Muscle regeneration controlled by a designated DNA dioxygenase (RNA-Seq)

DNA methylation plays critical roles in regulating muscle cell fate determination and myogenesis. Tet dioxygenases are responsible for active DNA demethylation. The functions of Tet proteins in muscle regeneration have not been well characterized. Here we find that Tet2 is required for complete regeneration after muscle injury. Loss of Tet2 in myoblasts leads to reduced fusion index and thinner myofibers. Tet2 activates transcription of key differentiation modulator Myogenin (MyoG) further promoting myoblast differentiation and fusion. Re-expressing of MyoG in Tet2 KO myoblasts rescues the differentiation and fusion defects. Further mechanistic analysis reveals that Tet2 facilitates the recruitment of H3K4me1 and H3K27ac, increases the chromatin accessibility, and MyoD binding on MyoG enhancer. These functions are specifically executed by Tet2, but not Tet1 and Tet3. We identified the Tet2 specific function during myogenesis and shed new lights on DNA methylation and pioneer transcription factor transcription activation. Overall design: Examine RNA expression, DNA methylation and chromatin accessibility differences between Tet2 KO and WT mouse primary myoblasts.

DNA甲基化（DNA methylation）在调控肌细胞命运决定与肌发生过程中发挥关键作用。Tet双加氧酶（Tet dioxygenases）负责介导主动DNA去甲基化过程。目前对于Tet蛋白在肌肉再生中的功能尚未得到充分阐释。本研究发现，Tet2对于肌肉损伤后的完全再生是必需的。成肌细胞中Tet2的缺失会降低细胞融合指数，并导致肌纤维变细。Tet2可激活关键分化调节因子肌分化蛋白（Myogenin，简称MyoG）的转录，进而促进成肌细胞的分化与融合。在Tet2敲除（Tet2 KO）的成肌细胞中重新表达MyoG，可挽救其分化与融合缺陷。进一步的机制分析显示，Tet2可促进H3K4me1与H3K27ac的招募，提升染色质开放性，并增强MyoD在MyoG增强子区域的结合能力。上述功能仅由Tet2特异性执行，而非Tet1与Tet3。本研究明确了Tet2在肌发生过程中的特异性功能，为DNA甲基化与先锋转录因子的转录激活机制提供了新的见解。实验设计：检测Tet2敲除（Tet2 KO）与野生型（Wild Type, WT）小鼠原代成肌细胞之间的RNA表达、DNA甲基化及染色质开放性差异。