Satellite cells (SC) are muscle stem cells positioned between the basal lamina and the sarcolemma of muscle fibers, and they serve as a pool to regenerate muscle tissue that has been damaged. Whereas embryonic stem (ES) cells have been extensively characterized at the transcriptional and epigenetic levels, our understanding of satellite cells is rudimentary. Although SC are essential for muscle regeneration, they have proven refractory to characterization owing to a number of technical impediments, including their paucity in muscle tissue, the inability of ES cells to differentiate into skeletal muscle, and the inability of SC to maintain ?stemness? in culture. Recently, we have begun characterizing a novel satellite cell model?iPax7 cells--in which the master regulator of satellite cell identity, Pax7, is inducibly expressed in ES cells. iPax7 cells have been shown to repopulate the satellite cell niche in animals, but they have not been characterized at the molecular or genomic level. Our goals are as follows. First, we will more deeply characterize the novel iPax7 model for satellite cells. Second, we will for the first time attempt to obtain a satellite cell-specific gene expression signature. We will identify markers of SC, which will be of further value for understanding and characterizing satellite cells and other myogenic precursors. Third, we will unravel how Pax7, a master regulator required for specification and maintenance of SC identity, assembles key regulatory elements. This will enable us to devise a gene regulatory network or ?blueprint? for satellite cells, allowing us to understand how Pax7 controls expression of other transcription factors that specify stem cell identity. We will leverage our recently acquired epigenetic data and Pax7 ChIP-seq data, as well as our rich data set mapping the genomic landscape of myoblasts and myotubes, to pursue two Specific Aims.
In Aim 1, we will compare iPax7 cells with muscle-derived satellite cells and ES cells using genome-wide epigenetic and transcriptome-level comparisons. These studies will allow us to identify satellite cell-specific signatures. We will test the hypothesis that a set of transcription factors collaborates with Pax7 to dictate satellite cell identity.
In Aim 2, we will examine the role of Pax7 in modulating the epigenomic landscape of myogenic precursors and SC by dissecting its role at regulatory elements. Our studies will lay important groundwork for a comprehensive understanding of the transcriptional networks and epigenomic changes that accompany the restriction from pluripotency to skeletal muscle identity. Additionally, these studies may reveal essential mechanistic roles for Pax7 in satellite cell maintenance, a poorly understood area. Understanding and utilizing these myogenic precursors, which can be manipulated at will, may prove invaluable in a clinical setting, opening novel avenues to improve their regenerative capacity and suggesting possible therapeutic opportunities for treatment of muscle wasting and disease.
We have begun to define the regulatory networks and epigenetic modifications that promote skeletal muscle differentiation. In this proposal, we will use a novel model for skeletal muscle satellite cells to begin exploring the mechanisms and regulatory networks that establish satellite cell identity and that subsequently drive myogenic differentiation through assembly and activation of enhancers. Our findings will significantly expand our understanding of the satellite cell epigenome, shed light on the role of master transcriptional regulators in satellite cell maintenance, and suggest novel ways to improve the ability of these cells to regenerate, thereby opening therapeutic windows to treat age-and disease-associated muscle wasting.
| Lilja, Karin C; Zhang, Nan; Magli, Alessandro et al. (2017) Pax7 remodels the chromatin landscape in skeletal muscle stem cells. PLoS One 12:e0176190 |
