TY - JOUR

T1 - Slow myosin heavy chain 1 is required for slow myofibril and muscle fibre growth but not for myofibril initiation

AU - Hau, Hoi Ting A.

AU - Kelu, Jeffrey J.

AU - Ochala, Julien

AU - Hughes, Simon M.

N1 - Funding Information: The mechanisms coordinating the many cell biological processes required for cell growth are poorly understood. For example, in the century since D'Arcy Thompson formulated the problem in print (Thompson, 1917), how cells match their plasma membrane surface area to the volume of their cytoplasm so as to control cell shape, and how cells generate the appropriate quantity of cytoskeletal elements to support the size and shape of the cell remain mysteries. Skeletal muscle fibres may provide an instructive example of cell volume control because a) their form is constrained to an approximate cylinder by their contractile function, b) their cytoplasm is essentially filled with a specialised actomyosin cytoskeleton and c) they are one of the few cell types that can both greatly increase and decrease in size during adult life. Despite these propitious characteristics for insight into the cell size problem, muscle has been hard to analyse because most fibres are multinucleate. Mutation of murine MYH4, encoding MyHC IIB, the major myosin in mouse limb muscles, led to reduced muscle size, but this was accompanied by reduced fibre number, compensatory hypertrophy and an unquantified change in nucleation (Allen et al., 2001). By analysing the early steps in growth of the unusual mononucleate slow fibres of the zebrafish, we reveal the dependence of several other aspects of cell growth on the accumulation of Smyhc1, the major MyHC of these fibres. Smyhc1 mRNA accumulation precisely parallels the initiation of slow muscle fibre terminal differentiation (Hinits and Hughes, 2007; Hinits et al., 2007). Slow muscle fibres assemble myofibrils prior to their migration (Jana Koth and Simon M. Hughes, unpublished) yet, despite lack of their major MyHC, SSFs migrate and orientate on the surface of the myotome in smyhc1 mutants in wild type numbers (Fig. 1 and (Li et al., 2020)). No compensatory accumulation of other MyHC isoforms was detected and myofibrils were greatly reduced. Despite attaining normal fibre length, the cross-sectional area, and thus volume, of SSFs fails to increase in the absence of normal myofibril assembly in smyhc1 mutants (Fig. 5). By 8 dpf, some SSFs become mis-orientated in smyhc1 mutants. Nevertheless, we and others have observed recovery of SSF morphology, function and presumably cell size in smyhc1 mutants once Smyhc2 and Smyhc3 become expressed (Li et al., 2020; Whittle et al., 2020). We hypothesise, therefore, that muscle fibre growth control can be likened to the situation with shopping and bags; the quantity of ‘purchased’ cytoskeleton dictates the extent of expansion of the sarcolemmal ‘bags’. How MyHC content or myofibril assembly is assessed and membrane trafficking and/or osmotic balance thereby controlled is of great interest.This work was funded by a PhD studentship grant 17GRO-PS48-0077 from Muscular Dystrophy UK and Medical Research Council (MRC) grants MR/N021231/1 and MR/W001381/1. SMH is an MRC Scientist with Programme Grant support. Funding Information: This work was funded by a PhD studentship grant 17GRO-PS48-0077 from Muscular Dystrophy UK and Medical Research Council (MRC) grants MR/N021231/1 and MR/W001381/1 . SMH is an MRC Scientist with Programme Grant support. Publisher Copyright: © 2023 The Authors

PY - 2023/7

Y1 - 2023/7

N2 - Slow myosin heavy chain 1 (Smyhc1) is the major sarcomeric myosin driving early contraction by slow skeletal muscle fibres in zebrafish. New mutant alleles lacking a functional smyhc1 gene move poorly, but recover motility as the later-formed fast muscle fibres of the segmental myotomes mature, and are adult viable. By motility analysis and inhibiting fast muscle contraction pharmacologically, we show that a slow muscle motility defect persists in mutants until about 1 month of age. Breeding onto a genetic background marking slow muscle fibres with EGFP revealed that mutant slow fibres undergo terminal differentiation, migration and fibre formation indistinguishable from wild type but fail to generate large myofibrils and maintain cellular orientation and attachments. In mutants, initial myofibrillar structures with 1.67 ​μm periodic actin bands fail to mature into the 1.96 ​μm sarcomeres observed in wild type, despite the presence of alternative myosin heavy chain molecules. The poorly-contractile mutant slow muscle cells generate numerous cytoplasmic organelles, but fail to grow and bundle myofibrils or to increase in cytoplasmic volume despite passive movements imposed by fast muscle. The data show that both slow myofibril maturation and cellular volume increase depend on the function of a specific myosin isoform and suggest that appropriate force production regulates muscle fibre growth.

AB - Slow myosin heavy chain 1 (Smyhc1) is the major sarcomeric myosin driving early contraction by slow skeletal muscle fibres in zebrafish. New mutant alleles lacking a functional smyhc1 gene move poorly, but recover motility as the later-formed fast muscle fibres of the segmental myotomes mature, and are adult viable. By motility analysis and inhibiting fast muscle contraction pharmacologically, we show that a slow muscle motility defect persists in mutants until about 1 month of age. Breeding onto a genetic background marking slow muscle fibres with EGFP revealed that mutant slow fibres undergo terminal differentiation, migration and fibre formation indistinguishable from wild type but fail to generate large myofibrils and maintain cellular orientation and attachments. In mutants, initial myofibrillar structures with 1.67 ​μm periodic actin bands fail to mature into the 1.96 ​μm sarcomeres observed in wild type, despite the presence of alternative myosin heavy chain molecules. The poorly-contractile mutant slow muscle cells generate numerous cytoplasmic organelles, but fail to grow and bundle myofibrils or to increase in cytoplasmic volume despite passive movements imposed by fast muscle. The data show that both slow myofibril maturation and cellular volume increase depend on the function of a specific myosin isoform and suggest that appropriate force production regulates muscle fibre growth.

UR - http://www.scopus.com/inward/record.url?scp=85158831298&partnerID=8YFLogxK

U2 - 10.1016/j.ydbio.2023.04.002

DO - 10.1016/j.ydbio.2023.04.002

M3 - Journal article

C2 - 37121308

AN - SCOPUS:85158831298

VL - 499

SP - 47

EP - 58

JO - Developmental Biology

JF - Developmental Biology

SN - 0012-1606

ER -