Non-Canonical Functions of SMAD2 and SMAD3 During Myogenic Differentiation and Fusion

• Main Author: Lamarche, Emilie
• Other Authors: Wiper, Nadine Louise
• Format: Others
• Language: en
• Published: Université d'Ottawa / University of Ottawa 2018
• Subjects: Myogenesis; Differentiation; TGFβ; SMAD2; SMAD3
• Online Access: http://hdl.handle.net/10393/37322; http://dx.doi.org/10.20381/ruor-21594

The transcription factors SMAD2 and SMAD3 are the effectors of classical transforming growth factor beta (TGFβ) signalling. This signalling cascade is involved in many cellular processes including proliferation and differentiation and is known to be a potent inhibitor of myogenic differentiation through SMAD3. We have previously shown that retinoic acid (RA) can upregulate SMAD3 in models of adipogenesis and mesenchymal stem cells and that SMAD3 can interact with the bZIP transcription factor C/EBPβ to disrupt its DNA binding. Forced expression of C/EBPβ inhibits myogenic differentiation but the mechanism has not been fully elucidated. Herein we show that RA increases Smad3 expression in myoblasts and that RA treatment antagonizes TGFβ-mediated inhibition of myogenic differentiation. TGFβ treatment increased C/EBPβ expression which was reversed by RA treatment. Further, RA was able to disrupt C/EBPβ occupancy of the Pax7 and Smad2 promoters in myoblasts. Loss of C/EBPβ in primary myoblasts using a conditional knockout model partially protected these cells from the anti-myogenic effects of TGFβ treatment. The TGFβ effector protein SMAD2 is expressed in myoblasts but its specific function in myogenesis has not been determined, as Smad2 knockout models are embryonic lethal. Thus, we created a novel Smad2 conditional knockout model where Smad2 is excised in PAX7-expressing muscle satellite cells. Herein we demonstrate a role for SMAD2 specifically in myogenic fusion. We describe a regeneration defect after acute injury and decreased fiber cross-sectional area at P21 (post-natal day 21) in Smad2cKO muscle, without affecting the numbers of PAX7-positive cells. Further, we reveal a mechanism whereby SMAD2 regulates KLF4 expression and mediates the KLF4-induced increased of the fusion gene Npnt. This work describes the pro-myogenic actions of RA-induced SMAD3 and the novel function of SMAD2 in terminal myogenic differentiation and fusion. This work also discusses future directions, implications and new insights into non-canonical SMAD actions.