Molecular Roadblocks for Cellular Reprogramming

Institute for Stem Cell Biology and Regenerative Medicine, Department of Pathology, and Cancer Biology Program, Stanford University School of Medicine, 265 Campus Drive, Stanford, CA 94305, USA.
Molecular cell (Impact Factor: 14.02). 09/2012; 47(6):827-38. DOI: 10.1016/j.molcel.2012.09.008
Source: PubMed


During development, diverse cellular identities are established and maintained in the embryo. Although remarkably robust in vivo, cellular identities can be manipulated using experimental techniques. Lineage reprogramming is an emerging field at the intersection of developmental and stem cell biology in which a somatic cell is stably reprogrammed into a distinct cell type by forced expression of lineage-determining factors. Lineage reprogramming enables the direct conversion of readily available cells from patients (such as skin fibroblasts) into disease-relevant cell types (such as neurons and cardiomyocytes) or into induced pluripotent stem cells. Although remarkable progress has been made in developing novel reprogramming methods, the efficiency and fidelity of reprogramming need to be improved in order increase the experimental and translational utility of reprogrammed cells. Studying the mechanisms that prevent successful reprogramming should allow for improvements in reprogramming methods, which could have significant implications for regenerative medicine and the study of human disease. Furthermore, lineage reprogramming has the potential to become a powerful system for dissecting the mechanisms that underlie cell fate establishment and terminal differentiation processes. In this review, we will discuss how transcription factors interface with the genome and induce changes in cellular identity in the context of development and reprogramming.

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    • "Cell identity is controlled in large part by the action of transcription factors (TFs) that recognize and bind specific sequences in the genome and regulate gene expression. While approximately half of all TFs are expressed in any one cell type (Vaquerizas et al., 2009), a small number of core TFs are thought to be sufficient to establish control of the gene expression programs that define cell identity (Buganim et al., 2013; Graf and Enver, 2009; Morris and Daley, 2013; Sancho-Martinez et al., 2012; Vierbuchen and Wernig, 2012; Yamanaka, 2012). It would be valuable to identify these core TFs for all cell types; an atlas of candidate core regulators would complement the Encyclopedia of Regulatory DNA Elements (ENCODE) (Rivera and Ren, 2013; Stergachis et al., 2013), guide exploration of the principles of transcriptional regulatory networks, enable more systematic research into the mechanistic and global functions of these key regulators of cell identity, and facilitate advances in direct reprogramming for clinically relevant cell types (Henriques et al., 2013; Iwafuchi-Doi and Zaret, 2014; Soufi et al., 2012; Xie and Ren, 2013). "
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    ABSTRACT: Hundreds of transcription factors (TFs) are expressed in each cell type, but cell identity can be induced through the activity of just a small number of core TFs. Systematic identification of these core TFs for a wide variety of cell types is currently lacking and would establish a foundation for understanding the transcriptional control of cell identity in development, disease, and cell-based therapy. Here, we describe a computational approach that generates an atlas of candidate core TFs for a broad spectrum of human cells. The potential impact of the atlas was demonstrated via cellular reprogramming efforts where candidate core TFs proved capable of converting human fibroblasts to retinal pigment epithelial-like cells. These results suggest that candidate core TFs from the atlas will prove a useful starting point for studying transcriptional control of cell identity and reprogramming in many human cell types.
    Full-text · Article · Oct 2015 · Stem Cell Reports
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    • "This new direct reprograming method featuring defined factors indicates that transdifferentiation can occur across germ layers. Transdifferentiation can be controlled through epigenetic regulation and gene activation [57], [58]. In 2012, Ladewig et al. reported that inhibiting GSK-3β and SMAD signaling during reprogramming increased the efficiency of human iN generation as well as the purity of the resulting iNs [59]. "
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    ABSTRACT: Wnts were previously shown to regulate the neurogenesis of neural stem or progenitor cells. Here, we explored the underlying molecular mechanisms through which Wnt signaling regulates neurotrophins (NTs) in the NT-induced neuronal differentiation of human mesenchymal stem cells (hMSCs). NTs can increase the expression of Wnt1 and Wnt7a in hMSCs. However, only Wnt7a enables the expression of synapsin-1, a synaptic marker in mature neurons, to be induced and triggers the formation of cholinergic and dopaminergic neurons. Human recombinant (hr)Wnt7a and general neuron makers were positively correlated in a dose- and time-dependent manner. In addition, the expression of synaptic markers and neurites was induced by Wnt7a and lithium, a glycogen synthase kinase-3β inhibitor, in the NT-induced hMSCs via the canonical/β-catenin pathway, but was inhibited by Wnt inhibitors and frizzled-5 (Frz5) blocking antibodies. In addition, hrWnt7a triggered the formation of cholinergic and dopaminergic neurons via the non-canonical/c-jun N-terminal kinase (JNK) pathway, and the formation of these neurons was inhibited by a JNK inhibitor and Frz9 blocking antibodies. In conclusion, hrWnt7a enhances the synthesis of synapse and facilitates neuronal differentiation in hMSCS through various Frz receptors. These mechanisms may be employed widely in the transdifferentiation of other adult stem cells.
    Full-text · Article · Aug 2014 · PLoS ONE
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    • "To solve this dilemma, novel strategies for generating functionally mature cells are in high demand. Recently, lineage reprogramming has emerged as an effective method for changing the fate of somatic cells (Vierbuchen and Wernig, 2012). In principle, one cell type can be converted directly to the final mature state of another cell type and can bypass its intermediate states during lineage reprogramming. "
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    ABSTRACT: Obtaining fully functional cell types is a major challenge for drug discovery and regenerative medicine. Currently, a fundamental solution to this key problem is still lacking. Here, we show that functional human induced hepatocytes (hiHeps) can be generated from fibroblasts by overexpressing the hepatic fate conversion factors HNF1A, HNF4A, and HNF6 along with the maturation factors ATF5, PROX1, and CEBPA. hiHeps express a spectrum of phase I and II drug-metabolizing enzymes and phase III drug transporters. Importantly, the metabolic activities of CYP3A4, CYP1A2, CYP2B6, CYP2C9, and CYP2C19 are comparable between hiHeps and freshly isolated primary human hepatocytes. Transplanted hiHeps repopulate up to 30% of the livers of Tet-uPA/Rag2(-/-)/γc(-/-) mice and secrete more than 300 μg/ml human ALBUMIN in vivo. Our data demonstrate that human hepatocytes with drug metabolic function can be generated by lineage reprogramming, thus providing a cell resource for pharmaceutical applications.
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