Pluripotency Factors on Their Lineage Move

Pluripotent stem cells are characterised by continuous self-renewal while maintaining the potential to differentiate into cells of all three germ layers. Regulatory networks of maintaining pluripotency have been described in great detail and, similarly, there is great knowledge on key players that regulate their differentiation. Interestingly, pluripotency has various shades with distinct developmental potential, an observation that coined the term of a ground state of pluripotency. A precise interplay of signalling axes regulates ground state conditions and acts in concert with a combination of key transcription factors. The balance between these transcription factors greatly influences the integrity of the pluripotency network and latest research suggests that minute changes in their expression can strengthen but also collapse the network. Moreover, recent studies reveal different facets of these core factors in balancing a controlled and directed exit from pluripotency. Thereby, subsets of pluripotency-maintaining factors have been shown to adopt new roles during lineage specification and have been globally defined towards neuroectodermal and mesendodermal sets of embryonic stem cell genes. However, detailed underlying insights into how these transcription factors orchestrate cell fate decisions remain largely elusive. Our group and others unravelled complex interactions in the regulation of this controlled exit. Herein, we summarise recent findings and discuss the potential mechanisms involved.

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Endogenous fluctuations of OCT4 and SOX2 bias pluripotent cell fate decisions

10.1101/299073 ◽

2018 ◽

Cited By ~ 4

Author(s):

Daniel Strebinger ◽

Cédric Deluz ◽

Elias T. Friman ◽

Subashika Govindan ◽

Andrea B. Alber ◽

...

Keyword(s):

Transcription Factors ◽

Cell Fate ◽

Es Cells ◽

Embryonic Stem ◽

Chromatin Accessibility ◽

Directed Differentiation ◽

Es Cell ◽

Endogenous Fluctuations ◽

Cell Fate Decisions ◽

Oct4 Protein

AbstractSOX2 and OCT4 are pioneer transcription factors playing a key role in embryonic stem (ES) cell self-renewal and differentiation. However, how temporal fluctuations in their expression levels bias lineage commitment is unknown. Here we generated knock-in reporter fusion ES cell lines allowing to monitor endogenous SOX2 and OCT4 protein fluctuations in living cells and to determine their impact on mesendodermal and neuroectodermal commitment. We found that small differences in SOX2 and OCT4 levels impact cell fate commitment in G1 but not in S phase. Elevated SOX2 levels modestly increased neuroectodermal commitment and decreased mesendodermal commitment upon directed differentiation. In contrast, elevated OCT4 levels strongly biased ES cell towards both neuroectodermal and mesendodermal fates. Using ATAC-seq on ES cells gated for different endogenous SOX2 and OCT4 levels, we found that high OCT4 levels increased chromatin accessibility at differentiation-associated enhancers. This suggests that small endogenous fluctuations of pioneer transcription factors can bias cell fate decisions by concentration-dependent priming of differentiation-associated enhancers.

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Quantifying the Stability of Coupled Genetic and Epigenetic Switches With Variational Methods

Frontiers in Genetics ◽

10.3389/fgene.2020.636724 ◽

2021 ◽

Vol 11 ◽

Author(s):

Amogh Sood ◽

Bin Zhang

Keyword(s):

Gene Regulation ◽

Transcription Factors ◽

Cell Fate ◽

Histone Modifications ◽

Regulatory Networks ◽

Variational Principles ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Cell Fate Decisions ◽

The Impact

The Waddington landscape provides an intuitive metaphor to view development as a ball rolling down the hill, with distinct phenotypes as basins and differentiation pathways as valleys. Since, at a molecular level, cell differentiation arises from interactions among the genes, a mathematical definition for the Waddington landscape can, in principle, be obtained by studying the gene regulatory networks. For eukaryotes, gene regulation is inextricably and intimately linked to histone modifications. However, the impact of such modifications on both landscape topography and stability of attractor states is not fully understood. In this work, we introduced a minimal kinetic model for gene regulation that combines the impact of both histone modifications and transcription factors. We further developed an approximation scheme based on variational principles to solve the corresponding master equation in a second quantized framework. By analyzing the steady-state solutions at various parameter regimes, we found that histone modification kinetics can significantly alter the behavior of a genetic network, resulting in qualitative changes in gene expression profiles. The emerging epigenetic landscape captures the delicate interplay between transcription factors and histone modifications in driving cell-fate decisions.

