Epigenetics of physical exercise is the study of epigenetic modifications to the cell genome resulting from physical exercise. Environmental factors, including physical exercise, have been shown to have a beneficial influence on epigenetic modifications. Generally, it has been shown that acute and long-term exercise has a significant effect on DNA methylation, an important aspect of epigenetic modifications.
The broader field of epigenetics studies heritable alterations to genes that do not involve changing the DNA sequence itself. The next section briefly discusses two important mechanisms involved in epigenetic modifications.
Epigenetic modifications Epigenetic modifications such as DNA methylation and histone modifications alter DNA accessibility and change chromatin structure, thereby regulating patterns of gene expression. These modifications can be heritable, thus passing from parent to offspring.
As mentioned previously, environmental factors can modulate epigenetic alterations. Factors such as diet, exposure to environmental toxins, and stress have all been shown to play a role in affecting epigenetic modifications, especially by influencing methylation patterns in the DNA. Physical exercise is one such factor that also has been shown to affect methylation and chromatin modifications.
DNA Methylation DNA methylation occurs when a methyl group is covalently attached to the C5 position of a cytosine nucleotide by a DNA methyltransferase (DNMT) enzyme. These dinucleotide repeats where cytosine is followed by an adjunct guanine nucleotide are referred to as CpG sites, with the 'p' indicating a phosphodiester linkage. Parts of the genome that contain a high amount of CpG sites are named CpG islands, and these islands often overlap with many of the core promoter regions found in the human genome. There are also CpG sites that are not a part of CpG islands, and these are constitutively methylated to indicate to transcription machinery to not allow transcription to be initiated there. In contrast, CpG islands are the areas in the genome where CpG sites are usually unmethylated until DNMTs methylate them, resulting in effects on gene expression.
The methylation of CpG islands typically results in the transcriptional silencing of a promoter, which can be done in two ways. Methylation can recruit histone deacetylases (HDAC), which work to form tightly condensed heterochromatin, which is transcriptionally inactive. Methylating a CpG island will also cause the methyl group to wedge into the DNA major groove, effectively blocking binding sites for proteins that can activate transcription.
Post-translational histone modifications
Histones are proteins that help organize DNA. Eight histone proteins assemble into an octomer that DNA wraps around to create a unit called a nucleosome. The ability of a histone to be modified by being marked on its N-terminal tail is essential for modulating gene expression. In its most basic sense, histones can be modified to become more or less condensed by histone deacetylases (HDACs) or histone acetyltransferases (HATs), respectively. When histones are acetylated and become less condensed as a result, it allows for the underlying DNA to be more accessible to be transcribed, increasing gene expression. Thus, acetylated histones can serve as binding sites for important transcription initiation factors and enzymes that have the ability to remodel chromatin. Meanwhile, increased condensation of histones via deacetylation results in the opposite effect, causing decreased gene expression. The regulation of gene expression can have implications on which genes are transcribed, ultimately affecting protein production in the human body.
Though histone tails can be marked in many different ways, four prominent modifications will be discussed in this article: acetylation, methylation (of histones), ubiquination, and phosphorylation.
In addition, methylated histones can both serve as binding sites for certain transcription factors due to their bromodomains and chromodomains, and also prevent the binding of such factors by hiding the recognition site of the transcription factor. Transcription factors, including promoters and enhancers, can subsequently modulate the rate at which DNA is transcribed to RNA, also affecting gene expression.
Muscle and endocrine effects due to physical exercise
Exercise of the human vastus lateralis (a thigh muscle) primarily affects the epigentic status of sites in enhancers within muscle cell chromatin, as described in detail below. In contrast, exercise of this skeletal muscle causes few or no epigenetic alterations in promoters or within genes of muscle cell chromatin.
Skeletal muscles are substantially composed of very large, multinucleated, contractile muscle fiber cells (myocytes). However, considerable numbers of small resident and infiltrating mononuclear cells are also present in skeletal muscles. In terms of volume, myocytes make up the great majority of skeletal muscle. However, in terms of nuclei present in skeletal muscle, myocyte nuclei may be only half of the nuclei present, while nuclei from resident and infiltrating mononuclear cells make up the other half.
Mononuclear cells of skeletal muscle
Mononuclear cells found in skeletal muscle tissue samples from mice and humans can be identified by messenger RNA transcription of cell type markers. Cameron et al. identified nine cell types in skeletal muscles. They include endothelial cells (45% of cells), fibro-adipogenic progenitors (FAPs)(20%), pericytes (14%) and endothelial-like pericytes (4%). Another 9% of mononuclear cells are muscle stem cells, adjacent to muscle fiber cells. Types of lymphoid cells (such as B-cells and T-cells) (3%) and myeloid cells such as macrophages (2%) make up most of the remaining mononuclear cells of skeletal muscle. In addition, Cameron et al. also identified two types of myocyte cells, Type I and Type II. Each of the different types of cells in skeletal muscle was found to express different sets of genes. The median number of genes expressed in each of the nine different cell types was 1,331 genes. When a biopsy is taken from a thigh muscle, however, the biopsy contains all the different cell types. Mixed together, in a biopsy of human thigh skeletal muscle, there are 13,026 to 13,108 genes with detected expression.
