• Drp1
• Dynein
• Oct4
• Wnt signaling
• mesenchymal-to-epithelial transition

• Mitochondria/metabolism
• Animals
• Epithelial-Mesenchymal Transition
• Kruppel-Like Factor 4
• Mice
• Mitochondrial Dynamics
• Cadherins/metabolism
• beta Catenin/metabolism
• beta Catenin/genetics
• Octamer Transcription Factor-3/metabolism
• Octamer Transcription Factor-3/genetics
• Dynamins/metabolism
• Cellular Reprogramming
• Dyneins/metabolism
• SOXB1 Transcription Factors/metabolism
• SOXB1 Transcription Factors/genetics
• Pluripotent Stem Cells/metabolism
• Pluripotent Stem Cells/cytology

• cell‐reprogramming‐responsive hydrogels
• dynamic responsiveness
• induced pluripotent stem cells
• induction of pluripotency
• phase‐separated biomolecular condensates

• Induced Pluripotent Stem Cells/cytology
• Induced Pluripotent Stem Cells/metabolism
• Animals
• Cellular Reprogramming
• Mice
• Humans
• Hydrogels/chemistry
• Biomolecular Condensates/metabolism
• Biomolecular Condensates/chemistry
• YAP-Signaling Proteins

• T2288102 Fundamental Research Center Project of National Natural Science Foundation of China
• 2018YFA0208900 National Key R&D Program of China
• XDB36000000 Strategic Priority Research Program of Chinese Academy of Sciences

• CYLD
• ECM organization
• EMT
• MET
• TGF-beta
• dedifferentiation
• pluripotency
• somatic cell reprogramming

• ΟΠΣ 5000432 State Scholarships Foundation
• Cooperative Grants Synergasia I #969 Greek General Secretariat for Research and Technology (GSRT)
• EL0084 European Economic Area
• MIS 5002802 The Greek Research Infrastructure for Personalised Medicine (pMED-GR)
• MIS 93225 Bodossaki Foundation

• Humans
• Mice
• Animals
• Cellular Reprogramming
• Transcription Factors/metabolism
• Embryonic Stem Cells/metabolism
• DNA/metabolism
• Point Mutation
• Octamer Transcription Factor-3/genetics
• Octamer Transcription Factor-3/metabolism
• SOXB1 Transcription Factors/genetics
• SOXB1 Transcription Factors/metabolism
• Cell Differentiation/genetics
• Induced Pluripotent Stem Cells/metabolism
• SOXF Transcription Factors/genetics
• SOXF Transcription Factors/metabolism

• Research Grants Council of Hong Kong General Research Funds
• Collaborative Research Fund
• Health and Medical Research Fund
• National Natural Science Foundation of China
• Innovation Technology Commission Funding

• AhR
• cell differentiation
• mesenchymal stem cell

• Humans
• Osteogenesis/genetics
• Polychlorinated Dibenzodioxins/toxicity
• Cell Differentiation
• Fibroblast Growth Factors/metabolism
• Mesenchymal Stem Cells/metabolism
• Cells, Cultured
• Osteoblasts

• P30 ES025128 NIEHS NIH HHS
• T32 ES007046 NIEHS NIH HHS
• National Institute of Environmental Health Sciences
• Molecular Pathways to Pathogenesis in Toxicology
• Center for Human Health and the Environment

• cell biology
• computational biology and bioinformatics
• stem cells

• Cell Differentiation/genetics
• Lymphoid Enhancer-Binding Factor 1/genetics
• Lymphoid Enhancer-Binding Factor 1/metabolism
• Mechanotransduction, Cellular
• Signal Transduction
• Transcription Factors/metabolism
• beta Catenin/genetics
• beta Catenin/metabolism

• DP2 AT012345 NCCIH NIH HHS
• Ministry of Education Tier-3 program
• Eidgenössische Technische Hochschule Zürich
• Paul Scherrer Institut
• National Science Foundation

• Cancer cell reprogramming
• Cancer-iPSCs
• Induced pluripotent cancer cells
• Induced pluripotent stem cells

