Regulation of Myosin V Interaction with Cargo

描述(申请人提供)：细胞器运动是细胞分裂和细胞分化的标志。在每种细胞类型中，每种细胞器的位置、数量和形态都被改变，以实现特定的细胞功能。细胞器的长距离运动发生在微管上，而细胞器在最终目的地的定位依赖于基于肌动蛋白的马达。在所有真核生物中，肌球蛋白V马达在基于肌动蛋白的运动中起着关键作用，而肌球蛋白V功能的破坏会导致人类疾病。例如，肌球蛋白Va转运的部分缺陷会导致以神经和色素缺陷为特征的格里斯塞利综合征。肌球蛋白VB的部分缺陷会导致微绒毛包涵体病，其特征是婴儿腹泻，危及生命。目前，这两种疾病都没有有效的药物治疗方法。确定基于肌球蛋白V的运输是如何实现的，可能为在正确的时间和地点处理货物中的每一种情况提供重要的新见解。肌球蛋白V的球状尾部结构域(GTD)通过细胞器特异的接头蛋白附着在其货物上。GTD和接头蛋白的调节有助于指定货物附着。我们确定了酿酒酵母肌球蛋白V马达Myo2的GTD的高分辨率结构，并确定了两个不同的货物结合区域；一个与酵母液泡结合，另一个与分泌小泡结合。我们发现空泡结合部位与一种新的蛋白质Vac17相互作用。分泌囊泡结合部位直接与Rab GTP酶结合。重要的是，我们发现Rab GTP酶结合位点在人肌球蛋白Va和肌球蛋白Vb中是保守的。因此，我们在酵母中的研究揭示了人肌球蛋白V马达上的Rab GTP酶结合位点。酵母Myo2和人肌球蛋白Va和Vb的货物结合域的一般保守，以及Rab GTP酶直接作用于CARN的事实。我们的主要目标是：1)确定单个Myo2接头蛋白的结合是增强还是抑制其他接头蛋白的结合。2)鉴定和鉴定调节Rab GTP酶与Myo2 GTD结合的蛋白质。3)确定调节肌球蛋白V从货物中脱离的机制。

DESCRIPTION (provided by applicant): Organelle movement is a hallmark of cell division and cellular differentiation. In every cell-type, the localization, number and morphology of each type of organelle is modified to achieve specific cellular functions. Long-range movement of organelles occurs on microtubules, while positioning of organelles at their final destination relies on actin-based motors. Myosin V motors play a critical role in actin based movement in all eukaryotes, and disruption of myosin V function causes disease in humans. For example, partial defects in myosin Va based transport cause Griscelli's syndrome, characterized by neurological and pigmentation defects. Partial defects in myosin Vb cause microvillus inclusion disease, characterized by infantile, life threatening diarrhea. Currently there are no effective drug-based treatments for either disease. Determination of how myosin V based transport is achieved may provide important new insights into treating theseetach from cargoes at the correct time and place. The globular tail domain (GTD) of myosin V attaches to its cargoes through organelle-specific adaptor proteins. Regulation of the GTD and adaptor proteins contributes to specifying cargo attachment. We determined a high-resolution structure of the GTD of Myo2, a Saccharomyces cerevisiae myosin V motor, and identified two distinct cargo binding regions; one required for binding to the yeast vacuole, the other required for binding to secretory vesicles. We found that the vacuole binding site interacts with a novel protein, Vac17. The secretory vesicle binding site attaches directly to a Rab GTPase. Importantly, we found that the Rab GTPase binding site is conserved in human myosin Va and myosin Vb. Thus, our studies in yeast revealed the Rab GTPase binding site on human myosin V motors. The general conservation of the cargo binding domains of yeast Myo2 and human myosin Va and Vb, and the fact that Rab GTPases act directly to attach carns. Our major goals are to: 1) Determine whether the binding of an individual Myo2 adaptor protein enhances or inhibits binding of other adaptor proteins. 2) Identify and characterize proteins that regulate the attachment of Rab GTPases to the GTD of Myo2. 3) Determine mechanisms that regulate the detachment of myosin V from cargoes.