title: Human myostatin negatively regulates human myoblast growth and differentiation.
abstract: Myostatin, a member of the transforming growth factor-β superfamily, has been implicated in the potent negative regulation of myogenesis in murine models. However, little is known about the mechanism(s) through which human myostatin negatively regulates human skeletal muscle growth. Using human primary myoblasts and recombinant human myostatin protein, we show here that myostatin blocks human myoblast proliferation by regulating cell cycle progression through targeted upregulation of p21. We further show that myostatin regulates myogenic differentiation through the inhibition of key myogenic regulatory factors including MyoD, via canonical Smad signaling. In addition, we have for the first time demonstrated the capability of myostatin to regulate the Notch signaling pathway during inhibition of human myoblast differentiation. Treatment with myostatin results in the upregulation of Hes1, Hes5, and Hey1 expression during differentiation; moreover, when we interfere with Notch signaling, through treatment with the γ-secretase inhibitor L-685,458, we find enhanced myotube formation despite the presence of excess myostatin. Therefore, blockade of the Notch pathway relieves myostatin repression of differentiation, and myostatin upregulates Notch downstream target genes. Immunoprecipitation studies demonstrate that myostatin treatment of myoblasts results in enhanced association of Notch1-intracellular domain with Smad3, providing an additional mechanism through which myostatin targets and represses the activity of the myogenic regulatory factor MyoD. On the basis of these results, we suggest that myostatin function and mechanism of action are very well conserved between species, and that myostatin regulation of postnatal myogenesis involves interactions with numerous downstream signaling mediators, including the Notch pathway.
authors.affiliation: Growth, Development and Metabolism Program, Singapore Institute for Clinical Sciences, Singapore.
authors.forename: Craig
authors.initials: C
authors.lastname: McFarlane
authors.forename: Gu Zi
authors.initials: GZ
authors.lastname: Hui
authors.forename: Wong Zhi Wei
authors.initials: WZ
authors.lastname: Amanda
authors.forename: Hiu Yeung
authors.initials: HY
authors.lastname: Lau
authors.forename: Sudarsanareddy
authors.initials: S
authors.lastname: Lokireddy
authors.forename: Ge
authors.initials: G
authors.lastname: Xiaojia
authors.forename: Vincent
authors.initials: V
authors.lastname: Mouly
authors.forename: Gillian
authors.initials: G
authors.lastname: Butler-Browne
authors.forename: Peter D
authors.initials: PD
authors.lastname: Gluckman
authors.forename: Mridula
authors.initials: M
authors.lastname: Sharma
authors.forename: Ravi
authors.initials: R
authors.lastname: Kambadur
mesh_terms: D000293:Adolescent; D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins / Q000235:genetics / Q000378:metabolism; D002219:Carbamates / Q000494:pharmacology; D002453:Cell Cycle; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism; D002454:Cell Differentiation; D049109:Cell Proliferation; D002478:Cells, Cultured; D002648:Child; D004268:DNA-Binding Proteins / Q000378:metabolism; D004151:Dipeptides / Q000494:pharmacology; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D047468:Immunoprecipitation; D051379:Mice; D024510:Muscle Development; D018485:Muscle Fibers, Skeletal / Q000378:metabolism; D018482:Muscle, Skeletal / Q000254:growth & development* / Q000378:metabolism; D017570:MyoD Protein / Q000378:metabolism; D032446:Myoblasts / Q000166:cytology* / Q000378:metabolism*; D018006:Myogenic Regulatory Factors / Q000378:metabolism; D055435:Myostatin / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D051880:Receptors, Notch / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D051785:Smad Proteins / Q000378:metabolism; D000072056:Transcription Factor HES-1; D016212:Transforming Growth Factor beta; D054462:p21-Activated Kinases / Q000235:genetics / Q000378:metabolism
chemical_list: D051792:Basic Helix-Loop-Helix Proteins; D002219:Carbamates; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D004151:Dipeptides; D018398:Homeodomain Proteins; D017570:MyoD Protein; D018006:Myogenic Regulatory Factors; D055435:Myostatin; D051880:Receptors, Notch; D011994:Recombinant Proteins; D012097:Repressor Proteins; D051785:Smad Proteins; D000072056:Transcription Factor HES-1; D016212:Transforming Growth Factor beta; D054462:p21-Activated Kinases; C485631:HEY1 protein, human; C410009:L 685458; C077478:HES5 protein, human; C078493:HES1 protein, human