Functional substitutions of amino acids that differ between GDF11 and GDF8 impact skeletal development and skeletal muscle.
Growth differentiation factor 11 (GDF11) and GDF8 (MSTN) are closely related TGF{beta} family proteins that interact with nearly identical signaling receptors and antagonist proteins. However, GDF11 appears to activate SMAD2/3 in vitro and in vivo more potently than GDF8. The ligands possess divergent structural properties, whereby substituting unique GDF11 amino acids into GDF8 enhanced activity of the resulting chimeric GDF8. We investigated potentially distinct endogenous activities of GDF11 and GDF8 in vivo by genetically modifying their mature signaling domains. Full recoding of the GDF8 mature domain to that of GDF11 yielded mice lacking GDF8, with GDF11 levels [~]50-fold higher than normal, and exhibiting modestly decreased muscle mass, with no apparent negative impacts on health or survival to adulthood. Substitution of two specific amino acids in the fingertip region of GDF11 with the corresponding GDF8 residues resulted in prenatal axial skeletal transformations, consistent with Gdf11-deficient mice, without apparent perturbation of skeletal or cardiac muscle development or homeostasis. These experiments uncover distinctive features between the GDF11 and GDF8 mature domains in vivo and identify specific requirement for GDF11 in early-stage skeletal development. Summary StatementReplacement of amino acids unique to GDF11 and GDF8 into the native locus of the other ligand yields measurable, differential skeletal and muscle phenotypes, revealing distinct features between the ligands.