Molecular Genetics of Cytoplasmic Dyein

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DESCRIPTION (provided by applicant): The dyneins are a class of motor protein involved in many of aspects of microtubule-dependent movement. Cytoplasmic dynein, in particular, is responsible for diverse forms of vesicular, macromolecular, and mitotic movement, as well as organization of the microtubule cytoskeleton and directed cell movement. How the dyneins function to produce force, and how cytoplasmic dynein is selectively targeted to a diversity of subcellular structures represent two of the major outstanding issues in the field. This proposal pursues Aims directed at each or these issues based on progress during the preceding project period. Aim I deals with the stalk, a 15 nm long projection extending from the motor domain, which is responsible for microtubule binding. This Aim will determine the structure of the stalk at atomic resolution; it will further identify conformational changes within its putative antiparallel coiled-coil D-helix responsible for long-range allosteric coupling of microtubule binding and ATP hydrolysis. Aim II addresses mechanisms involved in cytoplasmic dynein cargo binding. Our recent studies have revealed a general role for the ZW10 protein complex in linking dynactin and dynein to diverse subcellular structures. We will define the mechanism responsible for ZW10 membrane binding; focusing on the Golgi apparatus, we will determine the role of ZW10 relative to Rab6, spectrin, and other factors implicated in dynein cargo binding, with the goal of a complete elucidation of the Golgi cargo binding pathway. Aim III will involve dynein-mediated virus motility. We will focus on adenovirus as a structurally simple pathogenic form of dynein cargo. We will extend and complete our efforts to identify the interacting partners between dynein and the adenovirus capsid. Together these studies have important implications for understanding many aspects of normal and abnormal cell and developmental biology. The final aim has particular significance for the control of acute viral infection and the design of gene delivery vectors and strategies.