Metavinculin Regulation of Cell Cytoskeleton Remodeling in Response to Substrate

 描述（由申请人提供）：心血管疾病是美国的主要死亡原因，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灶性粘附介导，细胞外基质硬度增加导致主动脉结构变化，从而促进高血压的发生。目前尚不清楚粘着斑如何调节血管平滑肌细胞对刚度增加的反应。粘着斑蛋白黏着斑蛋白及其肌肉特异性剪接变体后黏着斑蛋白是力传递的关键组分。因此，我们的长期研究目标是阐明血管平滑肌细胞通过后黏着斑蛋白和黏着斑蛋白对细胞外基质硬度变化的反应机制。这项建议的目的是表征后黏着斑蛋白尾部结构特征的重要肌动蛋白细胞骨架重塑后，细胞外基质硬度的变化。我们的假设是，在肌动蛋白或磷脂结合后释放的后黏着斑蛋白C-末端发夹，与黏着斑蛋白尾部C-末端碱基结合形成异二聚体，这是血管平滑肌细胞中捆绑肌动蛋白纤维所不可或缺的。这一假设将使用新的实验和计算方法的组合来解决，具体目标如下：（1）表征后黏着斑蛋白和黏着斑蛋白分布对底物硬度变化的响应，（2）确定后黏着斑蛋白尾部结构修饰对黏着斑蛋白尾部与肌动蛋白和磷脂的结合的影响，以及（3）确定后黏着斑蛋白尾溶液结构并定位参与肌动蛋白诱导的后黏着斑蛋白-黏着斑蛋白异二聚体形成的后黏着斑蛋白残基。在完成上述工作后，我们希望能够明确后黏着斑蛋白-黏着斑蛋白异二聚体在细胞骨架重塑中的作用，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度增加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，从而有助于降低发病率，死亡率和心血管疾病的差异。

 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.