Warshaw, D. | bioRxiv Science

bioRxiv2020

Imaging ATP Consumption in Resting Skeletal Muscle: One Molecule at a Time

Muscle contraction is driven by sarcomere shortening and powered by cyclic hydrolysis of ATP by myosin molecular motors. However, myosin in relaxed muscle continues to slowly hydrolyze ATP, analogous to an idling engine. Utilizing super-resolution microscopy to directly image single molecule fluorescent ATP turnover in relaxed rat soleus skeletal muscle sarcomeres, we observed two rates of myosin ATP consumption that differed 5-fold. These distinct hydrolysis rates were spatially segregated, with the slower or "super relaxed" rate localized predominantly to the sarcomere C-zone, where Myosin Binding Protein-C (MyBP-C), a known modulator of muscle contractile function, exists. This super relaxed hydrolysis rate and its location suggest that MyBP-C can sequester myosin motors to regulate muscle metabolism and heat production in resting muscle and force generation upon activation. One Sentence SummarySuper relaxed skeletal muscle myosin is stabilized by Myosin-Binding Protein C as imaged by single ATP molecule consumption.

biophysics↗

bioRxiv2020

Mutations in MYLPF cause a novel segmental amyoplasia that manifests as distal arthrogryposis

We identified ten persons in six consanguineous families with Distal Arthrogryposis (DA) who had congenital contractures, scoliosis, and short stature. Exome sequencing revealed that each affected person was homozygous for one of two different rare variants (c.470G>T, p.(Cys157Phe) or c.469T>C, p.(Cys157Arg)) affecting the same residue of myosin light chain, phosphorylatable, fast skeletal muscle (MYLPF). In a seventh family, a c.487G>A, p.(Gly163Ser) variant in MYLPF arose de novo in a father, who transmitted it to his son. In an eighth family comprised of seven individuals with dominantly-inherited DA, a c.98C>T, p.(Ala33Val) variant segregated in all four persons tested. Variants in MYLPF underlie both dominant and recessively inherited DA. Mylpf protein models suggest that the residues associated with dominant DA interact with myosin whereas the residues altered in families with recessive DA only indirectly impair this interaction. Pathological and histological exam of a foot amputated from an affected child revealed complete absence of skeletal muscle (i.e., segmental amyoplasia). To investigate the mechanism for this finding, we generated an animal model for partial MYLPF impairment by knocking out zebrafish mylpfa. The mylpfa mutant had reduced trunk contractile force and complete pectoral fin paralysis, demonstrating that mylpf impairment most severely affects limb movement. mylpfa mutant muscle weakness was most pronounced in an appendicular muscle and was explained by reduced myosin activity and fiber degeneration. Collectively, our findings demonstrate that partial loss of MYLPF function can lead to congenital contractures, likely as a result of degeneration of skeletal muscle in the distal limb.

genetics↗