CELL ADHESION INDUCED DIFFERENTIATION IN EYE TISSUE

在眼睛发育过程中，特定的细胞类型与祖细胞有所不同 不同胚胎来源的细胞，以及细胞与细胞的相互作用 形成一种高效、工作的感觉器官，其方式 是不被理解的。之后，眼细胞类型必须保持其 为眼睛的健康和手术提供不同的表型。 几种导致失明的疾病是辨证所致 变化，例如，视网膜的黄斑变性，白内障 晶状体与视网膜色素上皮的转分化 (增殖性玻璃体视网膜疾病)。钙粘附素依赖于钙离子 哺乳动物发育所必需的细胞间黏附分子。 L最近展示了钙粘附素对细胞间黏附分子的控制 视网膜色素上皮细胞的分化状态；异位 E-钙粘附素在大鼠视网膜色素上皮细胞系中的表达 RPE-J，导致包括重组在内的表型转换 组成蛋白和诱导合成特定的mRNAs和 产生基础-侧向Na+，K+-ATPase极性、Ankyrin亚型的蛋白质 开关、桥粒连接复合体组装和角蛋白细丝 集合。利用这些观察，我们现在建议分析机制 用于确定视网膜色素上皮的分化状态 并将这些观察扩展到眼睛的其他细胞类型。我们的特定 拟议研究的目标是：(I)识别钙粘附素序列 视网膜色素上皮分化所需。嵌合体 内源性视网膜色素上皮之间会产生钙粘附素 钙粘附素和E-钙粘附素在RPE-J细胞中的表达 定义诱导表型变化所需的序列。(Ii) 识别可能转导钙粘素的钙粘素相关分子- 视网膜色素上皮的诱导分化信号。之后 确定在钙粘附素诱导过程中使用的序列 不同的计划，生化和酵母两种杂交方法将 用于识别与这些序列相互作用的蛋白质。(Iii) 分析钙粘附素诱导晶状体上皮细胞分化的作用。像视网膜一样 色素上皮细胞、晶状体上皮细胞不内源性 表达E-钙粘附素。我们将测试钙粘附素诱导的通用性 这种眼球细胞类型的分化程序。(四)确定小说 钙粘附素，可建立和维持分化状态 晶状体上皮细胞和视网膜细胞类型。我们已经启动了一项搜索 对于在眼组织中表达的新型钙粘附素。初步 来自我的实验室的证据表明，有钙粘素分子 在视网膜中表达，以前没有在那里描述过。我们 建议使用RT-PCR方法对新的钙粘附素进行广泛的筛选，并 克隆编码这些新钙粘附素的全长cDNA。结果是 这些研究将提供钙粘附素的详细机制框架。 在眼睛形态发生过程中诱导分化的变化，并将识别 差异化计划中失败的潜在站点 眼组织的病理生理过程。

During eye development, specific cell types differentiate from progenitor cells of diverse embryonic origin, and cell-cell interactions orchestrate the formation of a highly efficient, working sensory organ, in ways which are not understood. Later, the ocular cell types must maintain their differentiated phenotype for proper health and operation of the eye. Several diseases leading to loss of sight result from differentiation changes, for example, macular degeneration in the retina, cataracts of the lens and transdifferentiation of the retinal pigment epithelium (proliferative vitreoretinal disorders). Cadherins are calcium dependent cell-cell adhesion molecules that are essential for mammalian development. l have recently shown that cadherin cell-cell adhesion molecules control the differentiation state in retinal pigment epithelial cells; ectopic expression of E-cadherin in a rat retinal pigment epithelium cell line, RPE-J, results in a phenotypic transformation that includes reorganization of constitutive proteins and induced synthesis of specific mRNAS and proteins producing basal-lateral Na+, K+-ATPase polarity, ankyrin isoform switching, desmosome junctional complex assembly and keratin filament assembly. Using these observations, we now propose to analyze mechanisms for determining the differentiated state of the retinal pigment epithelium and extend these observations to other cell types of the eye. Our specific goals in the proposed studies are to: (i) identify cadherin sequences required for retinal pigment epithelium differentiation. Chimeric cadherins will be generated between endogenous retinal pigment epithelium cadherin and E-cadherin and expressed by transfection in RPE-J cells to define the sequences required for inducing phenotypic changes. (ii) Identify cadherin-associated molecules which may transduce cadherin- induced differentiation cues of the retinal pigment epithelium. After defining the sequences which are utilized during the cadherin-induced differentia~on program, biochemical and yeast two hybrid methods will be used to identify proteins which interact with these sequences. (iii) Analyze cadherin-induced differentiation the lens epithelium. Like retinal pigment epithelial cells, lens epithelial cells do not endogenously express E-cadherin. We will test the generality of the cadherin-induced differentiation program in this ocular cell type. (iv) Identify novel cadherins which may establish and maintain the differentiated state of lens epithelial cells and retinal cell types. We have initiated a search for novel cadherins which are expressed in ocular tissues. Preliminary evidence from my laboratory shows that there are cadherin molecules expressed in the retina which have not been previously described there. We propose an extensive screen for novel cadherins using RT-PCR methods, and to clone full length cDNAs encoding these novel cadherins. The results of these studies will provide a detailed mechanistic framework of cadherin- induced differentiation changes during eye morphogenesis and will identify potential sites in the differentiation program which fail during pathophysiological processes in eye tissues.