Structure and function of Myosin VI

描述(由申请人提供)：VI类肌球蛋白可能是非常规肌球蛋白中最非常规的一类。与所有其他特征的肌球蛋白相比，它们以相反的方向(减去肌动蛋白细丝的末端定向)运输。Myosin VI还对其短杠杆臂进行了不同寻常且有争议的延伸，以增加其步长。它重新排列其转化器构象，以实现较大的行程，并通过未知的机制进行二聚化。肌球蛋白VI参与了许多细胞功能，但对维持毛细胞的立毛纤毛和突变是必不可少的 肌球蛋白VI可导致耳聋。虽然它可以作为一个进行性的肌球蛋白马达，在内吞作用中运输货物，但它也可以作为一个依赖于应变的锚，参与组织结构，如高尔基体。肌球蛋白VI在细胞中被发现是一种单体，但作为二聚体的功能最好。我们已经提出，这表明了一种新的调控形式，可能与肌球蛋白VII和X~共享，即货物启动的二聚作用。本研究的目的是：(1)进一步描述肌球蛋白VI的杠杆臂延伸、负责二聚化的区域和肌球蛋白VI的门控机制2开始探索肌球蛋白VIIa和X的结构方面，它们似乎受到货物引发的二聚作用的调节，就像肌球蛋白VI~(3)一样。 使用生化和光学陷阱分析进一步鉴定导致耳聋的肌球蛋白VI突变~和(4)使用细胞分析(Caco-2细胞)和突变体来确定肌球蛋白VI的锚定和运输作用。 公共卫生相关性：肌球蛋白VI的突变会导致耳聋，并可能导致心肌病和肠道上皮细胞缺陷。肌球蛋白VI的许多设计特征与其他肌球蛋白类别不同，并未被完全理解，许多问题存在争议。这项研究将描绘肌球蛋白VI设计背后的结构适应，并详细说明导致人类耳聋的突变的影响。此外，它还试图确定不同的突变是否会导致不同的细胞缺陷。

DESCRIPTION (provided by applicant): Class VI myosins are perhaps the most unconventional of the unconventional myosin classes. They traffic in the reverse direction (minus end-directed on the actin filament) as compared to all other characterized myosins. Myosin VI also has an unusual and controversial extension of its short lever arm to increase its step size. It rearranges its converter conformation to achieve a large strok and dimerizes by an unknown mechanism. Myosin VI is involved in a number of cellular functions, but is essential for maintenance of the stereocilia of the hair cells and thus mutations in myosin VI can result in deafness. While it can function as a processive myosin motor, transporting cargoes in endocytosis, it also can function as a strain-dependent anchor that is involved in organizing structures such as the Golgi apparatus. Myosin VI is found as a monomer in cells, but functions optimally as a dimer. We have proposed that this is indicative of a novel form of regulation that may be shared by myosins VII and X~ namely, cargo-initiated dimerization. The aims of the study are: (1) further delineate the lever arm extension of myosin VI, the regions responsible for dimerization, and the mechanism of myosin VI gating~ (2) begin to probe structural aspects of myosins VIIa and X, which appear to be regulated in cells by cargo-initiated dimerization, as is the case for myosin VI~ (3) further characterize myosin VI mutations that result in deafness using both biochemical and optical trap assays~ and (4) use cellular assays (Caco-2 cells) and mutants to define anchoring vs. transport roles of myosin VI. PUBLIC HEALTH RELEVANCE: Mutations in myosin VI lead to deafness, and potentially to cardiomyopathy and deficits in the intestinal epithelium. Many of the design feature of myosin VI are unlike other myosin classes and are not fully understood, with a number of issues being controversial. This study will delineate the structural adaptation underlying myosin VI design and detail the impact of mutations that lead to deafness in humans. Furthermore, it seeks to determine if different mutations result in differential cellular deficits.