Role of Dystroglycan in Myoblast Adhesion

Dystroglycan is a ubiquitously expressed cellular adhesion protein, linking the actin cytoskeleton to the extracellular matrix. Previous studies have shown dystroglycan to be important in maintaining the mechanical integrity of cells, as well as functioning as a scaffold for the convergence of various intracellular signalling pathways. The protein is also crucial in formation of the basal membrane during embryogenesis, as well as in various subsequent developmental processes. Given its structural and mechano-transducive properties, this work reveals further roles for dystroglycan, particularly with respect to cell-substrate adhesion and matrix degradation. In order to study the involvement of dystroglycan in cells of skeletal muscle origin, H-2Kb-tsA58 myoblasts were infected with retroviral constructs, to produce cells stably overexpressing dystroglycan, or an shRNA-dystroglycan oligonucleotide to deplete dystroglycan by RNAi. Overexpression of dystroglycan induced the formation of filopodial protrusions, and cell motility increased and directional persistence decreased. Knockdown of dystroglycan decreased motility but did not affect persistence. A role for dystroglycan in early cell adhesion was also identified, with dystroglycan being crucial for the formation of adhesive puncta, and affecting adhesion and spreading on various substrates. In addition, a biochemical interaction between dystroglycan and vinculin was observed,· and modulating dystroglycan protein levels disrupted vinculin-based adhesions in polarised myoblasts. Dystroglycan was also found to be a novel component of podosomeslinvadapodia, which were observed for the first time in striated muscle cells. Depletion of dystroglycan by RNAi significantly decreased the ability of cells to produce podosomes, whilst overexpression completely abolished podosome formation. Biochemical analysis revealed an SH3-mediated interaction between dystroglycan and the crucial podosome protein Tks5, and supported a tripartite dystroglycanrrks5/Src complex regulating podosome formation. The data presented here augments the importance of dystroglycan in cell adhesion, and reveals a variety of novel roles for this multifunctional membrane protein. This raises the exciting possibility of further roles for dystroglycan in cell biology, and its involvement in diseases such as muscular dystrophy and cancer.EThOS - Electronic Theses Online ServiceGBUnited Kingdo