Browsing by Subject "MyoD Protein"
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Item Intrinsic Specificity of Binding and Regulatory Function of Class II bHLH Transcription Factors(2016-11-28) Casey, Bradford Harris; Krämer, Helmut; Johnson, Jane E.; Konopka, Genevieve; MacDonald, Raymond J.Embryonic development begins with a single cell, and gives rise to the many diverse cells which comprise the complex structures of the adult animal. Distinct cell fates require precise regulation to develop and maintain their functional characteristics. Transcription factors provide a mechanism to select tissue-specific programs of gene expression from the shared genome. ASCL1, ASCL2, and MYOD are class II basic Helix-Loop-Helix (bHLH) transcription factors which play crucial roles in lineage specification in the developing embryo. In vivo, these factors bind to distinct genomic sites, and regulate distinct transcriptional programs. The mechanisms by which they select their cognate binding sites remain poorly defined. Here, we utilize an inducible system to express these master regulatory factors in embryonic stem cells to characterize early events in bHLH factor binding and function in a common cellular context, removed from their role as endogenous master regulators of lineage specification. Using genome-wide sequencing approaches, we demonstrate that these factors maintain distinct binding when ectopically expressed in a common context. We observe that they initiate distinct transcriptional programs, which include key regulators in lineage specification. By comparing chromatin accessibility of bHLH binding sites, we reveal a shared ability for these factors to bind nucleosome-occupied sites, and meet the criteria which define pioneer transcription factors. We further characterize epigenetic features of the empirically observed genome-wide binding sites of these factors, and compare these findings to the conventional understanding of bHLH factor function. This work represents the first comprehensive approach to direct comparison of early events in the binding and transcriptional profiles of ASCL1, ASCL2, and MYOD.Item The Role of Twist2 in Physiologic and Pathologic Myogenesis(2019-05-01) Li, Stephen; Amatruda, James F.; Morrison, Sean J.; Hobbs, Helen H.; Olson, Eric N.Skeletal muscle is a highly regenerative tissue required for vertebrate life. It composes a significant portion of body mass and enables the physiologic processes of movement and breathing. Given its importance, skeletal muscle is also highly susceptible to aging and diseases such as cancer. Aging-related muscle atrophy (sarcopenia) is a process that affects nearly every person, contributing to debilitations and reductions in quality of life. As people age, fast-twitch muscle fibers selectively atrophy resulting in weakness. Normally, muscle regenerates through a population of stem cells called satellite cells, which differentiate and non-selectively fuse to existing myofibers in order to repair the damaged muscle tissue. However, several recent studies have suggested that the contribution of satellite cells to muscle during homeostasis and aging is minimal. Thus, it's possible that loss of an alternative muscle precursor that fuses specifically with fast-twitch fibers may be a mechanism by which aging-related muscle atrophy occurs. Through the technique of lineage-tracing (fate-mapping) of the transcription factor Twist2, we have identified a novel muscle progenitor that fuses specifically to type IIb/x (fast-twitch, glycolytic) muscle fibers during both aging and homeostasis. Additionally, loss of Twist2+ cells result in specific atrophy of fast-twitch myofibers. I show that Twist2 plays a role in regulating fiber-type specificity through upregulation of the membrane receptor Nrp1. Additionally, the Nrp1 chemo-repulsive ligand, Sema3a, is expressed by both type I and IIa fibers. This Sema3a-Nrp1 signaling mechanisms prevents Twist2+ cells from fusing to type I and IIa fibers, and exogenous overexpression of Sema3a in type IIb fibers impairs the contribution of Twist2+ cells to these fibers. I also found that Twist2 is highly amplified in rhabdomyosarcoma (RMS), a pediatric soft tissue sarcoma expressing hallmarks of the skeletal muscle lineage. Twist2 overexpression was capable of reversibly repressing myogenesis and promoting dedifferentiation of myotubes. Through integrated genomic analyses, I show that Twist2 epigenetically remodels the chromatin landscape to redirect MyoD binding from myogenic loci to oncogenic loci, enabling MyoD to adopt novel functions. Our findings identify the previously unknown roles of Twist2 in regulating mammalian muscle biology.