CDK4-E2F3 signals enhance oxidative skeletal muscle fiber numbers and function to affect myogenesis and metabolism
Young Jae Bahn, … , Alexandra C. McPherron, Sushil G. Rane
Young Jae Bahn, … , Alexandra C. McPherron, Sushil G. Rane
Published July 3, 2023
Citation Information: J Clin Invest. 2023;133(13):e162479. https://doi.org/10.1172/JCI162479.
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Research Article Metabolism Muscle biology

CDK4-E2F3 signals enhance oxidative skeletal muscle fiber numbers and function to affect myogenesis and metabolism

Abstract

Understanding how skeletal muscle fiber proportions are regulated is vital to understanding muscle function. Oxidative and glycolytic skeletal muscle fibers differ in their contractile ability, mitochondrial activity, and metabolic properties. Fiber-type proportions vary in normal physiology and disease states, although the underlying mechanisms are unclear. In human skeletal muscle, we observed that markers of oxidative fibers and mitochondria correlated positively with expression levels of PPARGC1A and CDK4 and negatively with expression levels of CDKN2A, a locus significantly associated with type 2 diabetes. Mice expressing a constitutively active Cdk4 that cannot bind its inhibitor p16INK4a, a product of the CDKN2A locus, were protected from obesity and diabetes. Their muscles exhibited increased oxidative fibers, improved mitochondrial properties, and enhanced glucose uptake. In contrast, loss of Cdk4 or skeletal muscle–specific deletion of Cdk4’s target, E2F3, depleted oxidative myofibers, deteriorated mitochondrial function, and reduced exercise capacity, while increasing diabetes susceptibility. E2F3 activated the mitochondrial sensor PPARGC1A in a Cdk4-dependent manner. CDK4, E2F3, and PPARGC1A levels correlated positively with exercise and fitness and negatively with adiposity, insulin resistance, and lipid accumulation in human and rodent muscle. All together, these findings provide mechanistic insight into regulation of skeletal muscle fiber–specification that is of relevance to metabolic and muscular diseases.

Authors

Young Jae Bahn, Hariom Yadav, Paolo Piaggi, Brent S. Abel, Oksana Gavrilova, Danielle A. Springer, Ioannis Papazoglou, Patricia M. Zerfas, Monica C. Skarulis, Alexandra C. McPherron, Sushil G. Rane

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Figure 3

Mitochondrial phenotype in Cdk4R/R and Cdk4KO muscle.

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Mitochondrial phenotype in Cdk4R/R and Cdk4KO muscle. (A) Immunofluoresc...
(A) Immunofluorescence showing MyhcIIa expression in TA muscles of Cdk4WT and Cdk4KO mice (n = 5 mice each group). MyhcIIa (green), laminin (red), DAPI (blue). Enlarged images of the areas in the white squares are shown below. Scale bars: 500 μm (top); 200 μm (bottom). (B) RNA-Seq heatmap of soleus muscle transcripts in Cdk4WT and Cdk4KO mice (n = 3 mice each group). Downregulated (Down) mRNAs are indicated with a blue bar. (C) Gene ontology (GO) analysis of downregulated genes in Cdk4WT and Cdk4KO muscle. (D) Histochemical analyses of metabolically active SDH-positive fibers in TA muscles of Cdk4WT and Cdk4KO mice (n = 5 mice each group). Scale bars: 500 μm. (E) Representative electron microscopy images showing mitochondria in Cdk4WT, Cdk4R/R, and Cdk4KO muscle (top). Scale bars: 500 nm. Immunostaining showing cytochrome oxidase–positive fibers in Cdk4WT, Cdk4R/R, and Cdk4KO muscle (bottom) (n = 4–5 mice each group). Scale bars: 200 μm. (F) Mitochondrial DNA copy numbers, (G) ATP content, and (H) citrate synthase activity in QA muscles of Cdk4WT, Cdk4R/R and Cdk4KO mice. (I) mRNA levels of mitochondria markers (Ndufs2,Uqcrc2 and Atp5a1) in QA muscles from Cdk4WT, Cdk4R/R, and Cdk4KO mice (n = 5–6 mice each group). (J) Heatmap of mitochondrial transcripts in Cdk4WT and Cdk4R/R muscle (n = 3 mice each group). (K) Protein expression of Pgc-1α, Uqcrc2, and Myh7 in TA muscles of Cdk4WT, Cdk4R/R, and Cdk4KO mice (n = 3 mice each group). (L) Effects of Cdk4 inhibitor (IDCX) on of mitochondrial genes Tfam, Ndufs2, Atp5a1, and Uqcrc2 in C2C12 myotubes. (M) Muscle endurance in Cdk4WT and Cdk4KO mice. Between 6 and 8 mice per group were used in each experiment, unless mentioned otherwise. Data are shown as the mean ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001, #P < 0.05 vs WT, by 2-tailed Student’s t test.