Experimental and computational efforts are constantly made to elucidate mechanisms controlling cell fate decisions during development and reprogramming. One powerful computational method is to consider cell commitment and reprogramming as movements in an energy landscape. Here, we develop Computation of Energy Landscapes of Logical Gene Networks (CELLoGeNe), which maps Boolean implementation of gene regulatory networks (GRNs) into energy landscapes. CELLoGeNe removes inadvertent symmetries in the energy landscapes normally arising from standard Boolean operators. Furthermore, CELLoGeNe provides tools to visualize and stochastically analyze the shapes of multi-dimensional energy landscapes corresponding to epigenetic landscapes for development and reprogramming. We demonstrate CELLoGeNe on two GRNs governing different aspects of induced pluripotent stem cells, identifying experimentally validated attractors and revealing potential reprogramming roadblocks. CELLoGeNe is a general framework that can be applied to various biological systems offering a broad picture of intracellular dynamics otherwise inaccessible with existing methods.

•

CELLoGeNe – Computation of Energy Landscapes of Logical Gene Networks

• 1331AFG075 Agency for Science, Technology and Research
• Agency for Science, Technology and Research

The Wnt/β-catenin pathway is involved in development, cancer, and embryonic stem cell (ESC) maintenance; its dual role in stem cell self-renewal and differentiation is still controversial. Here, by applying an in vitro system enabling inducible gene expression control, we report that moderate induction of transcriptionally active exogenous β-catenin in β-catenin null mouse ESCs promotes epiblast-like cell (EpiLC) derivation in vitro. Instead, in wild-type cells, moderate chemical pre-activation of the Wnt/β-catenin pathway promotes EpiLC in vitro derivation. Finally, we suggest that moderate β-catenin levels in β-catenin null mouse ESCs favor early stem cell commitment toward mesoderm if the exogenous protein is induced only in the “ground state” of pluripotency condition, or endoderm if the induction is maintained during the differentiation. Overall, our results confirm previous findings about the role of β-catenin in pluripotency and differentiation, while indicating a role for its doses in promoting specific differentiation programs.

•

Moderate β-catenin levels promote EpiLCs derivation in vitro

• Chromatin accessibility
• Gene dosage compensation
• Gene regulatory networks
• X chromosome inactivation
• X chromosome reactivation
• X chromosome upregulation
• iPSC reprogramming

• Alleles
• Aneuploidy
• Animals
• Cellular Reprogramming/genetics
• Chromatin/metabolism
• Gene Regulatory Networks
• Induced Pluripotent Stem Cells/metabolism
• Mice
• RNA-Seq
• Single-Cell Analysis
• Transcription Factors/metabolism
• Transcription, Genetic
• X Chromosome
• X Chromosome Inactivation

• The Research Foundation–Flanders (FWO)
• KU Leuven
• European Union’s Horizon 2020
• The Research Foundation–Flanders (FWO)

• Macrophages
• Neurons
• Pluripotency
• Stem cells
• Trans-differentiation
• miRNA

• Biological Phenomena
• Cell Differentiation/genetics
• Cell Line
• Humans
• Induced Pluripotent Stem Cells/metabolism
• Transcriptome

• R01 CA253368 NCI NIH HHS
• U54 GM128729 NIGMS NIH HHS
• Z01 ES071006 Intramural NIH HHS
• NIEHS
• NIMHD

• Cell reprogramming
• Epigenetics
• Reprogramming barriers/roadblocks
• Transcription factors
• induced Pluripotent Stem Cells

During somatic reprogramming, Yamanaka’s pioneer factors regulate a complex sequence of molecular events leading to the activation of a network of pluripotency factors, ultimately resulting in the acquisition and maintenance of a pluripotent state. Here, we show that, contrary to the pluripotency factors studied so far, overexpression of Mybl2 inhibits somatic reprogramming. Our results demonstrate that Mybl2 levels are crucial to the dynamics of the reprogramming process. Mybl2 overexpression changes chromatin conformation, affecting the accessibility of pioneer factors to the chromatin and promoting accessibility for early immediate response genes known to be reprogramming blockers. These changes in the chromatin landscape ultimately lead to a deregulation of key genes that are important for the mesenchymal-to-epithelial transition. This work defines Mybl2 level as a gatekeeper for the initiation of reprogramming, providing further insights into the tight regulation and required coordination of molecular events that are necessary for changes in cell fate identity during the reprogramming process.

