Somatic Reprogramming Research Articles

Progress on abnormal development of cloned pigs generated by somatic cell transfer nuclear

Cloning, also known as somatic cell nuclear transfer (SCNT), is an asexual reproduction technique that reprograms differentiated cells to the totipotent state, and generates offspring with a genotype identical to the donor cells. Pig cloning technique holds great promise for propagating excellent breeding boars, generating genetically modified pigs, protecting rare and endangered pigs and studying the mechanisms of somatic cell nucleus reprogramming. However, cloned pigs suffer from various developmental defects, including low birth rate, low birth weight, and high stillbirth occurrence, neonatal mortality and congenital malformations, which severely hamper their applications. Errors in epigenetic reprogramming of donor nucleus are considered as the main causes of low cloning efficiency and abnormal embryonic development in cloned embryos and animals. However, most studies to correct the errors in epigenetic reprogramming of cloned pig embryos have not substantially improved the birth and survival rates of cloned pigs. In this review, we summarize the abnormal phenotypes, causes of abnormal development of cloned pigs and effective methods for improving pig cloning efficiency, thereby providing a reference for the future research to improve the development and survival rates of cloned pig embryos and cloned pigs.

Somatic Reprograming by Nuclear Transfer.

Somatic cell nuclear transfer (SCNT) is a powerful technique, although challenging, to study reprograming into the totipotent state of differentiated nuclei in mammals. This procedure was initially applied in farm animals, then rodents, and more recently in primates. Nuclear transfer of embryonic stem cells is known to be more efficient, but many types of somatic cells have now been successfully reprogramed with this procedure. Moreover, SCNT reprograming is more effective on a per cell basis than induced Pluripotent Stem Cells (iPSC) and provides interesting clues regarding the underlying processes. In this chapter, we describe the protocol of nuclear transfer in mouse that combines cell cycle synchronization of the donor cells, enucleation of metaphase II oocyte and Piezo-driven injection of a donor cell nucleus followed by activation of the reconstructed embryos and nonsurgical transfer into pseudo-pregnant mice. Moreover, this protocol includes two facultative steps to erase the epigenetic "memory" of the donor cells and improve chromatin remodeling by histones modifications targeting.

Tumorigenic and Immunogenic Properties of Induced Pluripotent Stem Cells: a Promising Cancer Vaccine.

Induced pluripotent stem cells (iPSCs) are mainly characterized by their unlimited proliferation abilities and potential to develop into almost any cell type. The creation of this technology has been of great interest to many scientific fields, especially regenerative biology. However, concerns about the safety of iPSC application in transplantation have arisen due to the tumorigenic and immunogenic properties of iPSCs. This review will briefly introduce the developing history of somatic reprogramming and applications of iPSC technology in regenerative medicine. In addition, the review will highlight two challenges to the efficient usage of iPSCs and the underlying mechanisms of these challenges. Finally, the review will discuss the expanding application of iPSC technology in cancer immunotherapy as a potential cancer vaccine and its advantages in auxiliary treatment compared with oncofetal antigen-based and embryonic stem cell (ESC)-based vaccines.

Therapeutic effects of interleukin-37 and induced cardiosphere on treating myocardial ischemia-reperfusion injury

BackgroundsMyocardial ischemia-reperfusion injury (MI-RI) has many adverse complications with high mortality rate. In the current study, we investigated the therapeutic advantages of delivering Interleukin-37 (IL-37) by induced cardiospheres (iCS), generated from adult skin fibroblasts via somatic reprogramming, in treating the mice model MI-RI. MethodsThe mouse model of MI-RI was established and the iCS cells with IL-37 overexpression (iCS-IL37) were transplanted into the mice via tail-vein injection. Left ventricular (LV) dimensions and LV pressure-volume measurements were assessed by parasternal long-axis echocardiography and hemodynamic assessment. The infarct size was determined by histology analysis. And the inflammatory responses were analyzed by using enzyme-linked immunosorbent assay (ELISA). ResultsThe LV function was significantly improved after the iCS-IL37 transplantation when compared to the vehicle control group and iCS group, including the end-systolic pressure and dP/dtMax. Furthermore, the infarct size was significantly decreased after the iCS-IL37 transplantation. The protein levels of pro-inflammatory cytokines, including tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β), were down-regulated by the iCS-IL37 transplantation. ConclusionThe present study indicated that the iCS with IL-37 overexpression had therapeutic effects on the mice model of MI-RI.

