Metavinculin Regulation of Cell Cytoskeleton Remodeling in Response to Substrate

 描述(申请人提供)：心血管疾病是美国主要的死亡原因，高血压是心血管疾病的重要危险因素。通过细胞-细胞外基质接触或局部黏附，细胞外基质硬度增加导致主动脉结构改变，从而促进高血压的发生。目前尚不清楚局部粘连如何调节血管平滑肌细胞对僵硬增加的反应。粘着斑蛋白vinculin及其肌肉特异性剪接变异体metavinculin是力传递的关键成分。因此，我们的长期研究目标是阐明血管平滑肌细胞通过Metavinculin和vinculin反应细胞外基质硬度变化的机制。这项建议的目的是表征Metavinculin尾部的结构特征，这些结构特征对于细胞外基质硬度变化时肌动蛋白细胞骨架的重塑非常重要。我们的假设是，在肌动蛋白或磷脂结合后释放的Metavinculin C-末端发夹，与vinculin尾部C-末端碱基结合形成异源二聚体，这是在血管平滑肌细胞中捆绑肌动蛋白纤维所必需的。这一假说将使用新的实验和计算方法相结合来解决，具体目的如下：(1)表征Metavinculin和vinculin分布响应底物硬度的变化，(2)确定Metavinculin尾部结构改变对vinculin尾部与肌动蛋白和磷脂结合的影响，以及(3)确定Metavinculin尾部溶液结构并定位参与肌动蛋白诱导的Metavinculin异二聚体形成的Metavinculin残基。在拟议的工作完成后，我们希望确定Metavinculin-vinculin异二聚体在细胞骨架重构中的作用，并建立异二聚体的结构模型。这项研究的结果将有助于揭示由于血管僵硬增加而导致的血管重塑机制，并提出新的干预措施来预防高血压的发展，进而有助于降低心血管疾病的发病率、死亡率和差异。

 DESCRIPTION (provided by applicant): Cardiovascular diseases are the leading cause of death in the United States, and hypertension is an important risk factor for cardiovascular diseases. Mediated through cell-extracellular matrix contact, or focal adhesion, increased extracellular matrix stiffness causes aorta structural changes, thus contributes to hypertension development. It remains unclear how focal adhesion regulates responses of vascular smooth muscle cells to increased stiffness. The focal adhesion protein vinculin and its muscle specific splice variant metavinculin are the key components for force transmission. Therefore, our long-term research goal is to delineate the mechanism by which vascular smooth muscle cells respond to changes in extracellular matrix stiffness through metavinculin and vinculin. The goal of this proposal is to characterize metavinculin tail structural features important for actin cytoskeleton remodeling upon changes in extracellular matrix stiffness. Our hypothesis is that metavinculin C-terminal hairpin, released upon actin or phospholipid binding, binds toward vinculin tail C-terminal base to form a heterodimer, which is indispensable for bundling actin fibers in vascular smooth muscle cells. This hypothesis will be addressed using a combination of novel experimental and computational approaches with the following Specific Aims: (1) to characterize metavinculin and vinculin distribution in response to changes in substrate stiffness, (2) to determine the effect of metavinculin tail structure modification on the association of vinculin tail with actin and phospholipid, and (3) to determine metavinculin tail solution structur and locate metavinculin residues that are involved in actin induced metavinculin-vinculin heterodimer formation. Upon completion of the proposed work, we expect to define the role of metavinculin-vinculin heterodimer in cell cytoskeleton remodeling and build structural models for the heterodimer. The results of this study will help reveal vascular remodeling mechanism due to increased vascular stiffness and suggest new interventions to prevent hypertension development, and in turn help reduce morbidity, mortality, and disparity in cardiovascular diseases.