Unconventional Myosins of the Hair Cell

描述(由申请人提供)：非常规肌球蛋白是一类重要的分子马达蛋白。它们在真核生物中参与了大量不同的功能，包括囊泡和mRNAs的运输，肌动蛋白的重组，以及主要的结构功能。几种肌球蛋白(IC、VI、VILA和XV)在毛细胞功能中起着重要作用。为了确定这些马达在毛细胞中的具体细胞作用，将使用化学遗传学方法(已经验证了肌球蛋白-1c在缓慢适应中的作用)。突变的肌球蛋白将被设计成扩大它们的ATP结合口袋，这将使它们对合成的ADP类似物敏感。表达这些致敏马达的敲入或转基因小鼠将允许研究每个马达在缓慢适应、肌动蛋白束发育或顶端内吞作用中的假设作用(S)。这种方法提供了几个优点；最重要的是对特定的、选择性的肌球蛋白功能的抑制进行了严格的时间控制。选择性地将特定的肌球蛋白‘锁定’到肌动蛋白细丝上(刚性复合体)的能力使得能够更严格地确定体内功能，而不会出现过度表达造成的伪影。

DESCRIPTION (provided by applicant): Unconventional myosins comprise an important class of molecular motor proteins. They are involved in a myriad of different functions in eukaryotes, including transport of vesicles and mRNAs, actin reorganization, and predominantly structural functions. Several myosins (Ic, VI, VIla, and XV) have essential roles in hair-cell function. To ascertain the specific cellular roles of these respective motors in hair cells, a chemical-genetic approach (already validated for myosin-1c in slow adaptation) will be utilized. Mutant myosins will be engineered to enlarge their ATP binding pockets, which will sensitize them to synthetic ADP analogs. Knock-in or transgenic mice expressing these sensitized motors will allow the study of each motor's hypothesized role(s) in slow adaptation, actin bundle development, or apical endocytosis. This approach provides several advantages; foremost of which is tight temporal control over the specific, selective inhibition of myosin function. The ability to selectively 'lock down' a specific myosin onto actin filaments (rigor complex) enables a more rigorous determination of in vivo function without the artifacts caused by overexpression.