YTHDF2/3 Are Required for Somatic Reprogramming through Different RNA Deadenylation Pathways

N6-methyladenosine (m6A), the most abundant reversible modification on eukaryote messenger RNA, is recognized by a series of readers, including the YT521-B homology domain family (YTHDF) proteins, which are coupled to perform physiological functions. Here, we report that YTHDF2 and YTHDF3, but not YTHDF1, are required for reprogramming of somatic cells into induced pluripotent stem cells (iPSCs). Mechanistically, we found that YTHDF3 recruits the PAN2-PAN3 deadenylase complex and conduces to reprogramming by promoting mRNA clearance of somatic genes, including Tead2 and Tgfb1, which parallels the activity of the YTHDF2-CCR4-NOT deadenylase complex. Ythdf2/3 deficiency represses mesenchymal-to-epithelial transition (MET) and chromatin silencing at loci containing the TEAD motif, contributing to decreased reprogramming efficiency. Moreover, RNA interference of Tgfb1 or the Hippo signaling effectors Yap1, Taz, and Tead2 rescues Ythdf2/3-defective reprogramming. Overall, YTHDF2/3 couples RNA deadenylation and regulation with the clearance of somatic genes and provides insights into iPSC reprogramming at the posttranscriptional level.

Abstract 488: Establishing consistent methods for the generation of donor-derived iPSCs from T cell subsets

Abstract T cell-based immunotherapies offer a potent and specific solution to grievous diseases such as cancer and viral infections. Expansion and re-infusion of antigen specific T cells has seen success in the treatment of metastatic melanoma and prevention of viral infections such as CMV and EBV. Additionally, engineered T cell therapies, especially chimeric antigen receptor (CAR) T cells, have been met with great clinical success and spurred innovation within the field of cellular immunotherapies. While promising, these therapies rely on sourcing T cells from donor derived blood. This process is invasive to the patient, especially when multiple or prolonged blood draws are required to obtain sufficient T cell numbers. Additionally, T cells sourced from diseased patients are often exhausted and display significantly perturbed growth and dysfunctional responses to antigen stimulation. An appealing alternative is somatic reprogramming of T cells to generate induced pluripotent stem cells (iPSCs) and differentiated into rejuvenated T cells that retain the original TCR, particularly with tumor infiltrating lymphocytes or viral specific T cells. We had previously reported a Sendai-virus based CytoTune 2.1 Reprogramming specifically designed for translational and clinical research. In this study we examine the effect of various cellular phenotypes and optimize methods for consistent reprogramming of multiple donor T cell reprogramming. Additionally, pan CD3+ cells isolated using different methods and further enriched for CD4+ and CD8+ were successfully reprogrammed. The resulting iPSCs were extensively characterized to ensure that pluripotency was achieved and that TCR sequences were preserved. This approach of reprogramming T cells and specific cell subtypes to derive iPSC with a known TCR sequence enables their use for downstream application as a renewable source of T cells with known antigen specificity. Citation Format: Mark Landon, Chad C. MacArthur, Melissa E. Hernandez, Uma Lakshmipathy, Jerry Guzman. Establishing consistent methods for the generation of donor-derived iPSCs from T cell subsets [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 488.

Transient Dux expression facilitates nuclear transfer and induced pluripotent stem cell reprogramming.

Cloned animals generated by somatic cell nuclear transfer (SCNT) have been reported for many years; however, SCNT is extremely inefficient, and zygotic genome activation (ZGA) is required for SCNT-mediated somatic cell reprogramming. To identify candidate factors that facilitate ZGA in SCNT-mediated reprogramming, we performed siRNA-repressor and mRNA-inducer screenings, which reveal Dux, Dppa2, and Dppa4 as key factors enhancing ZGA in SCNT. We show that direct injection of ZGA inducers has no significant effect on SCNT blastocyst formation; however, following the establishment of an inducible Dux transgenic mouse model, we demonstrate that transient overexpression of Dux not only improves SCNT efficiency but also increases that of chemically induced pluripotent stem cell reprogramming. Moreover, transcriptome profiling reveals that Dux-treated SCNT embryos are similar to fertilized embryos. Furthermore, transient overexpression of Dux combined with inactivation of DNA methyltransferases (Dnmts) further promotes the full embryonic development of SCNT-derived animals. These findings enhance our understanding of ZGA-regulator function in somatic reprogramming.