•

Deletion and overexpression of MYBL2 pluripotency factor inhibit somatic reprogramming

• AP1
• ATAC-sequencing
• Jun
• Sox2
• chromatin landscape
• chromatin remodeling
• induced pluripotent stem cells
• mesenchymal-to-epithelial transition
• reprogramming blockers
• somatic reprogramming

• 3T3-L1 Cells
• Adipocytes/cytology
• Adipocytes/metabolism
• Adipose Tissue, White/metabolism
• Adiposity
• Animals
• Binding Sites
• Cell Differentiation
• Cell Proliferation
• Diet, High-Fat
• Gain of Function Mutation/genetics
• Homeostasis
• Humans
• Macrophage Activation
• Mice
• Mice, Knockout
• Obesity/genetics
• Obesity/pathology
• Organ Size
• RAW 264.7 Cells
• Serum Amyloid A Protein/metabolism
• TCF Transcription Factors/metabolism
• Transcription, Genetic
• Wnt Signaling Pathway
• beta Catenin/genetics

• National Natural Science Foundation of China
• Foundation of Shanghai Municipal Commission of Health and Family Planning
• National Key Research and Development Program of China
• Shanghai Rising-Star Program
• Shanghai Municipal Education Commission-Gaofeng Clinical Medicine Grant Support
• 973 Foundation

• Epithelialization
• Induced pluripotent stem cell (iPSC)
• LIN28
• LIN41
• Reprogramming efficiency
• WNT

Reprogramming of somatic cells to induced pluripotent stem cells, by overexpressing certain factors referred to as the reprogramming factors, can revolutionize regenerative medicine. To provide a coherent description of induced pluripotency from the gene regulation perspective, we use 35 microarray datasets to construct a reprogramming gene regulatory network. Comprising 276 nodes and 4471 links, the resulting network is, to the best of our knowledge, the largest gene regulatory network constructed for human fibroblast reprogramming and it is the only one built using a large number of experimental datasets. To build the network, a model that relates the expression profiles of the initial (fibroblast) and final (induced pluripotent stem cell) states is proposed and the model parameters (link strengths) are fitted using the experimental data. Twenty nine additional experimental datasets are collectively used to test the model/network, and good agreement between experimental and predicted gene expression profiles is found. We show that the model in conjunction with the constructed network can make useful predictions. For example, we demonstrate that our approach can incorporate the effect of reprogramming factor stoichiometry and that its predictions are consistent with the experimentally observed trends in reprogramming efficiency when the stoichiometric ratios vary. Using our model/network, we also suggest new (not used in training of the model) candidate sets of reprogramming factors, many of which have already been experimentally verified. These results suggest our model/network can potentially be used in devising new recipes for induced pluripotency with higher efficiencies. Additionally, we classify the links of the network into three classes of different importance, prioritizing them for experimental verification. We show that many of the links in the top ranked class are experimentally known to be important in reprogramming. Finally, comparing with other methods, we show that using our model is advantageous.

• Cellular Reprogramming/physiology
• Fibroblasts/metabolism
• Gene Expression Profiling
• Gene Expression Regulation
• Gene Regulatory Networks
• Humans
• Induced Pluripotent Stem Cells/metabolism

• ZIA LM092404 NLM NIH HHS
• National Institutes of Health

By post-transcriptionally regulating multiple target transcripts, microRNAs (miRNAs or miR) play important biological functions. H1 embryonic stem cells (hESCs) and NTera-2 embryonal carcinoma cells (ECCs) are two of the most widely used human pluripotent model cell lines, sharing several characteristics, including the expression of miRNAs associated to the pluripotent state or with differentiation. However, how each of these miRNAs functionally impacts the biological properties of these cells has not been systematically evaluated.