The functions of skeletal muscle include producing movement, maintaining body posture, controlling body temperature, and stabilizing joints. Skeletal muscle is also an endocrine organ. Under different physiological conditions, subsets of 654 different proteins as well as lipids, amino acids, metabolites and small RNAs are found in the secretome of skeletal muscles. A secretome is secreted by skeletal muscle cells, either locally (in skeletal muscle interstitium) or into the blood circulation, with a measurable plasma concentration. Secreted proteins often have an endocrine effect.
Skeletal muscle secretome alters with exercise
Williams et al. obtained biopsies of a thigh skeletal muscle (vastus lateralis muscle) of eight 23-year-old, originally sedentary, Caucasian males. Biopsies were taken both before and after a six-week-long endurance exercise training program. The exercise consisted of riding a stationary bicycle for one hour five days a week for six weeks.
Of the 13,108 genes with detected expression in the muscle biopsies, 641 genes were upregulated after endurance training and 176 genes were downregulated. Of the 817 genes whose expression was altered, 531 were identified as being in the secretome of skeletal muscle. Because many of the exercise-regulated genes are identified as secreted, this indicates that much of the effect of exercise has an endocrine rather than metabolic function. The main pathways found to be affected by secreted exercise-regulated proteins were related to cardiac, cognitive, kidney and platelet functions.
Exercise-trained effects are mediated by epigenetic mechanisms
Between 2012 and 2019 at least 25 reports indicated a major role of epigenetic mechanisms in skeletal muscle responses to exercise. Epigenetic alterations often occur by adding methyl groups to cytosines in the DNA or removing methyl groups from the cytosines of DNA, especially at CpG sites within enhancers. Methylations of cytosines can cause the DNA to be compacted into heterochromatin, thus inhibiting access of other molecules to the DNA. Epigenetic alterations also often occur through acetylations or deacetylations of the histone tails within chromatin. DNA in the nucleus generally consists of segments of 146 base pairs of DNA wrapped around eight tightly connected histones (and each histone also has a loose tail) in a structure called a nucleosome and one segment of DNA is connected to an adjacent DNA segment on a nucleosome by linker DNA. When histone tails are acetylated they usually cause loosening of the DNA around the nucleosome, leading to increased accessibility of the DNA.
Exercise-induced regulation of genes in muscles
Gene expression in muscle is largely regulated, as in tissues generally, by regulatory DNA sequences, especially enhancers. Enhancers are non-coding sequences in the genome that activate the expression of distant target genes, by looping around and interacting with the promoters of their target genes (see Figure "Regulation of transcription in mammals"). As reported by Williams et al., the average distance in the loop between the connected enhancers and promoters of muscle genes is 239,000 nucleotide bases.
Exercise-induced alteration to gene expression by enhancer DNA methylation or demethylation
Endurance muscle training alters muscle gene expression by epigenetic DNA methylation or de-methylation of CpG sites within enhancers.
In a study by Lindholm et al., twenty-three individuals who were about 27 years old and sedentary volunteered to have endurance training on only one leg during 3 months. The other leg was used as an untrained control leg. The training consisted of one-legged knee extension training for 3 month (45 min, 4 sessions per week). Skeletal muscle biopsies from the vastus lateralis (a thigh muscle) were taken both before training began and 24 hours after the last training session from each of the legs. The endurance-trained leg, compared to the untrained leg, had significant DNA methylation changes at 4,919 sites across the genome. The sites of altered DNA methylation were predominantly in enhancers. Transcriptional analysis, using RNA sequencing, identified 4,076 differentially expressed genes.
The transcriptionally upregulated genes were associated with enhancers that had a significant decrease in DNA methylation, while transcriptionally downregulated genes were associated with enhancers that had increased DNA methylation. Increased methylation was mainly associated with genes involved in structural remodeling of the muscle and glucose metabolism. Enhancers with decreased methylation were associated with genes functioning in inflammatory or immunological processes and in transcriptional regulation.
Exercise-induced long-term alteration of gene expression by enhancer histone acetylation or deacetylation
As indicated above, after exercise, epigenetic alterations of enhancers alter long-term expression of hundreds of muscle genes. This includes genes producing proteins secreted into the systemic circulation, many of which may act as endocrine messengers. Six sedentary, about 23 years old, Caucasian males provided vastus lateralis (a thigh muscle) biopsies before entering an exercise program (six weeks of 60-minute sessions of riding a stationary cycle, five days per week). Four days after this exercise program was completed, the expression of many genes was persistently epigenetically altered. The alterations included acetylations and deacetylations of the histone tails located in the enhancers controlling the genes with altered expression.