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Expression Profiles of MicroRNAs in Stem Cells Differentiation

Current Pharmaceutical Biotechnology ◽

10.2174/1389201021666200219092520 ◽

2020 ◽

Vol 21 (10) ◽

pp. 906-918

Author(s):

Hadi Rajabi ◽

Somayeh Aslani ◽

Alireza Abhari ◽

Davoud Sanajou

Keyword(s):

Stem Cells ◽

Transcription Factors ◽

Signaling Pathways ◽

Cell Fate ◽

Adult Stem Cells ◽

Expression Profiles ◽

Embryonic Stem ◽

Stem Cell Differentiation ◽

Cell Fate Decisions ◽

Effective Diagnosis

Stem cells are undifferentiated cells and have a great potential in multilineage differentiation. These cells are classified into adult stem cells like Mesenchymal Stem Cells (MSCs) and Embryonic Stem Cells (ESCs). Stem cells also have potential therapeutic utility due to their pluripotency, self-renewal, and differentiation ability. These properties make them a suitable choice for regenerative medicine. Stem cells differentiation toward functional cells is governed by different signaling pathways and transcription factors. Recent studies have demonstrated the key role of microRNAs in the pathogenesis of various diseases, cell cycle regulation, apoptosis, aging, cell fate decisions. Several types of stem cells have different and unique miRNA expression profiles. Our review summarizes novel regulatory roles of miRNAs in the process of stem cell differentiation especially adult stem cells into a variety of functional cells through signaling pathways and transcription factors modulation. Understanding the mechanistic roles of miRNAs might be helpful in elaborating clinical therapies using stem cells and developing novel biomarkers for the early and effective diagnosis of pathologic conditions.

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Zic3 Is Required for Maintenance of Pluripotency in Embryonic Stem Cells

Molecular Biology of the Cell ◽

10.1091/mbc.e06-07-0624 ◽

2007 ◽

Vol 18 (4) ◽

pp. 1348-1358 ◽

Cited By ~ 85

Author(s):

Linda Shushan Lim ◽

Yuin-Han Loh ◽

Weiwei Zhang ◽

Yixun Li ◽

Xi Chen ◽

...

Keyword(s):

Stem Cells ◽

Transcription Factors ◽

Embryonic Stem Cells ◽

Regulatory Networks ◽

Molecular Mechanisms ◽

Es Cells ◽

Embryonic Stem ◽

Es Cell ◽

Lineage Specification ◽

Mouse Es Cells

Embryonic stem (ES) cell pluripotency is dependent upon sustained expression of the key transcriptional regulators Oct4, Nanog, and Sox2. Dissection of the regulatory networks downstream of these transcription factors has provided critical insight into the molecular mechanisms that regulate ES cell pluripotency and early differentiation. Here we describe a role for Zic3, a member of the Gli family of zinc finger transcription factors, in the maintenance of pluripotency in ES cells. We show that Zic3 is expressed in ES cells and that this expression is repressed upon differentiation. The expression of Zic3 in pluripotent ES cells is also directly regulated by Oct4, Sox2, and Nanog. Targeted repression of Zic3 in human and mouse ES cells by RNA interference–induced expression of several markers of the endodermal lineage. Notably, the expression of Nanog, a key pluripotency regulator and repressor of extraembryonic endoderm specification in ES cells, was significantly reduced in Zic3 knockdown cells. This suggests that Zic3 may prevent endodermal marker expression through Nanog-regulated pathways. Thus our results extend the ES cell transcriptional network beyond Oct4, Nanog, and Sox2, and further establish that Zic3 plays an important role in the maintenance of pluripotency by preventing endodermal lineage specification in embryonic stem cells.

