The human myostatin precursor protein : structure, function and amyloid formation : implications for the muscle wastage disease sporadic inclusion body myositis : a dissertation presented in partial fulfillment of the requirements for the degree of Doctor of Philosophy in Biochemistry at Massey University, Palmerston North, New Zealand

Abstract

Myostatin is a major player in the regulation of mammalian muscle growth and development, maintaining the balance between proliferation and differentiation prenatally and the quiescence of satellite cells in adults. An absence or overexpression of myostatin results in double-muscling and cachexia respectively, placing myostatin as a promising target in the treatment of muscle wastage diseases. As a transforming growth factor-β superfamily member, myostatin is produced as a precursor protein, consisting of a propeptide region N-terminal to the growth factor domain. Cleavage of the precursor between the domains forms the myostatin latent complex, an inhibitory structure which is exported from the cell where a second cleavage event releases the active myostatin growth factor. The precursor protein, propeptide, and latent complex play important roles in the regulation of myostatin. However, their structure and function are poorly understood, and a possible role for the myostatin precursor protein in the muscle wastage disease sporadic inclusion body myositis, suggests that pre-growth factor forms of myostatin may be additional important therapeutic targets. This thesis presents an investigation into the structure and function of the myostatin precursor protein, the latent complex, and the propeptide region within these, with comparisons to a mutant form of myostatin responsible for the naturally-occurring double-muscled phenotype of the Piedmontese cattle breed. Results suggest that the diverse functions of the propeptide region are facilitated by regions of intrinsic disorder within a primarily structured domain, and that conformational alterations accompany the precursor to latent complex transition, resulting in a tighter inhibitory structure. Comparative analyses between the wild-type and mutant proteins suggest that the Piedmontese phenotype is due to a reduced capacity for covalent dimerisation and significant structural alterations within the type I receptor-binding domain. Investigation into misfolded myostatin precursor protein found that the precursor is able to form cytotoxic amyloid aggregates and mature fibrils under partially denaturing conditions, suggesting a possible mechanism for the role of the myostatin precursor in sporadic inclusion body myositis. Together, these novel results contribute important information towards an understanding of myostatin structure, function and regulation in both normal and disease scenarios.