Syndecan-3-mediated signalling in the regulation of myogenesis

Long-term maintenance of skeletal muscle is important for tissue health and function. Muscle stem cells, also termed satellite cells, are quiescent muscle progenitors residing within the muscle tissue and are indispensable for postnatal muscle growth and regeneration. Once activated by injury, satellite cells convert to proliferating myoblasts, which eventually differentiate and fuse to generate muscle fibres. Heparan sulphate proteoglycans, such as the syndecans, are expressed in satellite cells and contribute to regulation of several growth factor signalling pathways in addition to regulating cell-cell and cell-matrix adhesion. Previous studies have shown that genetic ablation of syndecan-3, a transmembrane heparan sulphate proteoglycan expressed in satellite cells and myoblasts, dramatically improves muscle regenerative potential in ageing and in pathological conditions such as muscular dystrophy. However, the molecular mechanisms underlying syndecan-3 function in muscle regeneration are poorly understood. In this project, I used an unbiased approach based on proteomics and bioinformatics to identify syndecan-3- regulated signalling pathways in myoblasts. I then validated the proteomics and bioinformatics results using traditional biochemistry and cell biology techniques. Interesting results were obtained from a global phosphoproteomic analysis of a control myoblast cell line compared to a syndecan-3 knockdown myoblast cell line. A general trend was observed in syndecan-3 knockdown myoblasts where the insulin/PI3K/mTOR signalling pathway was over-activated in serum-starved syndecan-3 knockdown myoblasts compared to control myoblasts. However, the same insulin/PI3K/mTOR signalling pathway showed a reduced response to serum stimulation in syndecan-3 knockdown myoblasts compared to control myoblasts. Therefore, I investigated the role of syndecan-3 in the regulation of insulin signalling in myoblasts. Results from insulin stimulation experiments in C2C12 and primary satellite cell-derived myoblasts revealed that syndecan-3 regulates the insulin signalling pathway likely by inhibiting AKT activation and consequently regulates the balance between myoblast proliferation and differentiation. These results indicate a novel role for syndecan3 in the regulation of insulin signalling during myogenesis.