Myosin Transducer Inter-Strand Communication Is Critical for Normal ATPase Activity and Myofibril Structure.

Abstract

Simple SummaryMyosin is a molecular motor protein that is critical for using stored energy to yield contraction in both skeletal muscle and in the heart. Mutations in myosin can result in inherited human hypertrophic cardiomyopathy, wherein the heart walls become thickened. This results in inability to adequately pump enough oxygenated blood to meet bodily demands, and can lead to abnormal heart beating patterns and the possibility of sudden death. Typically, over-active myosin can be causative for this disease. Here, we used the common fruit fly to study one of these cardiomyopathy-causing mutations by expressing the mutant protein in the indirect flight muscle of the fly. Surprisingly, we found that the mutation and associated mutations decrease myosin and muscle function instead of enhancing it. We show that at least some of these defects result from improper interactions within the myosin protein. This suggests that appropriate communications within myosin are critical for its function and further indicates that certain aspects of indirect flight muscle myosin are functionally distinct from those of myosin in the human heart. By understanding the intricate interactions within the myosin motor protein that are important for its normal role, therapeutics for improving mutant myosin function can be developed.The R249Q mutation in human β-cardiac myosin results in hypertrophic cardiomyopathy. We previously showed that inserting this mutation into Drosophila melanogaster indirect flight muscle myosin yields mechanical and locomotory defects. Here, we use transgenic Drosophila mutants to demonstrate that residue R249 serves as a critical communication link within myosin that controls both ATPase activity and myofibril integrity. R249 is located on a β-strand of the central transducer of myosin, and our molecular modeling shows that it interacts via a salt bridge with D262 on the adjacent β-strand. We find that disrupting this interaction via R249Q, R249D or D262R mutations reduces basal and actin-activated ATPase activity, actin in vitro motility and flight muscle function. Further, the R249D mutation dramatically affects myofibril assembly, yielding abnormalities in sarcomere lengths, increased Z-line thickness and split myofibrils. These defects are exacerbated during aging. Re-establishing the β-strand interaction via a R249D/D262R double mutation restores both basal ATPase activity and myofibril assembly, indicating that these properties are dependent upon transducer inter-strand communication. Thus, the transducer plays an important role in myosin function and myofibril architecture.