Histone methyltransferase MLL4 controls myofiber identity and muscle performance through MEF2 interaction
Lin Liu, Chenyun Ding, Tingting Fu, Zhenhua Feng, Ji-Eun Lee, Liwei Xiao, Zhisheng Xu, Yujing Yin, Qiqi Guo, Zongchao Sun, Wanping Sun, Yan Mao, Likun Yang, Zheng Zhou, Danxia Zhou, Leilei Xu, Zezhang Zhu, Yong Qiu, Kai Ge, Zhenji Gan
Lin Liu, Chenyun Ding, Tingting Fu, Zhenhua Feng, Ji-Eun Lee, Liwei Xiao, Zhisheng Xu, Yujing Yin, Qiqi Guo, Zongchao Sun, Wanping Sun, Yan Mao, Likun Yang, Zheng Zhou, Danxia Zhou, Leilei Xu, Zezhang Zhu, Yong Qiu, Kai Ge, Zhenji Gan
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Research Article Metabolism Muscle biology

Histone methyltransferase MLL4 controls myofiber identity and muscle performance through MEF2 interaction

Abstract

Skeletal muscle depends on the precise orchestration of contractile and metabolic gene expression programs to direct fiber-type specification and to ensure muscle performance. Exactly how such fiber type–specific patterns of gene expression are established and maintained remains unclear, however. Here, we demonstrate that histone monomethyl transferase MLL4 (KMT2D), an enhancer regulator enriched in slow myofibers, plays a critical role in controlling muscle fiber identity as well as muscle performance. Skeletal muscle–specific ablation of MLL4 in mice resulted in downregulation of the slow oxidative myofiber gene program, decreased numbers of type I myofibers, and diminished mitochondrial respiration, which caused reductions in muscle fatty acid utilization and endurance capacity during exercise. Genome-wide ChIP-Seq and mRNA-Seq analyses revealed that MLL4 directly binds to enhancers and functions as a coactivator of the myocyte enhancer factor 2 (MEF2) to activate transcription of slow oxidative myofiber genes. Importantly, we also found that the MLL4 regulatory circuit is associated with muscle fiber–type remodeling in humans. Thus, our results uncover a pivotal role for MLL4 in specifying structural and metabolic identities of myofibers that govern muscle performance. These findings provide therapeutic opportunities for enhancing muscle fitness to combat a variety of metabolic and muscular diseases.

Authors

Lin Liu, Chenyun Ding, Tingting Fu, Zhenhua Feng, Ji-Eun Lee, Liwei Xiao, Zhisheng Xu, Yujing Yin, Qiqi Guo, Zongchao Sun, Wanping Sun, Yan Mao, Likun Yang, Zheng Zhou, Danxia Zhou, Leilei Xu, Zezhang Zhu, Yong Qiu, Kai Ge, Zhenji Gan

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Figure 1

Slow myofiber–enriched MLL4 is required for type I muscle fiber formation.

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Slow myofiber–enriched MLL4 is required for type I muscle fiber formatio...
(A) Representative Western blot analysis of protein extracts prepared from WV and soleus muscles of WT mice using indicated antibodies. Quantification of the MLL4/tubulin, EZH2/tubulin, and SUZ12/tubulin signal ratios normalized (=1.0) to the WV and presented below the corresponding bands. n = 5–6 mice per group. (B) Representative Western blot analysis of MLL4 expression in GC muscles of indicated mice. n.s., nonspecific band. n = 3 mice per group. (C) Top: representative WGA staining of GC muscle from 8-week-old male Mll4-mKO and Mll4SET-mKO mice. Scale bar: 50 μm. Bottom: cross-sectional areas of GC myofibers were measured by ImageJ. n = 4–5 mice per group. (D) Expression of slow-twitch myosin gene (Myh7) and representative slow/fast-twitch troponin genes (qRT-PCR) in GC muscle from indicated genotypes. n = 5–8 mice per group. (E) Cross section of (top) soleus and (bottom) GC muscle from 8-week-old male Mll4-mKO and Mll4SET-mKO mice stained for MHC1 (green) and MHC2b (red). Scale bars: 250 μm. (F) Quantification of IF data shown in E. n = 3–5 mice per group. (G and H) Primary myoblasts isolated from Mll4SETfl/fl mice were infected with an adenovirus overexpressing Cre or control virus (Ctrl), followed by differentiation into myotubes. (G) Results of qRT-PCR and Western blot analysis in skeletal myotubes. n = 3 independent experiments. (H) Left: IF staining of skeletal myotubes was performed using antibodies directed against myosin-slow or myosin-fast. Scale bars: 100 μm. Right: quantification of the myosin-slow IF data expressed as mean percentage of total myotubes. n = 3 independent experiments. Values are represented as mean ± SEM. *P < 0.05 vs. corresponding controls, 2-tailed unpaired Student’s t test.