ViewpointMuscle Wasting: Cellular and Molecular MechanismsCan we talk about myoblast fusion?Hannah F. Dugdale and Julien OchalaHannah F. DugdaleCentre for Human and Applied Physiological Sciences, School of Basic & Medical Biosciences, Faculty of Life Sciences & Medicine, Guy’s Campus, King’s College London, London, United Kingdom and Julien OchalaCentre for Human and Applied Physiological Sciences, School of Basic & Medical Biosciences, Faculty of Life Sciences & Medicine, Guy’s Campus, King’s College London, London, United KingdomRandall Centre for Cell and Molecular Biophysics, School of Basic & Medical Biosciences, Faculty of Life Sciences & Medicine, Guy’s Campus, King’s College London, London, United KingdomDepartment of Biomedical Sciences, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, DenmarkPublished Online:17 Aug 2021https://doi.org/10.1152/ajpcell.00187.2021This is the final version - click for previous versionMoreSectionsPDF (987 KB)Download PDF ToolsExport citationAdd to favoritesGet permissionsTrack citations ShareShare onFacebookTwitterLinkedInWeChat Currently, there is no clinically approved treatment for genetic muscle disorders. As such, further understanding of the exact complex mechanisms that lead to such diseases are both warranted and essential for the design of potential therapeutic interventions.Muscle is not a simple and isolated organ, it constitutes ∼70% of total body mass and is arranged into bundles of multinucleated postmitotic fibers. Consequently, these fibers rely on specific progenitors to facilitate development, growth, and repair (1). The regulation and/or alteration of these progenitors have been widely studied in the context of genetic muscle diseases and in relation to activation, proliferation, and differentiation (2, 3). Surprisingly, fusogenic processes remain broadly underrepresented in comparison and as such, we have had to make some assumptions regarding myoblast fusion and resultant myonuclear numbers. Membrane fusion is an essential biological process required for life. During membrane fusion, two independent lipid membranes merge to create a singular continuous bilayer, which facilitates intracellular trafficking, organ formation, and even infection by enveloped viruses (4, 5). Cell-cell fusion is required for the development of multicellular organisms via the joining of cells (as the name suggests) during the formation of organs and importantly fertilization (4). Conversely, aberrant fusion can exacerbate and even cause a plethora of disorders including preeclampsia, cancer (6), and more recently one genetic muscle disorder (7). The latter, named Carey–Fineman–Ziter syndrome (CFZS), is associated with mutations in the MYMK gene encoding for a protein essential for myoblast fusion, myomaker (8).Myomaker (mymk) is the first skeletal muscle-specific protein identified as essential for myoblast fusion (8, 9). Mymk knockout (KO) models produce mononucleated fibers in zebrafish and mice, which causes paralysis at birth in the latter (8, 10). Clinically, all patients with CFZS present with facial dysmorphisms and muscle hypotonia. To date, seven missense mutations have been discovered (Fig. 1). Although two are yet to be categorized (c.235T > C and c.399 + 5G > A), the classification of the remaining five was carried out via overexpression in fibroblasts that were cocultured with C2C12 myoblasts (7, 12–14). As no fusogenic activity was observed in three of these mutations, they were consequently characterized as null (c.553T>C, c.298G>A, and c.2T>A). The final two mutations either retained residual fusion or displayed a reduced fusogenic capacity (c.271C>A and c.461T>C, respectively) ultimately resulting in the classification; hypomorphic. Unsurprisingly, the fusogenic perturbation present in these individuals results in the formation of fewer myofibers when compared with healthy individuals (12). These patients with Carey–Fineman–Ziter syndrome are a direct example of the involvement of myoblast fusion in the pathogenic roots of one specific disease. What about other genetic muscle disorders? Is there an indirect disruption of fusogenic processes that research studies have missed and may contribute to the complex pathomechanisms? Is it more common than one thinks? All these questions need answering.Figure 1.The top image is a schematic representation depicting stages of cell-cell fusion (cell recognition, hemifusion, pore formation, pore expansion, and cytosolic mixing). Here, two myoblast cells (one in pink and one in green) mix their phospholipid bilayer and ultimately form an early multinucleated myotube. The bottom right focuses on the location for myomaker, dysferlin, and myomerger/minion/myomixer based on a previously tentatively proposed model (11). The bottom left is a model of the transmembrane topology of myomaker adapted from the studies by Di Gioia et al. (7). Myomaker is 221 amino acids in length and possesses seven transmembrane domains, which are embedded in the plasma membrane. The N-terminal exits into the extracellular space and the C-terminal to the intracellular space (a region