Defective sarcomere organization and reduced larval locomotion and fish survival in slow muscle heavy chain 1 ( smyhc1 ) mutants

Abstract

Zebrafish skeletal muscles are broadly divided into slow-twitch and fast-twitch muscle fibers. The slow fibers, which express a slow fiber-specific myosin heavy chain 1 (Smyhc1), are the first group of muscle fibers formed during myogenesis.To uncover Smyhc1 function in muscle growth, we generated three mutant alleles with reading frame shift mutations in the zebrafish smyhc1 gene using CRISPR. The mutants showed shortened sarcomeres with no thick filaments and M-lines in slow fibers of the mutant embryos. However, the formation of slow muscle precursors and expression of other slow muscle genes were not affected and fast muscles appeared normal. The smyhc1 mutant embryos and larvae showed reduced locomotion and food intake. The mutant larvae exhibited increased lethality of incomplete penetrance. Approximately 2/5 of the homozygous mutants were viable and grew into reproductive adults. These adult mutants displayed a typical pattern of slow and fast muscle fiber distribution, and regained normal slow muscle formation. Together, our studies indicate that Smyhc1 is essential for myogenesis in embryonic slow muscles, and loss of Smyhc1 results in defective sarcomere assembly, reduces larval motility and fish survival, but has no visible impact on muscle growth in juvenile and adult zebrafish that escape the larval lethality. K E Y W O R D Sknockout, sarcomere, slow fibers, smyhc1, zebrafish | 1379 LI et aL.

MYH 3‐associated distal arthrogryposis zebrafish model is normalized with para‐aminoblebbistatin

“…Our results not only confirm that the smyhc1 R673H allele is more severe than a smyhc1 null allele (which was also recently described in Li et al , 2020), but also demonstrate the dosage sensitivity of the missense allele to the wild‐type allele, which modulates the effects of the disease‐causing R673H variant in our model system. Expression of the smyhc1 R673H allele in the absence of a wild‐type allele resulted in embryos with a lethal phenotype identical to smyhc1 R673H/R673H embryos.…”

“…Furthermore, in cultured cells, MYH3 expression also activates TGF‐β signaling pathways that are associated with bone fusions in other disease models (Zieba et al , 2017). Interestingly, we also note that smyhc1 is expressed in zebrafish somites at the 5–9 somite stage even prior to muscle development (Rauch et al , 2003; Bessarab et al , 2008; Li et al , 2020); its role at this early time period is unknown but may indicate a function independent from its role in muscle. More work is needed to understand the how MYH3 mediates the complex developmental relationship between muscle and bone.…”

“…To address the need for vertebrate models of DA, we created a zebrafish line in which the common human DA2A MYH3 mutation, R672H, was precisely edited into the corresponding amino acid of the slow myosin heavy chain 1 gene ( smyhc1 R673H [stl583]) (Roy et al , 2001). This gene is expressed in the somites as early as 5–9 somite stages, and in the lateral slow muscle of the trunk, as evidenced by previously conducted in situ hybridization (Rauch et al , 2003; Bessarab et al , 2008; Li et al , 2020). We simultaneously created a smyhc1 null allele ( smyhc1 − [stl582]), which allowed us to compare the effects of both mutant alleles on slow skeletal muscle and bone development.…”

“…Because smyhc1 is a myosin heavy chain gene critical for motor function, and because of the observed effects of smyhc1 mutations on gross anatomy described above, we assessed the effects of smyhc1 mutations on muscle function by quantifying the movement of fish at various times during development. The smyhc1 gene is expressed embryonically, as early as 10‐13 somites as evidenced by in situ hybridization (Rauch et al , 2003; Bessarab et al , 2008; Li et al , 2020); therefore, we first assessed embryonic movements by counting the number of light‐induced twitches beginning at 24 hpf when embryos first respond to light stimulation (Kokel & Peterson, 2011; Saint‐Amant & Drapeau, 1998; Fig 5A). The smyhc1 −/− embryos were completely paralyzed up to 48 hpf, after which they exhibited normal movements in response to light stimulation.…”

Genome-Wide Identification, Characterization and Expression Profiling of myosin Family Genes in Sebastes schlegelii

“…One early study was carried out to demonstrate the MYH classification, evolution, and expression in fish [19]. More recent studies focused on some specific members of myosin and characterized their expression and function in fish [20][21][22][23]. Despite the great process of characterization of myosin genes in teleost fish, the genomewide phylogenetic identification and characterization of complete myosin family genes in teleost is still poorly conducted.…”