title: β-catenin is essential for efficient in vitro premyogenic mesoderm formation but can be partially compensated by retinoic acid signalling.
abstract: Previous studies have shown that P19 cells expressing a dominant negative β-catenin mutant (β-cat/EnR) cannot undergo myogenic differentiation in the presence or absence of muscle-inducing levels of retinoic acid (RA). While RA could upregulate premyogenic mesoderm expression, including Pax3/7 and Meox1, only Pax3/7 and Gli2 could be upregulated by RA in the presence of β-cat/EnR. However, the use of a dominant negative construct that cannot be compensated by other factors is limiting due to the possibility of negative chromatin remodelling overriding compensatory mechanisms. In this study, we set out to determine if β-catenin function is essential for myogenesis with and without RA, by creating P19 cells with reduced β-catenin transcriptional activity using an shRNA approach, termed P19[shβ-cat] cells. The loss of β-catenin resulted in a reduction of skeletal myogenesis in the absence of RA as early as premyogenic mesoderm, with the loss of Pax3/7, Eya2, Six1, Meox1, Gli2, Foxc1/2, and Sox7 transcript levels. Chromatin immunoprecipitation identified an association of β-catenin with the promoter region of the Sox7 gene. Differentiation of P19[shβ-cat] cells in the presence of RA resulted in the upregulation or lack of repression of all of the precursor genes, on day 5 and/or 9, with the exception of Foxc2. However, expression of Sox7, Gli2, the myogenic regulatory factors and terminal differentiation markers remained inhibited on day 9 and overall skeletal myogenesis was reduced. Thus, β-catenin is essential for in vitro formation of premyogenic mesoderm, leading to skeletal myogenesis. RA can at least partially compensate for the loss of β-catenin in the expression of many myogenic precursor genes, but not for myoblast gene expression or overall myogenesis.
authors: Department of Biochemistry, Faculty of Medicine, University of Ottawa, Ottawa, Ontario, Canada.
authors: Jacob
authors: J
authors: Wong
authors: Virja
authors: V
authors: Mehta
authors: Anastassia
authors: A
authors: Voronova
authors: Josée
authors: J
authors: Coutu
authors: Tammy
authors: T
authors: Ryan
authors: Michael
authors: M
authors: Shelton
authors: Ilona S
authors: IS
authors: Skerjanc
mesh_terms: D000818:Animals; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects; D008648:Mesoderm / Q000187:drug effects / Q000254:growth & development* / Q000378:metabolism; D051379:Mice; D024510:Muscle Development / Q000187:drug effects / Q000235:genetics; D018482:Muscle, Skeletal / Q000187:drug effects / Q000254:growth & development / Q000378:metabolism; D032446:Myoblasts / Q000187:drug effects / Q000378:metabolism; D018006:Myogenic Regulatory Factors / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding / Q000187:drug effects / Q000235:genetics; D034741:RNA, Small Interfering / Q000378:metabolism; D055760:SOXF Transcription Factors / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects / Q000235:genetics; D014158:Transcription, Genetic / Q000187:drug effects; D014212:Tretinoin / Q000494:pharmacology; D060449:Wnt Signaling Pathway / Q000187:drug effects / Q000235:genetics; D051176:beta Catenin / Q000235:genetics / Q000378:metabolism*
chemical_list: D018006:Myogenic Regulatory Factors; D034741:RNA, Small Interfering; D055760:SOXF Transcription Factors; C102515:Sox7 protein, mouse; D051176:beta Catenin; D014212:Tretinoin; D004121:Dimethyl Sulfoxide