TRAF6 regulates satellite stem cell self-renewal and function during regenerative myogenesis
Sajedah M. Hindi, Ashok Kumar
Sajedah M. Hindi, Ashok Kumar
Published November 30, 2015
Citation Information: J Clin Invest. 2016;126(1):151-168. https://doi.org/10.1172/JCI81655.
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Research Article Muscle biology

TRAF6 regulates satellite stem cell self-renewal and function during regenerative myogenesis

Abstract

Satellite cells are a stem cell population within adult muscle and are responsible for myofiber regeneration upon injury. Satellite cell dysfunction has been shown to underlie the loss of skeletal muscle mass in many acquired and genetic muscle disorders. The transcription factor paired box-protein-7 (PAX7) is indispensable for supplementing the reservoir of satellite cells and driving regeneration in normal and diseased muscle. TNF receptor–associated factor 6 (TRAF6) is an adaptor protein and an E3 ubiquitin ligase that mediates the activation of multiple cell signaling pathways in a context-dependent manner. Here, we demonstrated that TRAF6-mediated signaling is critical for homeostasis of satellite cells and their function during regenerative myogenesis. Selective deletion of Traf6 in satellite cells of adult mice led to profound muscle regeneration defects and dramatically reduced levels of PAX7 and late myogenesis markers. TRAF6 was required for the activation of MAPKs ERK1/2 and JNK1/2, which in turn activated the transcription factor c-JUN, which binds the Pax7 promoter and augments Pax7 expression. Moreover, TRAF6/c-JUN signaling repressed the levels of the microRNAs miR-1 and miR-206, which promote differentiation, to maintain PAX7 levels in satellite cells. We also determined that satellite cell–specific deletion of Traf6 exaggerates the dystrophic phenotype in the mdx (a mouse model of Duchenne muscular dystrophy) mouse by blunting the regeneration of injured myofibers. Collectively, our study reveals an essential role for TRAF6 in satellite stem cell function.

Authors

Sajedah M. Hindi, Ashok Kumar

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Figure 6

TRAF6 regulates PAX7 expression and activation of MAPKs in myogenic cells.

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TRAF6 regulates PAX7 expression and activation of MAPKs in myogenic cell...
(A) Immunoblots demonstrate protein levels of PAX7 and TRAF6 in Traf6+/+ and Traf6–/– cultures. (B) Densitometry quantification of relative protein levels of PAX7 and TRAF6 in Traf6+/+ and Traf6–/– cultures. n = 3 in each group. (C) Relative mRNA levels of Pax7 and Traf6 in Traf6+/+ and Traf6–/– cultures assayed by performing qPCR. n = 4 in each group. (D) Primary myoblasts prepared from WT mice were transfected with scrambled shRNA or TRAF6 shRNA, and the mRNA levels of Pax7 and Traf6 were measured by performing qPCR. (E) Representative immunoblots from 2 independent experiments demonstrate the levels of phosphorylated and total ERK1/2, JNK1/2, p38, ERK5, and c-JUN in Traf6+/+ and Traf6–/– cultures. (F) Densitometry quantification of levels of phosphorylated and total ERK1/2, JNK1/2, p38, ERK5, and c-JUN in Traf6+/+ and Traf6–/– cultures. n = 4–5 in each group. (G) Immunoblots demonstrate the levels of phosphorylated c-JUN and an unrelated protein GAPDH in uninjured and injured TA muscle of Traf6fl/fl and TRAF6scko mice. Error bars represent SD. *P < 0.05 (vs. Traf6+/+ cultures) by unpaired t test. #P < 0.05 (vs. cultures transfected with scrambled shRNA) by unpaired t test. U, uninjured; I, injured.