PTPLB CONTROL OF CADHERIN FUNCTION & RETINA DEVELOPMENT

PTPLB 对钙粘蛋白功能的控制

基本信息

批准号：
6518587
负责人：
JACK E LILIEN
金额：
$ 19.75万
依托单位：
UNIVERSITY OF IOWA
依托单位国家：
美国
项目类别：
财政年份：
1999
资助国家：
美国
起止时间：
1999-02-01 至 2003-05-31
项目状态：
已结题

项目摘要

Our long-term objectives are to characterize the mechanisms and developmental ramifications of regulated changes in adhesion molecule function the developing retina. This proposal focuses on the regulation of N-cadherin function in the developing retina by the non-receptor protein tyrosine phosphatase PTPlB. Cadherin function depends on its linkage to the actin containing cytoskeleton, a linkage mediated by alpha- and beta-catenin. Phosphorylation of tyrosine residues on beta- catenin is associated with loss of the cadherin-cytoskeletal connection, and consequent loss of adhesive competence. PTPlB is bound to the cytoplasmic domain of N-cadherin and maintain the actin connection by dephosphorylating beta-catenin. Our specific aims are directed at defining the molecular basis for the interaction between N-cadherin and PTPlB, and the role that this interaction plays in retinal morphogenesis. Our specific aims are directed at defining the molecular basis for the interaction between N-cadherin and PTPlB, and the role that this interaction plays in retinal morphogenesis. We will 1. identify and functionally analyze the N-cadherin which interacts with PTP1B through analysis of binding in vitro and in situ. To localize the site, we will construct nested deletions of N-cadherin. We will also develop cell permeable peptides that mimic the N-cadherin binding domain thus interfere with the binding of PTPlB to cadherin in situ. 2. Identify the PTP1B domain essential for targeting to N-cadherin by analyzing binding of PTPlB to cadherin in vitro and in situ following site specific mutagenesis of tyrosine residues and construction of nested deletions. 3. Analyze and compare the role of cadherin-PTPlB interactions with the role of other N-cadherin effector interactions on neurite outgrowth and cell adhesion. We will determine the effect of cell permeable peptides which mimic different regions of the cytoplasmic of N-cadherin on neurite outgrowth on several substrates, as well as their effect on phosphorylation of beta-catenin and the integrity of the cadherin cytoskeletal connection. 4. Analyze the effect of temporally specific loss of cadherin/PTPlB interactions on morphogenesis of intact retina. Cell permeable peptides which perturb the interaction of N- cadherin with PTPlB will be compared to those which disrupt the binding of beta-catenin for their effect on morphogenesis of intact neural retina following addition at specific times during retina development.

我们的长期目标是表征机制和粘附分子变化的发育后果功能发育的视网膜。该提案重点介绍了法规非受体的N-钙粘蛋白功能在发育中的视网膜中的功能蛋白酪氨酸磷酸酶PTPLB。钙粘蛋白功能取决于其与含有细胞骨架的肌动蛋白的链接，由 α-和β-catenin。酪氨酸残基在β- Catenin与钙粘蛋白 - 骨骼连接的丧失有关，以及随之而来的粘合能力的丧失。 PTPLB与 N-钙粘蛋白的细胞质结构域，并通过去磷酸化β-catenin。我们的具体目标是针对的定义N-钙粘着蛋白与 PTPLB以及这种相互作用在视网膜中的作用形态发生。我们的特定目标旨在定义分子 N-钙黏着蛋白和PTPLB之间相互作用的基础，以及角色这种相互作用在视网膜形态发生中起作用。我们将1。识别并在功能上分析与与之相互作用的N-钙粘蛋白 PTP1B通过分析体外和原位的结合。本地化站点，我们将构建N-钙黏着蛋白的嵌套缺失。我们也会开发模仿N-钙粘蛋白结合结构域的细胞渗透肽因此，干扰了PTPLB与钙粘蛋白原位的结合。 2。确定靶向N-钙粘蛋白所必需的PTP1B域在体外和原位分析PTPLB与钙粘蛋白与钙粘蛋白的结合之后酪氨酸残基的特定地点诱变和建造嵌套的删除。 3。分析和比较钙粘蛋白-ptplb的作用与其他N-钙黏着蛋白效应子在上的作用的相互作用神经突生长和细胞粘附。我们将确定细胞渗透的肽，这些肽模仿细胞质的不同区域在几种底物上的神经突生长上的N-钙粘蛋白以及它们对β-catenin的磷酸化的影响及其完整性钙粘蛋白的细胞骨架连接。 4。分析时间的效果钙粘蛋白/PTPLB相互作用的特定损失在完整的形态发生上视网膜。细胞渗透的肽，这些肽会扰动N-的相互作用将带有PTPLB的钙粘蛋白与破坏结合的钙粘蛋白进行比较 β-catenin对完整神经形态发生的影响视网膜在视网膜开发期间的特定时间增加了视网膜。