Induced muscle progenitor cells [ATAC-seq]

The generation of myotubes from fibroblasts upon forced MyoD expression is a classic example of factor-induced reprogramming in mammals. We recently discovered that additional modulation of signaling pathways with small molecules facilitates reprogramming to more primitive induced muscle progenitor cells (iMPCs). However, the mechanisms by which a single transcription factor drives differentiated cells into distinct developmental states remain unknown. We therefore dissected the transcriptional and epigenetic dynamics of fibroblasts undergoing MyoD-dependent reprogramming to either myotubes or iMPCs using a novel MyoD transgenic model. To this end, we performed ATAC sequencing for Pax7-nGFP positive iMPCs/satellite cells, cells undergoing dedifferentiation (i.e. Dox+FRG) or transdifferentiation (i.e. Dox) and cells overexpressing a wild type MyoD or a mutant MyoD (i.e. MN) in the presence of FRG. Our analyses elucidate the role of MyoD in myogenic reprogramming and derive general principles by which transcription factors and signaling pathways cooperate to rewire cell identity. Our results may also inform on potential therapeutic applications of direct reprogramming. Overall design: ATAC-seq was performed for 14 samples in duplicate.

通过异位表达肌分化因子（MyoD）将成纤维细胞（fibroblasts）转化为肌管（myotubes），是哺乳动物中因子诱导重编程的经典研究范例。我们近期发现，借助小分子（small molecules）对信号通路（signaling pathways）进行额外调控，可将重编程产物提升至更原始的诱导性肌肉祖细胞（induced muscle progenitor cells，简称iMPCs）。然而，单个转录因子（transcription factor）如何将分化细胞导向不同发育状态的具体调控机制，目前仍未明确。基于此，我们借助新型MyoD转基因模型（transgenic model），解析了依赖MyoD的重编程过程中，成纤维细胞向肌管或iMPCs转化时的转录组与表观遗传（epigenetic）动态变化。为此，我们对以下样本开展了ATAC测序（ATAC sequencing）：Pax7-nGFP（Pax7-nGFP）阳性的iMPCs/卫星细胞（satellite cells）、经历去分化（dedifferentiation，即Dox+FRG组）或转分化（transdifferentiation，即Dox组）的细胞，以及在FRG处理条件下过表达野生型（wild type）MyoD或突变型（mutant）MyoD（即MN组）的细胞。本研究阐明了MyoD在肌源性重编程中的功能，并推导得出转录因子与信号通路协同重塑细胞身份（cell identity）的通用调控原则。本研究结果还可为直接重编程（direct reprogramming）的潜在治疗应用提供理论参考。实验整体设计：对14份样本分别进行两次生物学重复的ATAC-seq。