Signaling mechanisms utilized by myostatin and related ligands in skeletal muscle

Myostatin (MSTN, GDF-8) is a secreted signaling protein that we discovered many years ago as a transforming growth factor-ß (TGF-ß) family member that normally acts to limit skeletal muscle growth. A major focus of my laboratory has been to elucidate the molecular, cellular, and physiological mechanisms underlying MSTN activity, with the goal of developing strategies to target this pathway for clinical applications. Biochemical studies carried out by us and by others have identified many key regulatory and signaling components, which has led to the model that MSTN, along with activin A, regulates muscle growth and function by binding first to the type 2 receptors, ACVR2 and ACVR2B, and then engaging the type 1 receptors, ALK4 and ALK5. The formation of this receptor complex leads to activation of SMAD2 and SMAD3, which are the key mediators of the canonical signaling pathway for these ligands. To determine their roles in vivo, we have generated mice carrying flox alleles for each of these components and examined the effect of targeting them either individually or in combination specifically in skeletal myofibers. These studies have led to unexpected findings that have revealed major gaps in our understanding of how this system operates in vivo. One unexpected finding was that simultaneously targeting SMAD2 and SMAD3 led to only modest increases in muscle mass, implying that MSTN/activin A utilize both SMAD-dependent and SMAD-independent pathways in vivo. A second unexpected finding was that targeting the two type 1 receptors for MSTN/activin A has a substantially greater effect on muscle mass than targeting the two type 2 receptors. Our new findings, taken together with studies by other groups showing that muscle growth is regulated by other TGF-ß family members, specifically BMPs, that act antagonistically to MSTN and activin A, have led us to propose a new model that competition between MSTN/activin A and BMPs occurs primarily at the level of the type 2 receptors. The overall goal of this project will be to use a combination of genetic and pharmacological approaches to elucidate further how all of these regulatory components interrelate in muscle, specifically to determine whether different components of this regulatory system have distinct roles in regulating muscle growth and function, particularly with respect to metabolic function. The Specific Aims of this project will be to elucidate SMAD-dependent versus SMAD-independent effects in skeletal myofibers; to assess the effect of blocking SMAD-dependent and SMAD-independent pathways in skeletal muscle in mouse models of metabolic dysfunction and muscle wasting; and to elucidate how MSTN/activin A signaling and BMP signaling interrelate in myofibers. We believe that the studies outlined in this proposal will provide key insights into the mechanisms by which this complex network of regulatory and signaling components function in an integrated manner to regulate muscle homeostasis and will inform drug development efforts to maximize the anabolic potential of targeting this pathway in muscle to treat patients with muscle and metabolic diseases.