MYOSIN VI FUNCTION AND MECHANISM

描述（由申请人提供）：肌球蛋白VI具有许多生物化学和生物物理特性，这些特性使其与其他20多种基于肌动蛋白的马达区分开来。一个惊人的独特特征是它能够向肌动蛋白的尖端/负端移动，与所有其他家族成员相反的方向。这表明肌球蛋白VI在体内具有独特的功能。肌球蛋白VI参与了大量不同的细胞过程，包括内吞作用、运动性和细胞内定位，这些过程利用其沿沿着肌动蛋白丝移动和与肌动蛋白紧密结合（锚）的双重能力。这些不同的功能被认为需要不同于其他肌球蛋白的肌球蛋白VI的结构特征和生物物理性质。然而，几乎没有任何信息已经报道，证明是否有趣的和独特的性能在体外测量是重要的在体内。换句话说，我们对肌球蛋白VI的功能了解很多，但对它的实际功能知之甚少。提出的研究将解决这个问题，使用果蝇作为模型系统。到目前为止，肌球蛋白VI被认为是稳定肌动蛋白结构参与细胞重塑在果蝇精子细胞的发展。在这个过程中，肌球蛋白VI与组织特异性调节分子Androcam结合，而不是与其通常的调节因子钙调素结合。这种结合如何影响其功能尚不清楚。使用转基因果蝇线，表达突变体和截短/删除形式的肌球蛋白VI，以挽救肌球蛋白VI突变体的缺陷，揭示了在体外测量的一些独特的性能可能不会发挥关键作用，在其能力，参与肌动蛋白在体内的稳定。拟议的研究扩展了这些观察，检查精子细胞的发育和其他发育过程中，肌球蛋白VI工程，以了解在体内的各种细胞环境中的重要的结构和生化/生物物理特征。将进行GFP标记分子的定位研究和动力学测量，以了解不同的功能如何介导肌球蛋白VI参与各种各样的细胞功能。在体外的生物化学和生物物理特性的测量，以确定我们的操作，并改变调节Acam，影响肌球蛋白VI的特性将补充这些在体内的研究。这些研究将可能揭示肌球蛋白VI在体内的机制，这将是有价值的了解基本的细胞过程和疾病，其中肌球蛋白VI发挥作用。例如，肌球蛋白VI在几种癌症中上调，可能参与对转移重要的运动。此外，一些人类耳聋综合征和相关的肥厚性心肌病是由肌球蛋白VI突变引起的。肌球蛋白VI的研究有望阐明这些疾病的潜在缺陷，并可能揭示新的治疗方案。肌球蛋白VI的研究可能会导致在一些基本的细胞过程，如内吞作用，蛋白质定位和细胞和细胞内运动的分选途径参与的机制有更深入的了解。此外，肌球蛋白VI与几种疾病有关，如前列腺癌和卵巢癌、人类耳聋综合征和肥厚性心肌病。进一步了解肌球蛋白VI如何在体内工作，可能会提高我们对疾病机制的理解，并提供新的治疗途径。

DESCRIPTION (provided by applicant): Myosin VI has a number of biochemical and biophysical properties that distinguish it from the more than 20 other classes of actin-based motors. One strikingly unique feature is its ability to move towards the pointed/minus end of actin, the direction opposite all other family members. This suggests that myosin VI has unique functions in vivo. Myosin VI has been implicated in a large number of different cellular processes, including endocytosis, motility, and intracellular localization, processes that utilize its dual abilities to move along actin filaments and to bind tightly to actin (anchor). These different functions are thought to require structural features and biophysical properties of myosin VI that differ from other myosins. However, virtually no information has been reported that demonstrates whether the intriguing and unique properties measured in vitro are important in vivo. In other words, we know much about what myosin VI can do, but little about what it actually does. The studies proposed will address this question, using the fruitfly Drosophila as a model system. So far, myosin VI is known to stabilize an actin structure involved in cellular remodeling during Drosophila spermatid development. In this process, myosin VI binds to a tissue-specific regulatory molecule, Androcam, rather than its usual regulator, Calmodulin. How this binding affects it function is unknown. Using transgenic Drosophila lines that express mutant and truncated/deleted forms of myosin VI to rescue myosin VI mutant defects has revealed that some unique properties measured in vitro may not play a key role in its ability to participate in actin stabilization in vivo. The proposed studies extend these observations, examining both spermatid development and other developmental processes in which myosin VI works to understand the structural and biochemical/biophysical features that are important in vivo in a wide variety of cellular contexts. Localization studies and measurements of dynamics of GFP tagged molecules will be performed to understand how different features mediate myosin VI participation in a wide variety of cellular functions. Measurements in vitro of biochemical and biophysical properties to determine how our manipulations, and altered regulation by Acam, affect myosin VI's properties will complement these in vivo studies. These studies will likely reveal insights into myosin VI mechanisms in vivo that will be valuable in understanding basic cellular processes and diseases in which myosin VI plays a role. For example, myosin VI is up regulated in several cancers, likely participating in motility important for metastasis. Additionally, several Human deafness syndromes and associated hypertrophic cardiomyopathy are caused by mutations in myosin VI. Studies of myosin VI promise to illuminate the underlying defects in these diseases and potentially reveal new treatment options. Studies of myosin VI are likely to result in a greater understanding of mechanisms involved in a number of basic cellular processes, such as endocytosis, sorting pathways for protein localization and cellular and intracellular motility. In addition, myosin VI has been implicated in several diseases, such as prostate and ovarian cancers, human deafness syndromes, and hypertrophic cardiomyopathy. Further understanding of how myosin VI works in vivo is likely to enhance our understanding of disease mechanisms and provide for new routes of treatment.