Replicates in stem cell models—How complicated!

Current complexities in human pluripotent stem cell (hPSC)-based studies. hPSC studies begin with the recruitment of patients harboring disease-associated gene variant(s) that may increase susceptibility to disease development. Somatic reprogramming is then performed to derive patient-specific human induced pluripotent stem cells (hiPSCs), followed by step-wise directed differentiation to a relevant cell type before qualitative/quantitative assays are performed to assess for phenotypic or gene expression differences between the healthy and diseased hiPSCs.

The function of a heterozygous p53 mutation in a Li-Fraumeni syndrome patient.

p53 is one of the most extensively studied proteins in cancer research. Mutations in p53 generally abolish normal p53 function, and some mutants can gain new oncogenic functions. However, the mechanisms underlying p53 mutation-driven cancer remains to be elucidated. Our study investigated the function of a heterozygous p53 mutation (p.Asn268Glufs*4) in a Li-Fraumeni syndrome (LFS) patient. We used episomal technology to perform somatic reprogramming, and used molecular and cell biology methods to determine the p53 mutation levels in patient-originated induced pluripotent stem (iPS) cells at the RNA and protein levels. We found that p53 protein expression was not increased in this patient's somatic cells compared with those of a healthy control. p53 mutation facilitates the proliferation of tumor cells by inhibiting apoptosis and promoting cell division. It can inhibit the efficiency of somatic reprogramming by inhibiting OCT4 expression during reprogramming stage. Moreover, not all p53 mutant iPS cell lines have mutant p53 RNA sequences. A small percentage of mutant p53 mRNA is present in the somatic cells from the patient and his mother. In summary, this p53 mutation can promote tumor cell proliferation, inhibit somatic reprogramming, and exhibit random p53 allelic expression of heterozygous mutations in the patient and iPS cells which may be one of the reasons why the people with p53 mutations develop cancer at random. This finding suggested that mutant p53 allelic expression should be added to the risk forecasting of cancer.

Abstract A17: Polyploid giant cancer cells lead to evolution of drug resistance in high-grade ovarian cancer

Abstract Whole genomic duplications (WGD) have been reported in 37% to 56% of high-grade cancers. WGD leads to formation of polyploid giant cancer cells (PGCCs) with salient and atypical nuclear morphology noted by light microscopy. However, PGCCs are generally considered nonviable due to their inability to execute mitosis. We have recently reported that PGCCs are capable of generating mitosis-competent progeny cells via primitive amitotic division including budding, splitting, and burst. We have further demonstrated that PGCCs are part of the process of somatic reprogramming named the giant cell life cycle composed of four distinct but overlapping phases including initiation, self-renewal, termination, and stability. The giant cell life cycle resets the somatic cell from the mitotic cycle to stress-induced endoreplication cell cycle via WGD to form PGCCs. The PGCCs activate endogenous embryonic factors for the reprogramming, which leads to the birth of drug-resistant cells by clonal selection from newly generated heterogeneous cancer stem cells. However, the underlying genetic and epigenetic mechanisms during WGD remain unclear. By mapping DNA methylation and whole-genome sequencing, monitoring the telomere dynamics, and tracking the cellular and nuclear morphology using microscopic time-lapse imaging observations during the time course of giant cell life cycle, we hereby provide new evidence that the daughters cells derived from PGCCs have massive alterations at the whole-genome and epigenome levels. The reprogrammed resistant daughter cells are achieved, at least in part, by genomic instability via shortened telomeres. Targeting PGCCs and promoting their differentiation during the course of giant cell life cycle may represent a novel approach to intercept drug resistance. Citation Format: Na Niu, Xiaoran Li, Jun Yao, Anil Sood, Jinsong Liu. Polyploid giant cancer cells lead to evolution of drug resistance in high-grade ovarian cancer [abstract]. In: Proceedings of the AACR Special Conference on the Evolving Landscape of Cancer Modeling; 2020 Mar 2-5; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2020;80(11 Suppl):Abstract nr A17.

