Exploring networks underlying muscle stem cell identity - Resubmission - 1

项目摘要 肌肉干细胞，也被称为卫星细胞（SC），是一个祖细胞群，需要补充 因运动或因疾病或衰老引起的消耗而受损的肌肉。产生肌肉的能力 来自胚胎干细胞（ESC）的祖细胞将代表再生医学的重大进展。 然而，尽管最近取得了进展，但将小鼠和人ESC分化成具有生物学活性的细胞仍然非常具有挑战性。 肌肉祖细胞和SC代表肌肉组织内非常小的群体。这些限制 严重阻碍了对肌肉干细胞和关键调控过程的理解 控制SC身份。转录因子Pax 7在建立SC身份中起着关键作用，但由于 与获得SC相关的限制，我们对其分子功能的理解仍然不完整。 我们利用了一个系统，在这个系统中，ESC中的Pax 7表达能够产生肌源性前体， 为了重新填充SC生态位，我们已经开始研究染色质景观和表观遗传特征， 来了解它的独特属性通过这个系统，我们已经证明Pax 7可以 调节增强子功能和基因组拓扑结构。我们的目标是了解一个特定的染色质 环境使用最先进的基因组学、生物化学和蛋白质组学来指定卫星细胞身份 接近。在两个目标中，我们将：（I）研究Pax 7如何重新建模基因组，包装染色质， 长距离相互作用;（二）探索Pax 7如何与其他因素合作，以建立基因表达， 基因组重新布线在肌前体在体外和体内。我们将研究Pax 7相关的一个子集， 因子、增强子和鉴定的调控元件，并检查它们对基因组的功能影响。 结构、基因表达、细胞身份和分化。 众所周知，卫星细胞在衰老过程中更新效率较低，并且考虑到肌肉肿瘤 （横纹肌肉瘤）异常表达Pax 7融合蛋白以维持肌肉处于预分化状态， 我们的建议将有助于阐明异常Pax 7活性和干细胞 功能

Project Summary Muscle stem cells, also known as satellite cells (SC), are a progenitor population required to replenish muscle damaged by exercise or wasting caused by disease or aging. The ability to generate muscle progenitors from embryonic stem cells (ESC) would represent a major advance in regenerative medicine. However, despite recent progress, it remains very challenging to differentiate mouse and human ESC into muscle progenitors, and SC represent a very small population within muscle tissue. These limitations have significantly hampered progress toward understanding muscle stem cells, and the critical regulatory processes that govern SC identity. The transcription factor, Pax7, plays a critical role in establishing SC identity, but given the limitations associated with obtaining SC, our understanding of its molecular function remains incomplete. We have taken advantage of a system in which Pax7 expression in ESC generates myogenic precursors able to repopulate the SC niche, and we have begun investigating the chromatin landscape and epigenetic features of this population to understand its unique properties. With this system, we have shown that Pax7 can modulate enhancer function and genome topology. Our goal is to understand how a specific chromatin environment specifies satellite cell identity using state-of-the-art genomic, biochemical, and proteomic approaches. In two Aims, we will: (I) investigate how Pax7 re-models the genome, packaging chromatin for long-range interactions; (II) explore how Pax7 collaborates with other factors to establish gene expression and genome re-wiring in myogenic precursors in vitro and in vivo. We will inactivate a subset of Pax7-associated factors, enhancers, and identified regulatory elements and examine their functional impact on genome architecture, gene expression, cell identity, and differentiation. It is known that satellite cells are less efficiently renewed during aging, and given that muscle tumors (rhabdomyosarcomas) aberrantly express Pax7 fusion proteins to maintain muscle in a pre-differentiated state, our proposal will shed important light on pathological states resulting from aberrant Pax7 activity and stem cell function.