Towards a better understanding of the mechanisms underlying myosin-related congenital myopathies

Myosin is a family of proteins that plays a crucial role in generating force and motion by interacting with actin filaments in skeletal muscle. Myosin molecules notably contain heavy chain (MyHC) isoforms that have different functional capabilities. Mutations in one of its isoforms, MyHC I/β (encoded by the MYH7 gene) have been reported in humans, associated with muscle weakness and have led to two main distinct skeletal muscle diseases, Laing Distal Myopathy (LDM) and Myosin Storage Myopathy (MSM). The pathophysiological mechanisms by which subtle amino acid changes in the LMM region of MyHC I/β molecules leading to such variable skeletal muscle phenotypes in LDM and MSM patients remain poorly understood. Using a wide range of human MYH7 patient muscle biopsy samples, investigation of primary biophysical defects in the presence of defective MyHC I/β molecules including myosin filament length has revealed no change but rather a shift in myosin head positioning into a disordered relaxed state (DRX). On the road to generating a zebrafish LDM and MSM disease model, several genes were identified to be orthologous to human MYH7. Amongst orthologous genes, smyhc1 was targeted for genome editing using CRISPR/Cas9 to generate a loss of function model. Loss of smyhc1 led to early developmental defects, however, continued to grow to adulthood with no observable muscle defects. Smyhc1 null zebrafish are replaced and compensated by smyhc2 and smyhc3 in adult zebrafish. Work ongoing to generate large deletion of smyhc locus to understand the role of slow MyHC in sarcomere assembly during early developmental stages through to adulthood. It is concluded that in the presence of LDM mutations in the MyBP-C binding domain, myosin heads in the SRX state are destabilised, and zebrafish smyhc1 is orthologous to human MYH7 but only functions during the early stages of development. Continued work to generate knockout of smyhc locus may describe the function of smyhc2 and smyhc3 in later stages of development in the quest to model the progressive phenotype in LDM and MSM patients

Human Skeletal myopathy myosin mutations disrupt myosin head sequestration

Myosin heavy chains encoded by MYH7 and MYH2 are abundant in human skeletal muscle, and important for muscle contraction. However, it is unclear how mutations in these genes disrupt myosin structure and function leading to skeletal muscle myopathies termed myosinopathies. Here, we used multiple approaches to analyse the effects of common MYH7 and MYH2 mutations in the light meromyosin region of myosin (LMM). Analyses of expressed and purified MYH7 and MYH2 LMM mutant proteins combined with in-silico modelling showed that myosin coiled-coil structure and packing of filaments in vitro are commonly disrupted. Using muscle biopsies from patients, and Mant-ATP chase protocols to estimate the proportion of myosin heads that were super-relaxed, together with X-ray diffraction measurements to estimate myosin head order we found that basal myosin ATP consumption was increased and the myosin super-relaxed state was decreased in vivo. In addition, myofibre mechanics experiments to investigate contractile function showed myofibre contractility was not affected. These findings indicate that the structural remodelling associated with LMM mutations induces a pathogenic state in which formation of shutdown heads is impaired, thus increasing myosin head ATP demand in the filaments, rather than affecting contractility. These key findings will help design future therapies for myosinopathies

Human skeletal myopathy myosin mutations disrupt myosin head sequestration

International audienceMyosin heavy chains encoded by MYH7 and MYH2 are abundant in human skeletal muscle and important for muscle contraction. However, it is unclear how mutations in these genes disrupt myosin structure and function leading to skeletal muscle myopathies termed myosinopathies. Here, we used multiple approaches to analyze the effects of common MYH7 and MYH2 mutations in the light meromyosin (LMM) region of myosin. Analyses of expressed and purified MYH7 and MYH2 LMM mutant proteins combined with in silico modeling showed that myosin coiled coil structure and packing of filaments in vitro are commonly disrupted. Using muscle biopsies from patients and fluorescent ATP analog chase protocols to estimate the proportion of myosin heads that were super-relaxed, together with x-ray diffraction measurements to estimate myosin head order, we found that basal myosin ATP consumption was increased and the myosin super-relaxed state was decreased in vivo. In addition, myofiber mechanics experiments to investigate contractile function showed that myofiber contractility was not affected. These findings indicate that the structural remodeling associated with LMM mutations induces a pathogenic state in which formation of shutdown heads is impaired, thus increasing myosin head ATP demand in the filaments, rather than affecting contractility. These key findings will help design future therapies for myosinopathies

Human skeletal myopathy myosin mutations disrupt myosin head sequestration

Myosin heavy chains encoded by MYH7 and MYH2 are abundant in human skeletal muscle and important for muscle contraction. However, it is unclear how mutations in these genes disrupt myosin structure and function leading to skeletal muscle myopathies termed myosinopathies. Here, we used multiple approaches to analyze the effects of common MYH7 and MYH2 mutations in the light meromyosin (LMM) region of myosin. Analyses of expressed and purified MYH7 and MYH2 LMM mutant proteins combined with in silico modeling showed that myosin coiled coil structure and packing of filaments in vitro are commonly disrupted. Using muscle biopsies from patients and fluorescent ATP analog chase protocols to estimate the proportion of myosin heads that were super-relaxed, together with x-ray diffraction measurements to estimate myosin head order, we found that basal myosin ATP consumption was increased and the myosin super-relaxed state was decreased in vivo. In addition, myofiber mechanics experiments to investigate contractile function showed that myofiber contractility was not affected. These findings indicate that the structural remodeling associated with LMM mutations induces a pathogenic state in which formation of shutdown heads is impaired, thus increasing myosin head ATP demand in the filaments, rather than affecting contractility. These key findings will help design future therapies for myosinopathies.</p