Characterisation of extraembryonic endoderm-like cells from mouse embryonic fibroblasts induced using chemicals alone

BackgroundThe development of somatic reprogramming, especially purely chemical reprogramming, has significantly advanced biological research. And chemical-induced extraembryonic endoderm-like (ciXEN) cells have been confirmed to be an indispensable intermediate stage of chemical reprogramming. They resemble extraembryonic endoderm (XEN) cells in terms of transcriptome, reprogramming potential, and developmental ability in vivo. However, the other characteristics of ciXEN cells and the effects of chemicals and bFGF on the in vitro culture of ciXEN cells have not been systematically reported.MethodsChemicals and bFGF in combination with Matrigel were used to induce the generation of ciXEN cells derived from mouse embryonic fibroblasts (MEFs). RNA sequencing was utilised to examine the transcriptome of ciXEN cells, and PCR/qPCR assays were performed to evaluate the mRNA levels of the genes involved in this study. Hepatic functions were investigated by periodic acid-Schiff staining and indocyanine green assay. Lactate production, ATP detection, and extracellular metabolic flux analysis were used to analyse the energy metabolism of ciXEN cells.ResultsciXEN cells expressed XEN-related genes, exhibited high proliferative capacity, had the ability to differentiate into visceral endoderm in vitro, and possessed the plasticity allowing for their differentiation into induced hepatocytes (iHeps). Additionally, the upregulated biological processes of ciXEN cells compared to those in MEFs focused on metabolism, but their energy production was independent of glycolysis. Furthermore, without the cocktail of chemicals and bFGF, which are indispensable for the generation of ciXEN cells, induced XEN (iXEN) cells remained the expression of XEN markers, the high proliferative capacity, and the plasticity to differentiate into iHeps in vitro.ConclusionsciXEN cells had high plasticity, and energy metabolism was reconstructed during chemical reprogramming, but it did not change from aerobic oxidation to glycolysis. And the cocktail of chemicals and bFGF were non-essential for the in vitro culture of ciXEN cells.

Cellular Reprogramming Approaches to Engineer Cardiac Pacemakers.

The goal of this paper is to review present knowledge regarding biological pacemakers created by somatic reprogramming as a platform for mechanistic and metabolic understanding of the rare subpopulation of pacemaker cells, with the ultimate goal of creating biological alternatives to electronic pacing devices. Somatic reprogramming of cardiomyocytes by reexpression of embryonic transcription factor T-box 18 (TBX18) converts them into pacemaker-like. Recent studies take advantage of this model to gain insight into the electromechanical, metabolic, and architectural intricacies of the cardiac pacemaker cell across various models, including a surgical model of complete atrioventricular block (CAVB) in adult rats. The studies reviewed here reinforce the potential utility of TBX18-induced pacemaker myocytes (iPMS) as a minimally invasive treatment for heart block. Several challenges which must be overcome to develop a viable therapeutic intervention based on these observations are discussed.

SSRP1-mediated histone H1 eviction promotes replication origin assembly and accelerated development

In several metazoans, the number of active replication origins in embryonic nuclei is higher than in somatic ones, ensuring rapid genome duplication during synchronous embryonic cell divisions. High replication origin density can be restored by somatic nuclear reprogramming. However, mechanisms underlying high replication origin density formation coupled to rapid cell cycles are poorly understood. Here, using Xenopus laevis, we show that SSRP1 stimulates replication origin assembly on somatic chromatin by promoting eviction of histone H1 through its N-terminal domain. Histone H1 removal derepresses ORC and MCM chromatin binding, allowing efficient replication origin assembly. SSRP1 protein decays at mid-blastula transition (MBT) when asynchronous somatic cell cycles start. Increasing levels of SSRP1 delay MBT and, surprisingly, accelerate post-MBT cell cycle speed and embryo development. These findings identify a major epigenetic mechanism regulating DNA replication and directly linking replication origin assembly, cell cycle duration and embryo development in vertebrates.

A Link between Genetic Disorders and Cellular Impairment, Using Human Induced Pluripotent Stem Cells to Reveal the Functional Consequences of Copy Number Variations in the Central Nervous System-A Close Look at Chromosome 15.

