激活如何导致白内障的形成的关键是描绘SFK在正常晶状体分化和发育中的功能。我们将确定SFK在发育中的胚胎晶状体中调节钙粘附素功能的机制。为了将这些研究扩展到晶状体白内障的病理生理学，我们将使用两个Src诱导的晶状体疾病模型来研究SFKs异常激活的结构和功能靶点，重点是钙粘连蛋白连接失稳。一种是体外模型，将晶状体细胞培养物转化为对温度敏感的v-Src激酶，利用该模型，我们将分析在不同发育阶段，Src激活对晶状体钙粘蛋白连接的稳定性和功能的分子影响。这些研究将在整个晶状体培养模型中平行进行，该模型非常接近于应激诱发的白内障。在这个模型中，我们将能够将由于SFK的不适当激活而导致的分子变化与晶状体混浊的形成联系起来。我们推测，SFK影响晶状体细胞分化的一个机制是通过调节钙粘蛋白复合体，而对SFK信号通路的不适当调节通过破坏钙粘素连接而诱导晶状体白内障。我们建议1)确定在正常晶状体细胞分化中，Src家族激酶调节钙粘蛋白功能的机制；2)利用v-Src转化的晶状体细胞培养物作为应激性晶状体疾病的模型，确定Src激酶结构性激活干扰晶状体钙粘蛋白连接的结构和功能的机制；以及3)确定通过靶向钙粘蛋白连接而不适当地激活Src家族蛋白，从而导致晶状体白内障形成的机制。

DESCRIPTION (provided by applicant): Activation of Src tyrosine kinases occurs in response to oxidative, heat, and UV stress, all stress stimuli that promote the formation of lens cataract. We have found that inhibition of Src family tyrosine kinases (SFKs) blocks the development of lens opacity. In this proposal we will identify the molecular mechanisms by which SFK activation induces cataract formation. Our long term objectives are to identify the alterations in signaling pathways that lead to lens disease. The signaling intermediates in these networks are likely candidates for pharmaceutical intervention to suppress the formation of lens opacities. Key to understanding how the activation of SFKs leads to cataract formation is the delineation of SFK functions in normal lens differentiation and development. We will determine the mechanisms whereby SFKs regulate cadherin function in the developing embryonic lens. To extend these studies to the pathophysiology of lens cataract, we will examine the structural and functional targets of inappropriately activation of SFKs, focusing on cadherin junction destabilization, using two Src-induced lens disease models. One is an in vitro model in which lens cell cultures are transformed with a temperature sensitive v-Src kinase with which we will dissect the molecular affects of Src activation on the stabilization and function of lens cadherin junctions at different stages of development. These studies will be paralleled in a whole lens culture model that closely approximates stress-induced cataract. In this model we will be able to link the molecular changes that result from the inappropriate activation of SFKs with the formation of lens opacities. We hypothesize that one mechanism by which SFKs influence lens cell differentiation is through their regulation of cadherin complexes and that inappropriate regulation of the SFK signaling pathways induces lens cataracts by destabilizing cadherin junctions. We propose to 1) determine the mechanisms whereby Src family kinases regulate cadherin function in normal lens cell differentiation; 2) determine the mechanisms whereby constitutive activation of the Src kinase interferes with the structure and function of lens cadherin junctions, using v-Src transformed lens cell cultures as a model for stress-induced lens disease; and 3) determine the mechanisms whereby the inappropriate activation of Src family kinases, through their targeting of cadherin junctions, induces formation of lens cataracts.