We investigated the effects of 31 miRNAs on NTera-2 and H1 hESCs, by transfecting miRNA mimics. Following 3–4 days of culture, cells were stained for the pluripotency marker OCT4 and the G2 cell-cycle marker Cyclin B1, and nuclei and cytoplasm were co-stained with Hoechst and Cell Mask Blue, respectively. By using automated quantitative fluorescence microscopy (i.e., high-content screening (HCS)), we obtained several morphological and marker intensity measurements, in both cell compartments, allowing the generation of a multiparametric miR-induced phenotypic profile describing changes related to proliferation, cell cycle, pluripotency, and differentiation.

Despite the overall similarities between both cell types, some miRNAs elicited cell-specific effects, while some related miRNAs induced contrasting effects in the same cell. By identifying transcripts predicted to be commonly targeted by miRNAs inducing similar effects (profiles grouped by hierarchical clustering), we were able to uncover potentially modulated signaling pathways and biological processes, likely mediating the effects of the microRNAs on the distinct groups identified. Specifically, we show that miR-363 contributes to pluripotency maintenance, at least in part, by targeting NOTCH1 and PSEN1 and inhibiting Notch-induced differentiation, a mechanism that could be implicated in naïve and primed pluripotent states.

We present the first multiparametric high-content microRNA functional screening in human pluripotent cells. Integration of this type of data with similar data obtained from siRNA screenings (using the same HCS assay) could provide a large-scale functional approach to identify and validate microRNA-mediated regulatory mechanisms controlling pluripotency and differentiation.

The online version of this article (10.1186/s13287-019-1318-6) contains supplementary material, which is available to authorized users.

• Cell differentiation
• Human embryonic stem cells
• MicroRNA
• Microscopy, fluorescence
• Pluripotent stem cells
• Receptors, Notch

Cells have developed numerous adaptation mechanisms to external cues by controlling signaling-pathway activity, both qualitatively and quantitatively. The Wnt/β-catenin pathway is a highly conserved signaling pathway involved in many biological processes, including cell proliferation, differentiation, somatic cell reprogramming, development, and cancer. The activity of the Wnt/β-catenin pathway and the temporal dynamics of its effector β-catenin are tightly controlled by complex regulations. The latter encompass feedback loops within the pathway (e.g., a negative feedback loop involving Axin2, a β-catenin transcriptional target) and crosstalk interactions with other signaling pathways. Here, we provide a review shedding light on the coupling between Wnt/β-catenin activation levels and fluctuations across processes and cellular systems; in particular, we focus on development, in vitro pluripotency maintenance, and cancer. Possible mechanisms originating Wnt/β-catenin dynamic behaviors and consequently driving different cellular responses are also reviewed, and new avenues for future research are suggested.

• cancer
• canonical Wnt pathway
• development
• pluripotency
• signaling dynamics
• β-catenin

• Animals
• Cell Differentiation
• Humans
• Wnt Signaling Pathway
• beta Catenin/genetics
• beta Catenin/metabolism

• BB/L01386X/1 Biotechnology and Biological Sciences Research Council
• MR/N021444/1 Medical Research Council

• Comparative epigenomics
• Reprogramming
• iPS cells

The technology to derive embryonic and induced pluripotent stem cells from early embryonic stages and adult somatic cells, respectively, emerged as a powerful resource to enable the establishment of new in vitro models, which recapitulate early developmental processes and disease. Additionally, pluripotent stem cells (PSCs) represent an invaluable source of relevant differentiated cell types with immense potential for regenerative medicine and cell replacement therapies. Pluripotent stem cells support self-renewal, potency and proliferation for extensive periods of culture in vitro. However, the core pathways that rule each of these cellular features specific to PSCs only recently began to be clarified. The Wnt signaling pathway is pivotal during early embryogenesis and is central for the induction and maintenance of the pluripotency of PSCs. Signaling by the Wnt family of ligands is conveyed intracellularly by the stabilization of β-catenin in the cytoplasm and in the nucleus, where it elicits the transcriptional activity of T-cell factor (TCF)/lymphoid enhancer factor (LEF) family of transcription factors. Interestingly, in PSCs, the Wnt/β-catenin–TCF/LEF axis has several unrelated and sometimes opposite cellular functions such as self-renewal, stemness, lineage commitment and cell cycle regulation. In addition, tight control of the Wnt signaling pathway enhances reprogramming of somatic cells to induced pluripotency. Several recent research efforts emphasize the pleiotropic functions of the Wnt signaling pathway in the pluripotent state. Nonetheless, conflicting results and unanswered questions still linger. In this review, we will focus on the diverse functions of the canonical Wnt signaling pathway on the developmental processes preceding embryo implantation, as well as on its roles in pluripotent stem cell biology such as self-renewal and cell cycle regulation and somatic cell reprogramming.