Recent cutting-edge human genetics technology has allowed us to identify copy number variations (CNVs) and has provided new insights for understanding causative mechanisms of human diseases. A growing number of studies show that CNVs could be associated with physiological mechanisms linked to evolutionary trigger, as well as to the pathogenesis of various diseases, including cancer, autoimmune disease and mental disorders such as autism spectrum disorders, schizophrenia, intellectual disabilities or attention-deficit/hyperactivity disorder. Their incomplete penetrance and variable expressivity make diagnosis difficult and hinder comprehension of the mechanistic bases of these disorders. Additional elements such as co-presence of other CNVs, genomic background and environmental factors are involved in determining the final phenotype associated with a CNV. Genetically engineered animal models are helpful tools for understanding the behavioral consequences of CNVs. However, the genetic background and the biology of these animal model systems have sometimes led to confusing results. New cellular models obtained through somatic cellular reprogramming technology that produce induced pluripotent stem cells (iPSCs) from human subjects are being used to explore the mechanisms involved in the pathogenic consequences of CNVs. Considering the vast quantity of CNVs found in the human genome, we intend to focus on reviewing the current literature on the use of iPSCs carrying CNVs on chromosome 15, highlighting advantages and limits of this system with respect to mouse model systems.

RNA sequencing revealed the abnormal transcriptional profile in cloned bovine embryos

Somatic cell nuclear transfer (SCNT) has potential applications in agriculture and biomedicine, but the efficiency of cloning is still low. In this study, the transcriptional profiles in cloned and fertilized embryos were measured and compared by RNA sequencing. The 2-cell embryos were detected to identify the earliest transcriptional differences between embryos derived through IVF and SCNT. As a result, 364 genes showed decreased expression in cloned 2-cell embryos and were enriched in “intracellular protein transport” and “ubiquitin mediated proteolysis”. In blastocysts, 593 genes showed decreased expression in cloned blastocysts and were enriched in “RNA binding”, “nucleotide binding”, “embryo development”, and “adherens junction”. We identified 14 development related genes that were not activated in the cloned embryos. Then, 68 and 245 long non-coding RNAs were recognized abnormally expressed in cloned 2-cell embryos and cloned blastocysts, respectively. Furthermore, we found that incomplete RNA-editing occurred in cloned embryos and might be caused by decreased ADAR expression. In conclusion, our study revealed the abnormal transcripts and deficient RNA-editing sites in cloned embryos and provided new data for further mechanistic studies of somatic nuclear reprogramming.

Protective effects of induced cardiosphere on myocardial ischemia-reperfusion injury through secreting interleukin 10

BackgroundsMyocardial ischemia-reperfusion injury (MI-RI) has many adverse complications with high mortality rate. It has been demonstrated that the induced cardiospheres (iCS), generated from adult skin fibroblasts via somatic reprogramming, represents a novel source for cell therapy in myocardial infarction. However, whether the iCS could also be applied to treat MI-RI remains unclear. Thus, we investigated the therapeutic application of iCS in the mice model MI-RI. MethodsThe mice model of MI-RI was established and the iCS cells were transplanted to the mice via tail-vein injection. Left ventricular (LV) dimensions and LV pressure-volume measurements were assessed by parasternal long-axis echocardiography. The infarct size was determined by histology analysis. And the inflammatory responses were analyzed by using enzyme-linked immunosorbent assay (ELISA). ResultsThe LV function was significantly improved after the iCS transplantation when compared to the vehicle control group, including the end-systolic pressure and dP/dtMax. Furthermore, the infarct size was significantly decreased after the iCS transplantation. The protein levels of pro-inflammatory cytokines, including tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β), were down-regulated by the iCS transplantation while the IL-10 was up-regulated. The anti-inflammatory factor IL-10 was found to be expressed and secreted by the iCS cells and knocking down the IL-10 in iCS would significantly impair the therapeutic effects of iCS in the mice model of MI-RI. ConclusionThe present study indicated that the iCS had therapeutic effects on the mice model of MI-RI through secreting the IL-10.

Autophagy mediates temporary reprogramming and dedifferentiation in plant somatic cells.

