Tyrosine Phosphorylation in Lens Cell Differentiation

描述（由申请人提供）：Src酪氨酸激酶的激活响应于氧化、热和UV应激而发生，所有应激刺激均促进透镜白内障的形成。我们已经发现Src家族酪氨酸激酶（SFKs）的抑制阻断了透镜混浊的发展。在这个建议中，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透镜疾病的信号通路的改变。这些网络中的信号中间体可能是药物干预抑制透镜混浊形成的候选者。理解SFK激活如何导致白内障形成的关键是阐明SFK在正常透镜分化和发育中的功能。我们将确定SFKs调节胚胎透镜发育中钙粘蛋白功能的机制。为了将这些研究扩展到透镜白内障的病理生理学，我们将使用两种Src诱导的透镜疾病模型，研究SFKs不适当激活的结构和功能靶标，重点是钙粘蛋白连接不稳定。一种是体外模型，其中透镜细胞培养物用温度敏感的v-Src激酶转化，我们将在不同发育阶段剖析Src激活对透镜钙粘蛋白连接的稳定性和功能的分子影响。这些研究将在一个非常接近应激性白内障的全透镜培养模型中进行。在这个模型中，我们将能够将SFKs的不适当激活导致的分子变化与透镜混浊的形成联系起来。我们推测SFK影响透镜细胞分化的一种机制是通过调节钙粘蛋白复合物，并且SFK信号通路的不适当调节通过使钙粘蛋白连接不稳定而诱导透镜白内障。我们建议1）确定Src家族激酶在正常透镜细胞分化中调节钙粘蛋白功能的机制; 2）使用v-Src转化的透镜细胞培养物作为应激诱导的透镜疾病的模型，确定Src激酶的组成性激活干扰透镜钙粘蛋白连接结构和功能的机制;和3）确定Src家族激酶的不适当活化通过其钙粘蛋白连接的靶向诱导透镜白内障形成的机制。

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.