Myofibre Hyper-Contractility in Horses Expressing the Myosin Heavy Chain Myopathy Mutation, MYH1E321G

Abstract

Myosinopathies are defined as a group of muscle disorders characterized by mutations in genes encoding myosin heavy chains. Their exact molecular and cellular mechanisms remain unclear. In the present study, we have focused our attention on a MYH1-related E321G amino acid substitution within the head region of the type IIx skeletal myosin heavy chain, associated with clinical signs of atrophy, inflammation and/or profound rhabdomyolysis, known as equine myosin heavy chain myopathy. We performed Mant-ATP chase experiments together with force measurements on isolated IIx myofibres from control horses (MYH1E321G−/−) and Quarter Horses homozygous (MYH1E321G+/+) or heterozygous (MYH1E321G+/−) for the E321G mutation. The single residue replacement did not affect the relaxed conformations of myosin molecules. Nevertheless, it significantly increased its active behaviour as proven by the higher maximal force production and Ca2+ sensitivity for MYH1E321G+/+ in comparison with MYH1E321G+/− and MYH1E321G−/− horses. Altogether, these findings indicate that, in the presence of the E321G mutation, a molecular and cellular hyper-contractile phenotype occurs which could contribute to the development of the myosin heavy chain myopathy.

Systemic calcinosis in a Quarter Horse gelding homozygous for a myosin heavy chain 1 mutation

“…No triggering event was identified in our patient. Compared to MYH1 heterozygotes, homozygotes appear to be more commonly affected with IMM and display hypercontractility of type 2X fibers, which may trigger an auto‐immune response via cellular damage and exposure of affected myosin 36 . To date, it remains unclear why some horses with IMM develop systemic calcinosis.…”

Preponderant Myosin Super-Relaxed State In Skeletal Muscle From Endurance Athletes

“…All biopsies were immediately snap frozen in liquid nitrogen and stored at −80°C. At the time of analysis, single muscle fibres were chemically skinned as previously described (Ochala et al, 2021). They were stored at −20°C in a buffer solution containing glycerol and relaxing solution mixed 50/50.…”

“…They were stored at −20°C in a buffer solution containing glycerol and relaxing solution mixed 50/50. As previously published, single myofibres were individually attached to a TEM-grid (Sigma-Aldrich, Grids for transmission electron microscopy, G1403-1VL, UK), which was positioned on a microscopy slide (Thermo Scientific, Superfrost ® Plus, USA) (Ochala et al, 2021). Single nucleotide turnover was measured in flow chambers using fluorescent mant-ATP (250μM) as described previously (Stewart et al, 2010, Ochala et al, 2021).…”

“…As previously published, single myofibres were individually attached to a TEM-grid (Sigma-Aldrich, Grids for transmission electron microscopy, G1403-1VL, UK), which was positioned on a microscopy slide (Thermo Scientific, Superfrost ® Plus, USA) (Ochala et al, 2021). Single nucleotide turnover was measured in flow chambers using fluorescent mant-ATP (250μM) as described previously (Stewart et al, 2010, Ochala et al, 2021). Single muscle fibres with a relaxed sarcomere length of 2.2 μm were incubated in a rigor buffer containing 100 μL mant-ATP buffer for five minutes.…”

“…Single nucleotide turnover was measured in flow chambers using fluorescent mant-ATP (250μM) as described previously (Stewart et al, 2010, Ochala et al, 2021. Single muscle fibres with a relaxed sarcomere length of 2.2 µm were incubated in a rigor buffer containing 100 μL mant-ATP buffer for five minutes.…”