Single cysteine to tyrosine transition inactivates the growth inhibitory function of Piedmontese myostatin

Myostatin, a member of the transforming growth factor-beta superfamily, is a secreted growth factor that is proteolytically processed to give COOH-terminal mature myostatin and NH2-terminal latency-associated peptide in myoblasts. Piedmontese cattle are a heavy-muscled breed that express a mutated form of myostatin in which cysteine (313) is substituted with tyrosine. Here we have characterized the biology of this mutated Piedmontese myostatin. Northern and Western analyses indicate that there is increased expression of myostatin mRNA and precursor myostatin protein in the skeletal muscle of Piedmontese cattle. In contrast, a decrease in mature myostatin was observed in Piedmontese skeletal muscle. However, there is no detectable change in the circulatory levels of mature myostatin in Piedmontese cattle. Myoblast proliferation assay performed with normal and Piedmontese myostatin indicated that mature wild-type myostatin protein inhibited the proliferation of C2C12 myoblasts. Piedmontese myostatin, by contrast, failed to inhibit myoblast proliferation. In addition, when added in molar excess, Piedmontese myostatin acted as a potent "competitive inhibitor" molecule. These results indicate that, in Piedmontese myostatin, substitution of cysteine with tyrosine results in the distortion of the "cystine knot" structure and a loss of biological activity of the myostatin. This mutation also appears to affect either processing or stability of mature myostatin without altering the secretion of myostatin.