Regulation of cadherin-based cell adhesions by talin proteolysis and arginylation

描述（由申请人提供）：基于钙粘蛋白的粘附连接（AJ）是一种关键的细胞-细胞粘附装置，其失调导致发育失败和癌症。已知AJs的形成依赖于局部肌动蛋白网络的组装，但其调控机制尚不清楚。我们的初步数据表明，塔林，一种蛋白质，其作用与整联蛋白为基础的细胞-基质粘附的肌动蛋白网络的调节，也参与了AJ通过一种新的塔林蛋白水解片段包含黏着斑蛋白和肌动蛋白结合域和二聚化区域（称为VAD片段）。该片段在体内也经历翻译后酰基化。我们假设蛋白水解改变了塔林的结构域结构，使产生的VAD片段定位于钙粘蛋白为基础的AJ。在那里，该片段增强了局部肌动蛋白与其功能结构域的组装并稳定了AJ。此外，乙酰基化改变了VAD片段对其结合伴侣的亲和力，并增强了其在AJs中的生理功能。在这项研究中，我们将测试这一假设在这三个具体的目标：（1）talin VAD片段是如何招募到钙粘蛋白介导的AJ，它是如何增强肌动蛋白装配？(2)钙粘蛋白为基础的AJs的形成依赖于VAD片段吗？(3)乙酰基化如何增强VAD片段的功能？通过这项研究，我们将阐明一种新的调节机制，为钙粘蛋白为基础的细胞间粘附介导的新的处理塔林，这是传统上被认为只与整合素为基础的粘附。此外，该项目将提供新的见解如何保守的，但知之甚少的翻译后酰基化修饰影响细胞中的蛋白质功能。

DESCRIPTION (provided by applicant): The cadherin-based adherens junction (AJ) is a critical cell-cell adhesion apparatus, misregulation of which leads to development failures and cancers. The formation of AJs is known to be dependent on the assembly of a local actin network, but the regulation of this remains unclear. Our preliminary data suggest that talin, a protein known for its role in the regulation of the actin network associated with integrin-based cell-matrix adhesion, is also involved in the AJ through a novel talin proteolytic fragment containing the vinculin- and actin- binding domains and a dimerizing region (referred as the VAD fragment). This fragment also undergoes posttranslational arginylation in vivo. We hypothesize that the proteolysis alters the domain architecture of talin, allowing the resulting VAD fragment to localize to the cadherin- based AJ. There the fragment enhances local actin assembly with its functional domains and stabilizes the AJ. Furthermore, arginylation changes the affinities of the VAD fragment to its binding partners and enhances its physiological function in AJs. In this proposed study we will test this hypothesis in these three specific aims: (1) How is the talin VAD fragment recruited to the cadherin-mediated AJ, and how does it enhance actin assembly? (2) Is the formation of cadherin-based AJs dependent on the VAD fragment? (3) How does arginylation enhance the function of the VAD fragment? Through this study, we will elucidate a novel regulatory mechanism for cadherin-based cell-cell adhesions mediated by the novel processing of talin, which was traditionally considered only associated with integrin-based adhesions. Also this project will provide new insight into how the conserved but poorly understood posttranslational arginylation modification affects protein function in cells.