Study of Cell Adhesion Through an Analysis of Myosin 7

肌球蛋白VII（M7）是一种肌动蛋白为基础的运动蛋白，已知在人类，小鼠和斑马鱼的听力和平衡中发挥重要作用。简单的真核生物Dictyosteoblasts表达M7同源物（DdM 7）和无效突变体分析表明，它在细胞基质粘附所需的吞噬作用和细胞迁移，一个新的功能肌球蛋白中发挥关键作用。来自哺乳动物系统的新证据与M7直接参与粘附一致，表明这类肌球蛋白的作用在整个进化过程中一直是保守的。我们对Ddm 7的分析表明，这种肌球蛋白在将受体组织成高亲和力复合物（粘附复合物）中起作用，该复合物能够结合表面，并且不参与蛋白质向质膜的运输。此外，我们发现Ddm 7的尾部区域对于这种粘附复合物的形成是必不可少的。我们工作的目标是通过回答以下主要问题来确定基于DdM 7的粘附的分子基础：1）将DdM 7连接到粘附复合物的蛋白质是什么？生物化学和遗传学方法都将用于鉴定与DdM 7结合并参与细胞粘附的蛋白质;以及2）接头蛋白如何与DdM 7相互作用以形成粘附复合物。将采用互补分析来鉴定将该肌球蛋白连接至粘附复合物所需的DdM 7分子的区域。鉴于M7功能在整个进化过程中的保守性，对M7在遗传上易处理的系统（如网骨藻）中功能的详细分子分析将提供一个独特的机会，以深入了解M7在听觉和前庭系统的特化细胞中的粘附功能以及M7的突变（如在Usher综合征IB型中发生的突变）如何导致耳聋。

Myosin VII (M7) is an actin-based motor protein known to play an essential role in hearing and balance in humans, mice and zebrafish. The simple eukaryote Dictyostelium expresses a M7 homologue (DdM7) and null mutant analysis has revealed that it plays a critical role in cell substrate adhesion required for phagocytosis and cell migration, a novel function for a myosin. Emerging evidence from mammalian systems is consistent with M7's direct involve involvement in adhesion, indicating that the role of this class of myosins has been conserved throughout evolution. Our analysis of Ddm7 suggests that this myosin plays a role in organizing receptors into a high affinity complex (the adhesion complex) that is capable of binding to surfaces and is not involved in the transport of proteins to the plasma membrane. Furthermore, we have found that the tail region of Ddm7 is essential for the formation of this adhesion complex. The goal of our work is to identify the molecular basis of DdM7-based adhesion by answering the following major questions: 1) What are the proteins that link DdM7 to the adhesion complex? Both biochemical and genetic approaches will be used to identify proteins that bind to DdM7 and participate in cell adhesion; and 2) How do the linker proteins interact with DdM7 to make an adhesion complex. A complementation analysis will be employed to identify regions of the DdM7 molecule required for linking this myosin to the adhesion complex. Given the conservation of M7 function throughout evolution, the detailed molecular analysis of M7 function in a genetically tractable system such as Dictyostelium shall offer a unique opportunity to gain insight into how M7-based adhesion functions in the specialized cells of the auditory and vestibular system and how mutations in M7, such as occur in Usher's syndrome type IB, might lead to deafness.