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Embryonic stem cells become mechanoresponsive upon exit from ground state of pluripotency

Open Biology ◽

10.1098/rsob.180203 ◽

2019 ◽

Vol 9 (1) ◽

pp. 180203 ◽

Cited By ~ 2

Author(s):

C. M. Verstreken ◽

C. Labouesse ◽

C. C. Agley ◽

K. J. Chalut

Keyword(s):

Mechanical Stress ◽

Cell Fate ◽

Ground State ◽

Es Cells ◽

Embryonic Stem ◽

Cell Fate Decisions ◽

Mechanical Signals ◽

Broad Array ◽

Transcriptional Changes ◽

The Impact

Stem cell fate decisions are driven by a broad array of signals, both chemical and mechanical. Although much progress has been made in our understanding of the impact of chemical signals on cell fate choice, much less is known about the role and influence of mechanical signalling, particularly in embryonic stem (ES) cells. Many studies use substrates with different stiffness to study mechanical signalling, but changing substrate stiffness can induce secondary effects which are difficult to disentangle from the direct effects of forces/mechanical signals. To probe the direct impact of mechanical stress on cells, we developed an adaptable cell substrate stretcher to exert specific, reproducible forces on cells. Using this device to test the response of ES cells to tensile strain, we found that cells experienced a transient influx of calcium followed by an upregulation of the so-called immediate and early genes. On longer time scales, however, ES cells in ground state conditions were largely insensitive to mechanical stress. Nonetheless, as ES cells exited the ground state, their susceptibility to mechanical signals increased, resulting in broad transcriptional changes. Our findings suggest that exit from ground state of pluripotency is unaffected by mechanical signals, but that these signals could become important during the next stage of lineage specification. A better understanding of this process could improve our understanding of cell fate choice in early development and improve protocols for differentiation guided by mechanical cues.

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Controlling gene networks and cell fate with precision-targeted DNA-binding proteins and small-molecule-based genome readers

Biochemical Journal ◽

10.1042/bj20140400 ◽

2014 ◽

Vol 462 (3) ◽

pp. 397-413 ◽

Cited By ~ 10

Author(s):

Asuka Eguchi ◽

Garrett O. Lee ◽

Fang Wan ◽

Graham S. Erwin ◽

Aseem Z. Ansari

Keyword(s):

Transcription Factors ◽

Dna Binding ◽

Cell Fate ◽

Gene Networks ◽

Regulatory Networks ◽

Transcriptional Networks ◽

Cell Fate Decisions ◽

Differentiated Cells ◽

Dna Binding Factors ◽

Dna Binders

Transcription factors control the fate of a cell by regulating the expression of genes and regulatory networks. Recent successes in inducing pluripotency in terminally differentiated cells as well as directing differentiation with natural transcription factors has lent credence to the efforts that aim to direct cell fate with rationally designed transcription factors. Because DNA-binding factors are modular in design, they can be engineered to target specific genomic sequences and perform pre-programmed regulatory functions upon binding. Such precision-tailored factors can serve as molecular tools to reprogramme or differentiate cells in a targeted manner. Using different types of engineered DNA binders, both regulatory transcriptional controls of gene networks, as well as permanent alteration of genomic content, can be implemented to study cell fate decisions. In the present review, we describe the current state of the art in artificial transcription factor design and the exciting prospect of employing artificial DNA-binding factors to manipulate the transcriptional networks as well as epigenetic landscapes that govern cell fate.

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GATA-targeted compounds modulate cardiac subtype cell differentiation in dual reporter stem cell line

Stem Cell Research & Therapy ◽

10.1186/s13287-021-02259-z ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Mika J. Välimäki ◽

Robert S. Leigh ◽

Sini M. Kinnunen ◽

Alexander R. March ◽

Ana Hernández de Sande ◽

...

Keyword(s):