Somatic cells acclimate to changes in the environment by temporary reprogramming. Much has been learned about transcription factors that induce these cell-state switches in both plants and animals, but how cells rapidly modulate their proteome remains elusive. Here, we show rapid induction of autophagy during temporary reprogramming in plants triggered by phytohormones, immune, and danger signals. Quantitative proteomics following sequential reprogramming revealed that autophagy is required for timely decay of previous cellular states and for tweaking the proteome to acclimate to the new conditions. Signatures of previous cellular programs thus persist in autophagy-deficient cells, affecting cellular decision-making. Concordantly, autophagy-deficient cells fail to acclimatize to dynamic climate changes. Similarly, they have defects in dedifferentiating into pluripotent stem cells, and redifferentiation during organogenesis. These observations indicate that autophagy mediates cell-state switches that underlie somatic cell reprogramming in plants and possibly other organisms, and thereby promotes phenotypic plasticity.

Dysregulated MicroRNA Fingerprints and Methylation Patterns in Hepatocellular Carcinoma, Cancer Stem Cells, and Mesenchymal Stem Cells.

Hepatocellular carcinoma (HCC) is one of the top causes of cancer mortality worldwide. Although HCC has been researched extensively, there is still a need for novel and effective therapeutic interventions. There is substantial evidence that initiation of carcinogenesis in liver cirrhosis, a leading cause of HCC, is mediated by cancer stem cells (CSCs). CSCs were also shown to be responsible for relapse and chemoresistance in several cancers, including HCC. MicroRNAs (miRNAs) constitute important epigenetic markers that regulate carcinogenesis by acting post-transcriptionally on mRNAs, contributing to the progression of HCC. We have previously shown that co-culture of cancer cells with mesenchymal stem cells (MSCs) could induce the reprogramming of MSCs into CSC-like cells. In this review, we evaluate the available data concerning the epigenetic regulation of miRNAs through methylation and the possible role of this regulation in stem cell and somatic reprogramming in HCC.

Optimizing treatment of DNA methyltransferase inhibitor RG108 on porcine fibroblasts for somatic cell nuclear transfer.

Aberration in DNA methylation is believed to be one of the major causes of abnormal gene expression and inefficiency of somatic cell nuclear transfer (SCNT). RG108, a non-nucleoside DNA methyltransferase (DNMT) inhibitor, has been reported to facilitate somatic nuclear reprogramming and improved blastocyst formation. The aim of this study was to investigate interaction effect of RG108 treatment time (24-72hr) and concentrations (0.05-50µM) on donor cells, and further to optimize the treatment for porcine SCNT. Our results showed that RG108 treatment resulted in time-dependent decrease of genome-wide DNA methylation on foetal fibroblasts, which only happened after 72-hr treatment in our experiments, and no interaction effect between treatment time and concentration. Remarkable decrease of methylation in imprinted gene H19 and increased apoptosis was observed in 5 and 50µM RG108-treated cells. Furthermore, the blastocyst rates of SCNT embryos were increased as the fibroblasts treated with RG108 at 5 and 50µM, and additional treatment during cultivation of SCNT embryos would not provide any advantage for blastocyst formation. In conclusion, the RG108 treatment of 72hr and 5μM would be optimized time and concentration for porcine foetal fibroblasts to improve the SCNT embryonic development. In addition, combined treatment of RG108 on donor cells and SCNT embryos would not be beneficial for embryonic development.

Transcription Factor-Directed Re-wiring of Chromatin Architecture for Somatic Cell Nuclear Reprogramming toward trans-Differentiation.

MYOD-directed fibroblast trans-differentiation into skeletal muscle provides a unique model to investigate how one transcription factor (TF) reconfigures the three-dimensional chromatin architecture to control gene expression, which is otherwise achieved by the combinatorial activities of multiple TFs. Integrative analysis of genome-wide high-resolution chromatin interactions, MYOD and CTCF DNA-binding profile, and gene expression, revealed that MYOD directs extensive re-wiring of interactions involving cis-regulatory and structural genomic elements, including promoters, enhancers, and insulated neighborhoods (INs). Re-configured INs were hot-spots of differential interactions, whereby MYOD binding to highly constrained sequences at IN boundaries and/or inside INs led to alterations of promoter-enhancer interactions to repress cell-of-origin genes and to activate muscle-specific genes. Functional evidence shows that MYOD-directed re-configuration of chromatin interactions temporally preceded the effect on gene expression and was mediated by direct MYOD-DNA binding. These data illustrate a model whereby a single TF alters multi-loop hubs to drive somatic cell trans-differentiation.