INTERCELLULAR COMMUNICATION IN THE EYE LENS

描述（改编自申请人摘要）：最近的一项研究表明， 透镜间隙连接蛋白46（Cx46）缺陷的小鼠 白内障，表明间隙连接介导的通信的直接作用 保持透镜的透明度。 缝隙连接是一组 连接相邻细胞的细胞质的跨膜通道。 这些 通道允许小的代谢物、离子和第二信使从 细胞间的联系 脊椎动物眼睛内部的透镜纤维细胞 透镜既没有血液供应也没有细胞器。 因此，透镜存活率和 体内平衡独特地依赖于经由间隙细胞间通讯 在透镜表面与上皮细胞的连接。 小鸡透镜是 选择作为实验系统，因为它提供了许多优势， 与其他物种。 已知有三种连接蛋白参与了 透镜间隙连接通道的形成：透镜上皮细胞中的Cx43， Cx45.6和Cx56主要存在于透镜纤维中。 透镜纤维蛋白， 在动物的整个生命周期中， 磷酸化 透镜连接蛋白的磷酸化可能具有功能性 在透镜间隙连接通道的调节中的含义。 此外，委员会认为， 来自其他物种的证据表明， 涉及间隙连接通道组装和门控。 第一个目标是 建议是确定连接蛋白磷酸化的功能作用， 该透镜包括：识别磷酸化位点，研究 连接蛋白磷酸化的功能作用，并确定相关的 涉及的激酶。 第二个目标是通过实验来确定 通过不同的透镜连接蛋白正确靶向的信号， 异聚连接蛋白的形成。 最终目标涉及 表征透镜连接蛋白和MIP之间的关系， MIP在透镜间隙连接中的作用。 拟议的实验将是 在已建立的系统中进行，包括完整的鸡胚晶状体， 原代透镜培养物、连接蛋白缺陷细胞系和非洲爪蟾卵母细胞， 此外，本申请人新开发的一种新的逆转录病毒测定系统， 将采用其中外源连接蛋白将被引入到 原位鸡胚透镜，以确定透镜间隙的作用 透镜生理学和病理学的连接点。

DESCRIPTION (Adapted from applicant's abstract): A recent study shows that mice deficient in the lens gap junction protein connexin 46 (Cx46) develop cataracts, indicating the direct role of gap junction-mediated communication in maintaining lens transparency. Gap junctions are clusters of transmembrane channels that connect the cytoplasm of adjacent cells. These channels permit small metabolites, ions and second messengers to pass from cell to cell. Lens fiber cells within the interior of the vertebrate eye lens have nether a blood supply nor organelles. Thus, lens survival and homeostasis are uniquely dependent on intercellular communication via gap junctions with epithelial cells at the lens surface. The chick lens is chosen as an experimental system as it provides many advantages compared with other species. Three connexins are known to participate in the formation of lens gap junction channels: Cx43 in lens epithelial cells, Cx45.6 and Cx56 mainly in lens fibers. Lens fiber proteins, which survive the whole life span of animals, are susceptible to post-translational phosphorylation. Phosphorylation of lens connexins may have functional implications in the regulation of lens gap junction channels. Moreover, evidence from other species has shown that phosphorylation of connexins is related to gap junction channel assembly and gating. The first goal of this proposal is to determine the functional role of connexin phosphorylation in the lens which includes: identifying phosphorylation sites, studying functional roles of connexin phosphorylation, and determining the relevant kinases involved. The second objective involves experiments to determine the signals for the correct targeting by different lens connexins and the formation of heteromeric connexins. The final objective involves the characterization of relationships between lens connexins and MIP and the function of MIP in lens gap junctions. The proposed experiments will be performed in established systems, including intact embryonic chicken lenses, primary lens cultures, connexin-deficient cell lines and Xenopus oocytes, In addition, a novel retroviral assay system, newly developed by the applicant, will be employed in which exogenous connexins will be introduced into the embryonic chick lens in situ in order to define the roles of lens gap junctions in lens physiology and pathology.