Gene Expression ◽

Progenitor Cells ◽

Cell Fate ◽

Reporter Gene ◽

Gene Regulatory Networks ◽

Regulatory Networks ◽

Cell Fate Decisions ◽

Dual Reporter ◽

Gene Regulatory ◽

Reporter Gene Assays

AbstractBackgroundPharmacological modulation of cell fate decisions and developmental gene regulatory networks holds promise for the treatment of heart failure. Compounds that target tissue-specific transcription factors could overcome non-specific effects of small molecules and lead to the regeneration of heart muscle following myocardial infarction. Due to cellular heterogeneity in the heart, the activation of gene programs representing specific atrial and ventricular cardiomyocyte subtypes would be highly desirable. Chemical compounds that modulate atrial and ventricular cell fate could be used to improve subtype-specific differentiation of endogenous or exogenously delivered progenitor cells in order to promote cardiac regeneration.MethodsTranscription factor GATA4-targeted compounds that have previously shown in vivo efficacy in cardiac injury models were tested for stage-specific activation of atrial and ventricular reporter genes in differentiating pluripotent stem cells using a dual reporter assay. Chemically induced gene expression changes were characterized by qRT-PCR, global run-on sequencing (GRO-seq) and immunoblotting, and the network of cooperative proteins of GATA4 and NKX2-5 were further explored by the examination of the GATA4 and NKX2-5 interactome by BioID. Reporter gene assays were conducted to examine combinatorial effects of GATA-targeted compounds and bromodomain and extraterminal domain (BET) inhibition on chamber-specific gene expression.ResultsGATA4-targeted compounds 3i-1000 and 3i-1103 were identified as differential modulators of atrial and ventricular gene expression. More detailed structure-function analysis revealed a distinct subclass of GATA4/NKX2-5 inhibitory compounds with an acetyl lysine-like domain that contributed to ventricular cells (%Myl2-eGFP+). Additionally, BioID analysis indicated broad interaction between GATA4 and BET family of proteins, such as BRD4. This indicated the involvement of epigenetic modulators in the regulation of GATA-dependent transcription. In this line, reporter gene assays with combinatorial treatment of 3i-1000 and the BET bromodomain inhibitor (+)-JQ1 demonstrated the cooperative role of GATA4 and BRD4 in the modulation of chamber-specific cardiac gene expression.ConclusionsCollectively, these results indicate the potential for therapeutic alteration of cell fate decisions and pathological gene regulatory networks by GATA4-targeted compounds modulating chamber-specific transcriptional programs in multipotent cardiac progenitor cells and cardiomyocytes. The compound scaffolds described within this study could be used to develop regenerative strategies for myocardial regeneration.

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Transcriptional Networks Regulating Embryonic Stem Cell Fate Decisions

Regulatory Networks in Stem Cells ◽

10.1007/978-1-60327-227-8_8 ◽

2009 ◽

pp. 87-100

Author(s):

Emily Walker ◽

William L. Stanford

Keyword(s):

Stem Cell ◽

Embryonic Stem Cell ◽

Cell Fate ◽

Embryonic Stem ◽

Transcriptional Networks ◽

Stem Cell Fate ◽

Cell Fate Decisions

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Leveraging high-powered RNA-Seq datasets to improve inference of regulatory activity in single-cell RNA-Seq data

10.1101/553040 ◽

2019 ◽

Cited By ~ 1

Author(s):

Ning Wang ◽

Andrew E. Teschendorff

Keyword(s):

Transcription Factors ◽

Single Cell ◽

Cell Fate ◽

Regulatory Networks ◽

Large Scale ◽

Single Cells ◽

Differential Expression Analysis ◽

Dropout Rate ◽

Rna Seq ◽

Regulatory Activity

AbstractInferring the activity of transcription factors in single cells is a key task to improve our understanding of development and complex genetic diseases. This task is, however, challenging due to the relatively large dropout rate and noisy nature of single-cell RNA-Seq data. Here we present a novel statistical inference framework called SCIRA (Single Cell Inference of Regulatory Activity), which leverages the power of large-scale bulk RNA-Seq datasets to infer high-quality tissue-specific regulatory networks, from which regulatory activity estimates in single cells can be subsequently obtained. We show that SCIRA can correctly infer regulatory activity of transcription factors affected by high technical dropouts. In particular, SCIRA can improve sensitivity by as much as 70% compared to differential expression analysis and current state-of-the-art methods. Importantly, SCIRA can reveal novel regulators of cell-fate in tissue-development, even for cell-types that only make up 5% of the tissue, and can identify key novel tumor suppressor genes in cancer at single cell resolution. In summary, SCIRA will be an invaluable tool for single-cell studies aiming to accurately map activity patterns of key transcription factors during development, and how these are altered in disease.

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