title: Glycogen synthase kinase 3β represses MYOGENIN function in alveolar rhabdomyosarcoma.
abstract: MYOGENIN is a member of the muscle regulatory factor family that orchestrates an obligatory step in myogenesis, the terminal differentiation of skeletal muscle cells. A paradoxical feature of alveolar rhabdomyosarcoma (ARMS), a prevalent soft tissue sarcoma in children arising from cells with a myogenic phenotype, is the inability of these cells to undergo terminal differentiation despite the expression of MYOGENIN. The chimeric PAX3-FOXO1 fusion protein which results from a chromosomal translocation in ARMS has been implicated in blocking cell cycle arrest, preventing myogenesis from occurring. We report here that PAX3-FOXO1 enhances glycogen synthase kinase 3β (GSK3β) activity which in turn represses MYOGENIN activity. MYOGENIN is a GSK3β substrate in vitro on the basis of in vitro kinase assays and MYOGENIN is phosphorylated in ARMS-derived RH30 cells. Constitutively active GSK3β(S9A) increased the level of a phosphorylated form of MYOGENIN on the basis of western blot analysis and this effect was reversed by neutralization of the single consensus GSK3β phosphoacceptor site by mutation (S160/164A). Congruently, GSK3β inhibited the trans-activation of an E-box reporter gene by wild-type MYOGENIN, but not MYOGENIN with the S160/164A mutations. Functionally, GSK3β repressed muscle creatine kinase (MCK) promoter activity, an effect which was reversed by the S160/164A mutated MYOGENIN. Importantly, GSK3β inhibition or exogenous expression of the S160/164A mutated MYOGENIN in ARMS reduced the anchorage independent growth of RH30 cells in colony-formation assays. Thus, sustained GSK3β activity represses a critical regulatory step in the myogenic cascade, contributing to the undifferentiated, proliferative phenotype in alveolar rhabdomyosarcoma (ARMS).
authors.affiliation: Department of Biology, York University, 4700 Keele Street, Toronto, Ontario M3J 1P3, Canada.
authors.forename: M G
authors.initials: MG
authors.lastname: Dionyssiou
authors.affiliation: Department of Biology, York University, 4700 Keele Street, Toronto, Ontario M3J 1P3, Canada.
authors.forename: S
authors.initials: S
authors.lastname: Ehyai
authors.affiliation: Department of Kinesiology, York University, 4700 Keele Street, Toronto, Ontario M3J 1P3, Canada.
authors.forename: E
authors.initials: E
authors.lastname: Avrutin
authors.affiliation: Department of Kinesiology, York University, 4700 Keele Street, Toronto, Ontario M3J 1P3, Canada.
authors.forename: M K
authors.initials: MK
authors.lastname: Connor
authors.affiliation: 1] Department of Biology, York University, 4700 Keele Street, Toronto, Ontario M3J 1P3, Canada [2] Centre for Research in Mass Spectrometry, York University, Toronto, Ontario M3J 1P3, Canada [3] Muscle Health Research Centre, York University, Toronto, Ontario M3J 1P3, Canada [4] Centre for Research in Biomolecular Interactions, York University, Toronto, Ontario M3J 1P3, Canada.
authors.forename: J C
authors.initials: JC
authors.lastname: McDermott
mesh_terms: D000818:Animals; D019556:COS Cells; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D002522:Chlorocebus aethiops; D052283:Creatine Kinase, MM Form / Q000235:genetics / Q000378:metabolism; D004558:Electric Stimulation; D004789:Enzyme Activation; D015971:Gene Expression Regulation, Enzymologic; D015972:Gene Expression Regulation, Neoplastic; D005838:Genotype; D038362:Glycogen Synthase Kinase 3 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D000071679:Glycogen Synthase Kinase 3 beta; D051379:Mice; D009154:Mutation; D018008:Myogenin / Q000235:genetics / Q000378:metabolism*; D015514:Oncogene Proteins, Fusion / Q000235:genetics / Q000378:metabolism; D051761:Paired Box Transcription Factors / Q000235:genetics / Q000378:metabolism; D010641:Phenotype; D010766:Phosphorylation; D011401:Promoter Regions, Genetic; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D018232:Rhabdomyosarcoma, Alveolar / Q000201:enzymology* / Q000235:genetics / Q000473:pathology; D013997:Time Factors; D014158:Transcription, Genetic; D014162:Transfection
chemical_list: C503799:MYOG protein, human; D018008:Myogenin; D015514:Oncogene Proteins, Fusion; C471563:PAX3-FOXO1A fusion protein, human; D051761:Paired Box Transcription Factors; D047428:Protein Kinase Inhibitors; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; C000605507:Gsk3b protein, mouse; D051057:Proto-Oncogene Proteins c-akt; D038362:Glycogen Synthase Kinase 3; D052283:Creatine Kinase, MM Form