Abstract
Tendons are an essential part of the musculoskeletal system, connecting muscle and skeletal elements to enable force generation. The transcription factor scleraxis marks vertebrate tendons from early specification. Scleraxis-null mice are viable and have a range of tendon and bone defects in the trunk and limbs but no described cranial phenotype. We report the expression of zebrafish scleraxis orthologs: scleraxis homolog (scx)-a and scxb in cranial and intramuscular tendons and in other skeletal elements. Single mutants for either scxa or scxb, generated by clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9), are viable and fertile as adult fish. Although scxb mutants show no obvious phenotype, scxa mutant embryos have defects in cranial tendon maturation and muscle misalignment. Mutation of both scleraxis genes results in more severe defects in cranial tendon differentiation, muscle and cartilage dysmorphogenesis and paralysis, and lethality by 2–5 wk, which indicates an essential function of scleraxis for craniofacial development. At juvenile and adult stages, ribs in scxa mutants fail to mineralize and/or are small and heavily fractured. Scxa mutants also have smaller muscle volume, abnormal swim movement, and defects in bone growth and composition. Scleraxis function is therefore essential for normal craniofacial form and function and vital for fish development.—Kague, E., Hughes, S. M., Lawrence, E. A., Cross, S., Martin-Silverstone, E., Hammond, C. L., Hinits, Y. Scleraxis genes are required for normal musculoskeletal development and for rib growth and mineralization in zebrafish.
Highlights
The musculoskeletal system is formed by coordinated differentiation and morphogenesis of skeletal muscle, tendon, ligament, cartilage, bone, and associated joint cell
Whereas loss of scxb alone does not lead to severe phentoypes, scxb is required in the absence of scxa because loss of both leads to lethal jaw paralysis
In addition to confirming the expression of scxa in the craniofacial tendon precursors and the myotendinous junctions (MTJ) [23, 24], we demonstrate that scxa mRNA is expressed in the full extent of the vertical myosepta at juvenile stages
Summary
In the integrated musculoskeletal system, resolving the primary cause of defects requires identification of the earliest failures—in muscle, bone, or tendon—and determination of their secondary consequences during development of the entire system. With such detailed understanding, treatments for genetic or environmentally induced musculoskeletal pathologies may be more effective. Knockdown experiments in zebrafish with antisense morpholino oligonucleotides against both scxa and scxb were reported to have no effect on tnmd expression or create any craniofacial defects in the embryos [23]. Expression data implicate Scx in tendon development in various vertebrate groups, functional data derive mostly from mice trunk and limbs. Double mutant embryos have reduced muscle growth and function and paralysis of the jaw, leading to death at early juvenile stages
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