Novel MYBPC1 mutations cosegregate with a myopathy associated with muscle weakness, hypotonia and tremor

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Abstract Myosin Binding Protein-C (MyBP-C) comprises a family of accessory proteins that directly interact with both thick myosin and thin actin filaments. Three distinct isoforms have been characterized, including the cardiac (c), slow (s) skeletal and fast (f) skeletal. During the last forty years, numerous studies have focused on the biology of cMyBP-C primarily due to its high mutational prevalence in heart disease. On the contrary, the regulation and roles of the skeletal isoforms have been mainly inferred due to the structural similarity they share with cMyBP-C. Our group has been studying the biology of sMyBP-C aiming to understand its regulation, roles, and disease association. Our findings are highly novel and intriguing. First, we found that MYBPC1, the gene that encodes sMyBP-C, is heavily spliced giving rise to multiple variants that can be co-expressed in the same muscle and myofiber. Second, the presence of alternatively spliced insertions affects the ability of the NH2 and COOH termini to bind actin and myosin, and regulate the formation of actomyosin crossbridges in vitro. Third, the NH2-terminus of sMyBP-C undergoes extensive PKA- and PKC-mediated phosphorylation, which is altered in disease. Fourth, sMyBP-C has both structural and regulatory roles with its structural role in the organization and maintenance of thick myosin filaments preceding its regulatory role in modulating cross- bridge cycling. Fifth, four novel, dominant, missense mutations located in the NH2-terminal M-motif of sMyBP-C co-segregate with of a new myopathy characterized by muscle weakness, hypotonia, facial and body deformities, and high-frequency irregular tremor. Molecular modeling and biochemical studies indicated that the four myopathic mutations differentially affect the ability of the NH2-terminus of sMyBP-C to bind myosin, and the structure and stability of the M-motif. We therefore hypothesize that the four MYBPC1 mutations may differentially alter the biochemical and biophysical properties of sMyBP-C compromising its structural and regulatory roles, yet elicit similar myopathic phenotypes. We further propose that mutant sMyBP-C results in the formation of abnormal and deregulated cross-bridges, which in addition to causing a deficit in force production underlying muscle weakness, act as the primary pacemaker of the observed tremor. The goal of our proposal is to comprehensively study the pathogenesis of this novel form of MYBPC1-associated myopathy using a combination of sophisticated in vitro approaches (Aim 1) and novel preclinical models (Aims 2 & 3). The proposed studies are highly significant in terms of discovery science as we will mechanistically examine the etiologies of this myopathy, and impactful in terms of translational/clinical research for accurate disease diagnosis as well as appropriate and effective treatment design.