• Wnt/β-catenin pathway
• cell cycle
• embryonic stem cells
• pre-implantation development
• somatic cell reprogramming

• ES cells
• Wnt signaling
• iPS cells
• somatic reprogramming

• Embryonic stem cells
• GATA6
• JUP
• TCF1
• β-catenin

• Animals
• Cell Differentiation/genetics
• Cell Transdifferentiation/genetics
• Cellular Reprogramming/genetics
• Databases, Genetic
• Embryonic Stem Cells/metabolism
• Fibroblasts/metabolism
• Humans
• Induced Pluripotent Stem Cells/metabolism
• Internet
• Mice
• Myocytes, Cardiac/metabolism
• Software
• Transcriptome

Carcinomas are phenotypically arrayed along an epithelial–mesenchymal transition (EMT) spectrum, a developmental program currently exploited to understand the acquisition of drug resistance through a re‐routing of growth factor signaling. This review collates the current approaches employed in developing therapeutics against cancer‐associated EMT, and provides an assessment of their respective strengths and drawbacks. We reflect on the close relationship between EMT and chemoresistance against current targeted therapeutics, with a special focus on the epigenetic mechanisms that link these processes. This prompts the hypothesis that carcinoma‐associated EMT shares a common epigenetic pathway to cellular plasticity as somatic cell reprogramming during tissue repair and regeneration. Indeed, their striking resemblance suggests that EMT in carcinoma is a pathological adaptation of an intrinsic program of cellular plasticity that is crucial to tissue homeostasis. We thus propose a revised approach that targets the epigenetic mechanisms underlying pathogenic EMT to arrest cellular plasticity regardless of upstream cancer‐driving mutations.

• EMT spectrum
• cancer therapeutics
• cellular plasticity
• drug discovery
• drug resistance
• epithelial-mesenchymal transition

• deterministic models
• energy landscape
• reprogramming
• stem cell commitment

• The Swedish Research Council
• The Swedish Foundation for Strategic Research

• Sostdc1
• secreted factors
• somatic cell reprogramming

Mouse and rat embryonic stem cell (ESC) self-renewal can be maintained by dual inhibition of glycogen synthase kinase 3 (GSK3) and mitogen-activated protein kinase kinase (MEK). Inhibition of GSK3 promotes ESC self-renewal by abrogating T-cell factor 3 (TCF3)-mediated repression of the pluripotency network. How inhibition of MEK mediates ESC self-renewal, however, remains largely unknown. Here, we show that inhibition of MEK can significantly suppress lymphoid enhancer factor 1 (LEF1) expression in mouse ESCs. Knockdown or knockout of Lef1 partially mimics the self-renewal-promoting effect of MEK inhibitors. Moreover, depletion of both Tcf3 and Lef1 enables maintenance of undifferentiated mouse ESCs without exogenous factors, cytokines or inhibitors. Transcriptome resequencing analysis reveals that LEF1 is closely associated with endoderm specification in ESCs. Thus, our study adds support to the notion that the key to maintaining the ESC ground state is to shield ESCs from differentiative cues.

Summary: Depletion of Lef1 and Tcf3 shows that ESCs could be shielded from differentiative cues to maintain the ESC ground state.

