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Title: The role of MEGF10 in skeletal muscle myopathy
Author: Hughes, Ruth Elizabeth
Awarding Body: University of Leeds
Current Institution: University of Leeds
Date of Award: 2016
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Mutations in the transmembrane protein MEGF10 (multiple epidermal growth factor-like protein 10) cause early-onset myopathy, areflexia, respiratory distress and dysphagia (EMARDD), an autosomal recessive congenital myopathy, characterized by reduced muscle fibre size. MEGF10 is reportedly expressed in satellite cells, playing a role in regulating cell proliferation and differentiation. The pattern of expression and the function of MEGF10 have not been clearly described. Cultured myoblasts, isolated single fibres and a muscle hyperplasia model were used to investigate changes in MEGF10 expression. Western blotting, immunofluorescence and RNAseq showed that MEGF10 was expressed at low levels in cultured myoblasts, suggesting that these cells are a poor model for studying endogenous MEGF10. Overexpression of GFP-tagged wildtype MEGF10 and MEGF10 containing pathogenic mutations decreased myoblasts cell motility and fusion. Isolated single fibres showed the presence of quiescent (Pax7+), committed and activated (MyoD+ or myogenin+) satellite cells after 2 - 4 days in culture. However, these cells did not consistently express MEGF10. A muscle hyperplasia model showed no MEGF10 expression in CD34+/Pax7+ or MyoD+ positive satellite cells, but MEGF10+/CD34- cells were present on these fibres. Finally, the expressed and purified extracellular domain (ECD) of MEGF10 was heavily post- translationally modified and facilitated myoblast attachment to a non- adherent surface. Pathogenic mutations in the ECD did not have a significant effect on myoblast fusion, whilst the wildtype ECD inhibited fusion. In conclusion, the choice of skeletal muscle model system is important in obtaining meaningful results. The discovery of MEGF10 expression in, as yet, unidentified CD34- cells on isolated fibres requires further study. MEGF10 is likely to respond to differing cues within the complex muscle environment to regulate myogenesis through cellular changes affecting differentiation and motility. Whilst EMARDD is a rare condition, insights into MEGF10 function may give insight into mechanisms of pathogenesis in a range of skeletal muscle diseases.
Supervisor: Peckham, Michelle ; Johnson, Colin A. Sponsor: MRC
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available