that is essential for both fusion and mymk trafficking). M, approximate placement of the denoted mutations.Download figureDownload PowerPointTo highlight the importance of myoblast fusion and its underappreciation, we here give examples of two classes of disorders, i.e., limb-girdle muscular dystrophy and centronuclear myopathy. Both groups of muscle disorders are considered clinically and genetically heterogeneous and are characterized by muscle weakness and atrophy (15, 16). The primary mechanisms underlying DYSF-associated limb-girdle muscular dystrophy (LGMD2B), include altered dysferlin protein production resulting in defective membrane resealing (17). In addition, when the mutation is null, it encompasses a reduction in the incorporation of nuclei into myotubes that do not fuse properly and produce small fibers (18, 19). As dysferlin 1) upregulates myogenin, a binding partner of the MYMK promoter; and 2) interacts with myomerger [a membrane micropeptide, also known as myomixer or minion, found to be essential for myoblast fusion; with a primary involvement in pore formation (11)]. It is, therefore, reasonable to postulate that fusogenic impairments play an important role in the development of this complex disease. Similar inferences can be drawn for mouse models of CAV3-related limb-girdle muscular dystrophy (LGMD1C). Here, caveolin-3, a principal structural component of vesicular invaginations of the plasma membrane (known as caveolae) is affected (20). Similar to LGMD2B, the number of nuclei per muscle fiber following fusion analysis in culture also display impairments (21).The potential fusogenic role of dynamin-2 in the context of DMN2-linked congenital myopathies appears more complex than the aforementioned mutated fusion-associated proteins. Dynamin-2 is a large, ubiquitously expressed GTPase involved in, but not limited to vesicle release, actin cytoskeletal assembly, membrane fission, and nuclear positioning. Dynamin-2 has been linked to the pathophysiology of three classical forms of centronuclear myopathy (not merely DNM2-related centronuclear myopathy; 22–26). As such, the aim of phase 1 clinical trial using an antisense oligonucleotide-mediated Dynamin-2 knockdown (DYN101) is to reverse this (27, 28). However, in C2C12 and primary myoblasts where dynamin-2 is absent, significant abrogation of myoblast fusion was inferred from the reduced formation of multinucleated myofibers (29, 30). These findings should be carefully considered as reducing dynamin-2 expression may impair fusogenic capacity and therefore, induce an alternate muscle dysfunction in these patients. Moreover, these findings may also indicate a potential role for fusogenic proteins, which despite their upregulation does not lead to fusion per se and instead initiates or intensifies muscle dysfunction. This phenomenon has also been highlighted in Duchenne muscular dystrophy (the most common and severe form of genetic muscle disorder). Here, myomaker is required for progenitor fusion due to the presence of chronic muscle regeneration. Interestingly and unexpectedly however, upregulation of myomaker in diseased myofibers has been shown to contribute to the pathophenotype (31). Whether a secondary mechanism is present or not, these findings are indicative of a tightly regulated temporal expression for fusogenic proteins whereby dysregulation leads to dysfunctional musculature.Here, we have presented data outlining the complex role of fusogenic mechanisms in generating primary phenotype or exacerbating known symptoms of genetic muscle diseases. Despite these promising findings, our current understanding of fusion and resulting muscle phenotype is limited and inferred through experimentation in animal and cell culture models. Going forward, we believe that investigators should thoroughly study myoblast fusion further, not only by investigating myonuclear number but also by dissecting the different stages of myoblast fusion, as outlined in Fig. 1. In addition, direct measurements of known fusogenic markers including myomaker and myomerger should be established in muscle disease, including the aforementioned myopathies. 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JCI Insight 5: e136095, 2020. doi:10.1172/jci.insight.136095. Crossref | PubMed | ISI | Google ScholarAUTHOR NOTESCorrespondence: J. Ochala (julien.[email protected]ku.dk). Download PDF Previous Back to Top FiguresReferencesRelatedInformation CollectionsAJP-Cell CollectionsMuscle Wasting: Cellular and Molecular Mechanisms Cited ByImpaired activity of the fusogenic micropeptide Myomixer causes myopathy resembling Carey-Fineman-Ziter syndrome1 June 2022 | Journal of Clinical Investigation, Vol. 132, No. 11 More from this issue > Volume 321Issue 3September 2021Pages C504-C506 Crossmark Copyright & PermissionsCopyright © 2021 the American Physiological Society.https://doi.org/10.1152/ajpcell.00187.2021PubMed34288723History Received 20 May 2021 Accepted 7 July 2021 Published online 17 August 2021 Published in print 1 September 2021 Keywordsgenetic muscle disordersmyoblast fusionmyomakerskeletal muscle Metrics