• Differentiation
• Embryonic stem cells
• LEF1
• Self-renewal
• TCF3

• Cell reprogramming
• Induced pluripotent stem cells
• Leucine-rich repeat-containing G-protein coupled receptor 5

• Klf4
• Oct4
• Sox2
• collaborative binding
• enhancers
• induced pluripotent stem cells
• pluripotency
• reprogramming
• transcription factors

• Cellular reprogramming
• Ten-eleven translocated 1 (TET1) gene
• WNT signaling
• miR-29

• acetylation
• embryonic stem cells
• histone
• plasticity
• reprogramming

• Animals
• Carcinogenesis
• Cell Differentiation
• Cell Proliferation
• Chromatin/metabolism
• Gene Expression Regulation
• Humans
• Lysine Acetyltransferases/metabolism
• Neoplasms/physiopathology

• R01 GM067718 NIGMS NIH HHS
• CIHR

Treatment with several Wnt/β-catenin signaling pathway regulators can change the cellular reprogramming efficiency; however, the dynamics and role of endogenous Wnt/β-catenin signaling in reprogramming remain largely unanswered. Here we identify the upregulation of WNT2 and subsequent β-catenin nuclear accumulation as key events in reprogramming. Transient nuclear accumulation of β-catenin occurs early in MEF reprogramming. Wnt2 is strongly expressed in the early stage of reprogramming. Wnt2 knockdown suppresses the nuclear accumulation of β-catenin and reduces the reprogramming efficiency. WNT2 overexpression promotes β-catenin nuclear accumulation and enhances the reprogramming efficiency. WNT2 contributes to the promotion of cell proliferation. Experiments with several drugs that control the Wnt pathway also indicate the importance of β-catenin nuclear accumulation in reprogramming. Our findings reveal the role of WNT2/β-catenin signaling in reprogramming.

•

Nuclear accumulation of β-catenin occurs in the early stage of MEF reprogramming

• cell fates
• cis-regulatory elements
• embryonic stem cells
• induced pluripotent stem cells
• neural specification
• neural stem cells
• temporal regulation

• Cell proliferation
• Cell reprogramming
• Induced pluripotent stem cell (iPSC)

Stem cells balance their self-renewal and differentiation potential by integrating environmental signals with the transcriptional regulatory network. The maintenance of cell identity and/or cell lineage commitment relies on the interplay of multiple factors including signaling pathways, transcription factors, and the epigenetic machinery. These regulatory modules are strongly interconnected and they influence the pattern of gene expression of stem cells, thus guiding their cellular fate. Embryonic stem cells (ESCs) represent an invaluable tool to study this interplay, being able to indefinitely self-renew and to differentiate towards all three embryonic germ layers in response to developmental cues. In this review, we highlight those mechanisms of signaling to chromatin, which regulate chromatin modifying enzymes, histone modifications, and nucleosome occupancy. In addition, we report the molecular mechanisms through which signaling pathways affect both the epigenetic and the transcriptional state of ESCs, thereby influencing their cell identity. We propose that the dynamic nature of oscillating signaling and the different regulatory network topologies through which those signals are encoded determine specific gene expression programs, leading to the fluctuation of ESCs among multiple pluripotent states or to the establishment of the necessary conditions to exit pluripotency.

• Chemical biology
• ciPSC
• epigenetics
• iPSC
• patent
• reprogramming
• small molecule
• stem cell

• C/EBPα
• Cellular reprogramming
• Mbd3
• iPSCs
• p53

The differentiated state of somatic cells provides barriers for the derivation of induced pluripotent stem cells (iPSCs). To address why some cell types reprogram more readily than others, we studied the effect of combined modulation of cellular signaling pathways. Surprisingly, inhibition of transforming growth factor β (TGF-β) together with activation of Wnt signaling in the presence of ascorbic acid allows >80% of murine fibroblasts to acquire pluripotency after 1 week of reprogramming factor expression. In contrast, hepatic and blood progenitors predominantly required only TGF-β inhibition or canonical Wnt activation, respectively, to reprogram at efficiencies approaching 100%. Strikingly, blood progenitors reactivated endogenous pluripotency loci in a highly synchronous manner, and we demonstrate that expression of specific chromatin-modifying enzymes and reduced TGF-β/mitogen-activated protein (MAP) kinase activity are intrinsic properties associated with the unique reprogramming response of these cells. Our observations define cell-type-specific requirements for the rapid and synchronous reprogramming of somatic cells.

•

A three-compound mix drives rapid and efficient MEF reprogramming