Regulation of cytoplasmic dynein-mediated transport by the mammalian Bicaudal D protein.

Cytoplasmic dynein is a large motor complex, which is responsible for the microtubule-minus end-directed transport of numerous cargos, including cellular organelles, chromosomes and protein complexes. Due to the large size and complexity of the dynein motor, the knowledge about the mechanisms of dynein association with its cargo and the regulation of its motor activity is rather limited. Mammalian Bicaudal D1 and 2 proteins were identified in our previous studies as adaptors, linking the dynein motor to certain types of cytoplasmic vesicles. The aim of the current project is to gain insight into the mechanism of interaction between Bicaudal D2 protein and dynein-dynactin complexes and to address the role of different dynein and dynactin subunits in Bicaudal D2-dependent dynein transport. This will be achieved by biochemical purification of protein complexes, analysis of the protein-protein interactions by chemical cross-linking and mass spectrometry, and with binding studies using purified proteins. The role of dynein and dynactin components will be studied by using RNA interference-mediated knockdown in cultured cells. A novel assay, based on the inducible Bicaudal D- and dynein-dependent transport in cultured cells will be used to investigate the relevance of the identified interactions, as well as the relative importance of different dynein and dynactin subunits.