Muscle type-specific RNA polymerase II recruitment during PGC-1α gene transcription after acute exercise in adult rats.

Epigenetic regulation of gene expression differs between fast- and slow-twitch skeletal muscles in adult rats, although the precise mechanisms are still unknown. The present study investigates the differences in responses of RNA polymerase II (Pol II) and histone acetylation during transcriptional activation in the plantaris and soleus muscles of adult rats after acute treadmill running. We targeted the peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) gene to analyze epigenomic changes by chromatin immunoprecipitation. The mRNA expression of the PGC-1α-b isoform was significantly upregulated in both plantaris and soleus muscles 2 h after acute running, although the magnitude of the upregulation was more pronounced in the plantaris muscle. The sequences of proximal exons of the PGC-1α locus were expressed more in the plantaris muscle after acute running. Accumulation of Pol II was noted near the alternative exon 1 in both plantaris and soleus muscles in association with the enhanced distribution of acetylated histone 3. Accumulation of Pol II was also observed at the transcription start site, exon 2, and exon 3 in the plantaris muscle, but not the soleus muscle. It was noted that in the soleus muscle, acetylation of histone 3 at lysine 27 was enhanced throughout the PGC-1α locus in response to transcriptional activation, suggesting that elongating Pol II was capable of traveling through to the end of the locus. These results indicate that the mobility of Pol II during PGC-1α transcription differed between fast- and slow-twitch skeletal muscles, affecting the strength of the transcriptional activity.NEW & NOTEWORTHY Fast- and slow-twitch skeletal muscles have distinct characteristics in both force production and metabolism. Epigenetic regulations also largely differ in these muscles. Here we show that RNA polymerase II is distributed extensively at the proximal regions downstream of transcription start site during the transcriptional activation of PGC-1α in fast-twitch muscles, but it accumulates at the first exon in slow-twitch muscles. These findings will provide a basis to understand type-specific mechanisms in skeletal muscle.

Amount of daily exercise is an essential stimulation to alter the epigenome of skeletal muscle in rats.

Long-term running training causes epigenetic changes in the skeletal muscles. Here we tested the effects of the total amount or duration of running training on the distribution of histones in the rat plantaris muscle. Post-weaned young rats were assigned to 3 different training groups: Run-1, 30 min/day running exercise for 8 wk using an animal treadmill at 24 m/min; Run-2, 15 min/day for 8 wk; and Run-3, 60 min/day for 4 wk. Citrate synthase activity was not significantly changed by running training, although the slight increase was observed in Run-3. Genes that were previously defined as showing the typical responses to running training were targeted to measure the distribution of histones using chromatin immunoprecipitation. The distribution of acetylated histone 3 was elevated in Run-2 and Run-3, but not in Run-1. Incorporation of H3.3 into the nucleosome was stimulated in Run-1, whereas H3.3 distribution was unchanged in Run-2 or downregulated in Run-3. Significant downregulation of H3.3 expression was also detected in Run-3. We further checked the responses of the target genes during acute running. Target genes were transcriptionally activated and histone acetylation was stimulated at the loci in response to acute running. These results suggested that the exchange of the histone component to H3.3 was stimulated by running training, inhibiting the accumulation of acetylated histones in Run-1. Additionally, it was further suggested that the enhanced daily amount of running caused changes in the H3.3 expression, affecting the rate of the histone exchange in Run-3. NEW & NOTEWORTHY Chromatin remodeling in the skeletal muscle is a potent mechanism preventing disuse atrophy in later life that can be acquired via long-term exercise training. Here we demonstrated in rats that daily exercise amount is a key factor in the development of epigenetic changes in the skeletal muscle. To acquire a health benefit, our research suggests the importance of considering the time endurance for daily exercise bouts.

Running training experience attenuates disuse atrophy in fast-twitch skeletal muscles of rats.

Responsiveness to physiological stimuli, such as exercise and muscular inactivation, differs in individuals. However, the mechanisms responsible for these individual differences remain poorly understood. We tested whether a prior experience of exercise training affects the responses of skeletal muscles to unloading. Young rats were assigned to perform daily running training with a treadmill for 8 wk. After an additional 8 wk of normal habitation, the rats were hindlimb unloaded by tail suspension for 1 wk. Fast-twitch plantaris, gastrocnemius, and tibialis anterior muscles did not atrophy after unloading in rats with training experience, although soleus muscle lost weight similar to sedentary rats. We also analyzed the transcriptome in plantaris muscle with RNA sequencing followed by hierarchical clustering analysis and found that a subset of genes that were generally upregulated in sedentary rats after unloading were less responsive in rats with training experience. The distribution of histone 3 was diminished at the loci of these genes during the training period. Although the deposition of histone 3 was restored after an additional period of normal habitation, the incorporation of H3.3 variant was promoted in rats with training experience. This remodeling of nucleosomes closely correlated to the conformational changes of chromatin and suppressed gene expression in response to unloading. These results suggest that exercise training stimulated the early turnover of histone components, which may alter the responsiveness of gene transcription to physiological stimuli.NEW & NOTEWORTHY The present study demonstrates that disuse atrophy was suppressed in fast-twitch skeletal muscles of rats with training experience in early life. We also found a subset of genes that were less responsive to unloading in the muscle of rats with training experience. It was further determined that exercise training caused an early turnover of nucleosome components, which may alter the responsiveness of genes to stimulus in later life.

Prenatal myonuclei play a crucial role in skeletal muscle hypertrophy in rodents.

Multinucleated muscle fibers are formed by the fusion of myogenic progenitor cells during embryonic and fetal myogenesis. However, the role of prenatally incorporated myonuclei in the skeletal muscle fibers of adult animals is poorly understood. We demonstrated, using muscle-specific reporter mice, that the prenatal myonuclei remained in the adult soleus muscle, although cardiotoxin injection caused the loss of prenatal myonuclei. Overloading by the tendon transection of synergists failed to induce compensatory hypertrophy in regenerated soleus muscle fibers of adult rats, whereas unloading by tail suspension normally induced the fiber atrophy. Loss of hypertrophying function correlated with the lowered histone acetylation at the transcription start site of Igf1r gene, which was one of the genes that did not respond to the overloading. These parameters were improved by the transplantation of cells harvested from the juvenile soleus muscles of neonatal rats in association with enhanced histone acetylation of Igf1r gene. These results indicated that the presence of prenatal myonuclei was closely related to the status of histone acetylation, which could regulate the responsiveness of muscle fibers to physiological stimuli.