A molecular signature defining exercise adaptation with ageing and in vivo partial reprogramming in skeletal muscle

Abstract

Exercise promotes functional improvements in aged tissues, but the extent to which it simulates partial molecular reprogramming is unknown. Using transcriptome profiling from (1) a skeletal muscle-specific in vivo Oct3/4, Klf4, Sox2 and Myc (OKSM) reprogramming-factor expression murine model; (2) an in vivo inducible muscle-specific Myc induction murine model; (3) a translatable high-volume hypertrophic exercise training approach in aged mice; and (4) human exercise muscle biopsies, we collectively defined exercise-induced genes that are common to partial reprogramming. Late-life exercise training lowered murine DNA methylation age according to several contemporary muscle-specific clocks. A comparison of the murine soleus transcriptome after late-life exercise training to the soleus transcriptome after OKSM induction revealed an overlapping signature that included higher JunB and Sun1. Also, within this signature, downregulation of specific mitochondrial and muscle-enriched genes was conserved in skeletal muscle of long-term exercise-trained humans; among these was muscle-specific Abra/Stars. Myc is the OKSM factor most induced by exercise in muscle and was elevated following exercise training in aged mice. A pulse of MYC rewired the global soleus muscle methylome, and the transcriptome after a MYC pulse partially recapitulated OKSM induction. A common signature also emerged in the murine MYC-controlled and exercise adaptation transcriptomes, including lower muscle-specific Melusin and reactive oxygen species-associated Romo1. With Myc, OKSM and exercise training in mice, as well habitual exercise in humans, the complex I accessory subunit Ndufb11 was lower; low Ndufb11 is linked to longevity in rodents. Collectively, exercise shares similarities with genetic in vivo partial reprogramming. KEY POINTS: Advances in the last decade related to cellular epigenetic reprogramming (e.g. DNA methylome remodelling) toward a pluripotent state via the Yamanaka transcription factors Oct3/4, Klf4, Sox2 and Myc (OKSM) provide a window into potential mechanisms for combatting the deleterious effects of cellular ageing. Using global gene expression analysis, we compared the effects of in vivo OKSM-mediated partial reprogramming in skeletal muscle fibres of mice to the effects of late-life murine exercise training in muscle. Myc is the Yamanaka factor most induced by exercise in skeletal muscle, and so we compared the MYC-controlled transcriptome in muscle to Yamanaka factor-mediated and exercise adaptation mRNA landscapes in mice and humans. A single pulse of MYC is sufficient to remodel the muscle methylome. We identify partial reprogramming-associated genes that are innately altered by exercise training and conserved in humans, and propose that MYC contributes to some of these responses.

Keywords: DNA methylation; MYC; Yamanaka factors; ageing.

Figure 1

Comparison of Oct3/4, Klf4, Sox2, and Myc (OKSM) expression to late-life progressive weighted wheel running (PoWeR) in mouse muscle A DNA methylation age (mDNAge) derived from clocks developed by the Horvath laboratory applied to late-life PoWeR (n=10) and sedentary control (n=9) mouse gastrocnemius muscle B Principal component analysis (PCA) plots showing DESeq2-normalized gene count transcriptome data from OKSM expression (n=2) versus control (n=2) in mouse soleus muscle fibres (Wang et al., 2021) and the late-life PoWeR (n=4) versus the sedentary (n=5) transcriptome in soleus muscle (Dungan et al., 2022a) C Chord diagram showing significantly downregulated genes common across OKSM and late-life PoWeR in the soleus D Upregulated genes common across OKSM and late-life PoWeR in the soleus E Downregulated genes common across OKSM, late-life PoWeR, and long-term exercise training. All data reported as mean ± standard deviation

Figure 2

Identification of MYC-controlled genes in soleus muscle that also associate with late-life exercise and OKSM induction A Data from the MetaMEx human muscle and exercise meta-analysis tool from the Zierath laboratory (Pillon et al., 2020) showing the responsiveness of Oct3/4, Klf4, Sox2, and Myc to endurance and resistance exercise in humans. Input parameters: Male/Female, Young, Sedentary/Active/Athlete, Lean, Vastus Lateralis, Healthy, Immediate/0/3/4/5/6/8/18/24 hours. Adjusted p values are for each individual gene and condition. B Lisa analysis of transcription factors (TFs) controlling upregulated genes in the PoWeR-trained transcriptome C Myc gene expression in the soleus after 8 weeks of PoWeR from 22–24 months in mice (Dungan et al., 2022a) D Western blot showing MYC protein in skeletal muscle and heart 12 hours after removal of overnight doxycycline in biological replicate MYC and control mice, with corresponding Ponceau S stain beneath E PCA plot of full soleus DESeq2 normalized gene count transcriptome dataset from MYC (n=3) and control (n=4) mice F Telomerase complex genes controlled by MYC in the soleus (SOL) (n=3 control, n=4 MYC) and quadriceps (QUAD) (n=2 control, n=2 MYC) muscles G Upregulated genes in the soleus common to MYC and late-life PoWeR H Downregulated genes in the soleus common to MYC and late-life PoWeR I Gene expression signature common to MYC, PoWeR, and OKSM in the murine soleus. All data reported as mean ± standard deviation

Figure 3

A pulse of MYC alters the DNA methylome in skeletal muscle A Soleus muscle DNA methylation in promoter, exon, and intron regions of HSA-Myc (n=4) relative to control mice (n=3), p<0.0005 and 10x coverage per CpG site for MYC methylation data B Promoter CpG DNA methylation and C gene expression of Cfap298, Mitol (Marchf5), and Mettl1 after a pulse of MYC, p<0.0005 and 10x coverage per CpG site for MYC methylation data D Common gene-level intron hypo- and E hyper-methylation with MYC induction in the soleus compared to soleus myonuclear DNA methylation after 8 weeks of late-life PoWeR (Dungan et al., 2022a).*=multiple sites of regulation in one or both conditions, but only data from one site is shown. All data reported as mean ± standard deviation