Signals of Ezh2, Src, and Akt Involve in myostatin-Pax7 pathways regulating the myogenic fate determination during the sheep myoblast proliferation and differentiation.

Caihong Wei,Fuping Zhao,Lingyang Xu,Hangxing Ren,Li Zhang,Jian Lu,Xiaoning Zhang,Ruizao Liu,Li Li,Lixin Du,Maurilio Sampaolesi

doi:10.1371/journal.pone.0120956

Abstract

Myostatin and Pax7 have been well documented individually, however, the mechanism by which Myostatin regulates Pax7 is seldom reported. Here, based on muscle transcriptome analysis in Texel (Myostatin mutant) and Ujumqin (wild type) sheep across the five fetal stages, we constructed and examined the Myostatin-Pax7 pathways in muscle. Then we validated the signals by RNAi in the proliferating and differentiating sheep myoblasts in vitro at mRNA, protein, and cell morphological levels. We reveal that Myostatin signals to Pax7 at least through Ezh2, Src, and Akt during the sheep myoblast proliferation and differentiation. Other signals such as p38MAPK, mTOR, Erk1/2, Wnt, Bmp2, Smad, Tgfb1, and p21 are most probably involved in the Myostatin-affected myogenic events. Myostatin knockdown significantly reduces the counts of nucleus and myotube, but not the fusion index of myoblasts during cell differentiation. In addition, findings also indicate that Myostatin is required for normal myogenic differentiation of the sheep myoblasts, which is different from the C2C12 myoblasts. We expand the regulatory network of Myostatin-Pax7 pathways and first illustrate that Myostatin as a global regulator participates in the epigenetic events involved in myogenesis, which contributes to understand the molecular mechanism of Myostatin in regulation of myogenesis.

Highlights

Myostatin, a member of the transforming growth factor-β (TGF-β) family, is predominantly expressed and secreted by skeletal muscle and functions as a negative regulator of musclePLOS ONE | DOI:10.1371/journal.pone.0120956 March 26, 2015Ezh2, Src, and Akt Involve in Myostatin-Pax7 Pathways growth
Myostatin signaling through Smad2, Smad3 and Smad4 is regulated by the inhibitory Smad7 via a negative feedback mechanism [23, 24], and the p38 MAPK played an important role in regulation of GDF-8-activated Smad pathways [25]
Our findings expand the network of Myostatin-Pax7 pathways and first suggest that Myostatin as a global regulator may participate in the epigenetic event mediated by Ezh2 and Src for decision of muscle cell fate

Summary

Introduction

A member of the transforming growth factor-β (TGF-β) family, is predominantly expressed and secreted by skeletal muscle and functions as a negative regulator of musclePLOS ONE | DOI:10.1371/journal.pone.0120956 March 26, 2015Ezh, Src, and Akt Involve in Myostatin-Pax Pathways growth. Myostatin signaling through Smad, Smad and Smad is regulated by the inhibitory Smad via a negative feedback mechanism [23, 24], and the p38 MAPK played an important role in regulation of GDF-8-activated Smad pathways [25]. To elucidate comprehensively the mechanism by which Myostatin affects Pax expression to orchestrate the myogenic process, in this article, based on fetal muscle microarray analysis in Texel and Ujumqin sheep, we constructed the Myostatin-Pax pathways by the IPA program, and further swept and validated the potential signals involved in the sheep myoblasts comprehensively. Our findings expand the network of Myostatin-Pax pathways and first suggest that Myostatin as a global regulator may participate in the epigenetic event mediated by Ezh and Src for decision of muscle cell fate

Methods

Results

Discussion

Conclusion