Genome-wide DNA methylation and transcriptome integration reveal distinct sex differences in skeletal muscle.

Nearly all human complex traits and diseases exhibit some degree of sex differences, and epigenetics contributes to these differences as DNA methylation shows sex differences in various tissues. However, skeletal muscle epigenetic sex differences remain largely unexplored, yet skeletal muscle displays distinct sex differences at the transcriptome level. We conducted a large-scale meta-analysis of autosomal DNA methylation sex differences in human skeletal muscle in three separate cohorts (Gene SMART, FUSION, and GSE38291), totalling n = 369 human muscle samples (n = 222 males, n = 147 females). We found 10,240 differentially methylated regions (DMRs) at FDR < 0.005, 94% of which were hypomethylated in males, and gene set enrichment analysis revealed that differentially methylated genes were involved in muscle contraction and metabolism. We then integrated our epigenetic results with transcriptomic data from the GTEx database and the FUSION cohort. Altogether, we identified 326 autosomal genes that display sex differences at both the DNA methylation, and transcriptome levels. Importantly, sex-biased genes at the transcriptional level were overrepresented among the sex-biased genes at the epigenetic level (p-value = 4.6e-13), which suggests differential DNA methylation and gene expression between males and females in muscle are functionally linked. Finally, we validated expression of three genes with large effect sizes (FOXO3A, ALDH1A1, and GGT7) in the Gene SMART cohort with qPCR. GGT7, involved in muscle metabolism, displays male-biased expression in skeletal muscle across the three cohorts, as well as lower methylation in males. In conclusion, we uncovered thousands of genes that exhibit DNA methylation differences between the males and females in human skeletal muscle that may modulate mechanisms controlling muscle metabolism and health.