One week, but not 12?hours, of cast immobilization alters promotor DNA methylation patterns in the nNOS gene in mouse skeletal muscle
One week, but not 12?hours, of cast immobilization alters promotor DNA methylation patterns in the nNOS gene in mouse skeletal muscle
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一周而非 12 小时的石膏固定会改变小鼠骨骼肌中 nNOS 基因的启动子 DNA 甲基化模式
DOI：
10.1113/jp277019
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发表时间：
2019
期刊：
影响因子：
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通讯作者：
Higaki Yasuki
中科院分区：
文献类型：
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作者：
Tomiga Yuki;Ito Ai;Sudo Mizuki;Ando Soichi;Eshima Hiroaki;Sakai Kazuya;Nakashima Shihoko;Uehara Yoshinari;Tanaka Hiroaki;Soejima Hidenobu;Higaki Yasuki
dna甲基化可能在调节骨骼肌基因表达以适应运动和不运动中发挥重要作用。骨骼肌神经元型一氧化氮合酶（nNOS）是骨骼肌质量的关键调控因子；然而，目前尚不清楚nNOS的表达是否受DNA甲基化的调节。我们发现，固定1周后，小鼠腿部萎缩的慢抽动比目鱼肌中nNOS DNA甲基化水平升高，nNOS基因表达下调。这些变化在非萎缩性快缩指长伸肌中未被检测到。固定12小时后，nNOS基因表达降低，而nNOS DNA甲基化水平不变，表明短期肌肉不活动对nNOS基因表达的下调与DNA甲基化模式无关。这些发现有助于更好地理解通过nNOS/NO通路的表观遗传机制维持骨骼肌质量和预防肌肉萎缩。摘要dna甲基化是控制骨骼肌在各种环境刺激下（如运动和不运动）基因表达的一种机制。神经元型一氧化氮合酶（nNOS）调控骨骼肌萎缩。然而，在萎缩肌肉中调控nNOS表达的机制尚不清楚。我们假设萎缩性肌肉中的nNOS表达受比目鱼肌（Sol；慢肌纤维为主）和指长伸肌（EDL；快肌纤维为主）中nNOS启动子的DNA甲基化调控。石膏固定1周后，Sol肌明显萎缩，EDL肌无明显萎缩。我们发现，1周的石膏固定增加了Sol中nNOS DNA甲基化水平，尽管在EDL中仅检测到轻微变化。与萎缩Sol中DNA甲基化水平升高一致，总nNOS和nNOSµ（即骨骼肌中nNOS的主要剪接变体）的基因表达水平下降。萎缩Sol中nNOS蛋白的丰度和细胞膜（尤其是IIa型纤维）的免疫反应性也有所下降。石膏固定1周后，EDL中未观察到这些变化。此外，尽管没有明显的萎缩，但固定12小时后，总nNOS和nNOSµin Sol的基因表达水平降低。然而，nNOS DNA甲基化与基因表达之间没有关联。nNOS的另一个剪接变体nNOSβ基因在EDL中的表达在石膏固定下没有变化，而在Sol中未检测到其表达。我们得出结论，nNOS基因在石膏固定肌肉中表达的慢性适应可能与nNOS DNA甲基化有关。
Key pointsDNA methylation may play an important role in regulating gene expression in skeletal muscle to adapt to physical activity and inactivity.Neuronal nitric oxide synthase (nNOS) in skeletal muscle is a key regulator of skeletal muscle mass; however, it is unclear whether nNOS expression is regulated by DNA methylation.We found that 1 week of cast immobilization increased nNOS DNA methylation levels and downregulated nNOS gene expression in atrophic slow‐twitch soleus muscle from the mouse leg. These changes were not detected in non‐atrophic fast‐twitch extensor digitorum longus muscle.Twelve hours of cast immobilization decreased nNOS gene expression, whereas nNOS DNA methylation levels were unchanged, suggesting that downregulation of nNOS gene expression by short‐term muscle inactivity is independent of the DNA methylation pattern.These findings contribute to a better understanding of the maintenance of skeletal muscle mass and prevention of muscle atrophy by epigenetic mechanisms via the nNOS/NO pathway.AbstractDNA methylation is a mechanism that controls gene expression in skeletal muscle under various environmental stimuli, such as physical activity and inactivity. Neuronal nitric oxide synthase (nNOS) regulates muscle atrophy in skeletal muscle. However, the mechanisms regulating nNOS expression in atrophic muscle remain unclear. We hypothesized that nNOS expression in atrophic muscle is regulated by DNA methylation of the nNOS promotor in soleus (Sol; slow‐twitch fibre dominant) and extensor digitorum longus (EDL; fast‐twitch fibre dominant) muscles. One week of cast immobilization induced significant muscle atrophy in Sol but not in EDL. We showed that 1 week of cast immobilization increased nNOS DNA methylation levels in Sol, although only a minor change was detected in EDL. Consistent with the increased DNA methylation levels in atrophic Sol, the gene expression levels of total nNOS and nNOSµ (i.e. the major splicing variant of nNOS in skeletal muscle) decreased. The abundance of the nNOS protein and cell membrane (especially type IIa fibre) immunoreactivity also decreased in atrophic Sol. These changes were not observed in EDL after 1 week of cast immobilization. Furthermore, despite the lack of significant atrophy, 12 h of cast immobilization decreased gene expression levels of total nNOS and nNOSµ in Sol. However, no association was detected between nNOS DNA methylation and gene expression. The expression of the nNOSβ gene, another splicing variant of nNOS, in EDL was unchanged by cast immobilization, whereas its expression was not detected in Sol. We concluded that chronic adaptation of nNOS gene expression in cast immobilized muscle may involve nNOS DNA methylation.