Nanog is Dispensable for the Generation of Induced Pluripotent Stem Cells
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE53832 | GEO | 2014/01/28
SECONDARY ACCESSION(S): PRJNA233503
REPOSITORIES: GEO
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Project description:Cellular reprogramming from somatic cells to induced pluripotent stem cells (iPSCs) can be achieved through forced expression of the transcription factors Oct4, Klf4, Sox2 and c-Myc (OKSM). These factors, in combination with environmental cues, induce a stable intrinsic pluripotency network that confers indefinite self-renewal capacity on iPSCs. In addition to Oct4 and Sox2, the homeodomain-containing transcription factor Nanog is an integral part of the pluripotency network. Although Nanog expression is not required for the maintenance of pluripotent stem cells, it has been reported to be essential for the establishment of both embryonic stem cells (ESCs) from blastocysts and iPSCs from somatic cells. Here we revisit the role of Nanog in direct reprogramming. Surprisingly, we find that Nanog is dispensable for iPSC formation under optimized culture conditions. We further document that Nanog-deficient iPSCs are transcriptionally highly similar to wild-type iPSCs and support the generation of teratomas and chimeric mice. Lastly, we provide evidence that the presence of ascorbic acid in the culture media is critical for overcoming the previously observed reprogramming block of Nanog knockout cells. Comparison of Nanog KO iPSCs to WT pluripotent cells.
2014-01-28 | E-GEOD-53832 | biostudies-arrayexpress
Project description:Brief expression of pluripotency-associated factors such as OCT4, KLF4, SOX2 and c-MYC (OKSM), in combination with differentiation-inducing signals, was reported to trigger transdifferentiation of fibroblasts into alternative cell types. Here, we show that OKSM expression gives rise to both induced pluripotent stem cells (iPSCs) and iNSCs under conditions that were previously shown to induce only NSC transdifferentiation. Fibroblast-derived iNSC colonies silenced retroviral transgenes and reactivated silenced X chromosomes, both hallmarks of pluripotent stem cells. Moreover, lineage tracing via an Oct4-CreER labeling system demonstrated that virtually all iNSC colonies originate from cells transiently expressing Oct4, whereas ablation of Oct4-positive cells prevented iNSC formation. Lastly, use of an alternative transdifferentiation cocktail that lacks OCT4 and was reportedly unable to support induced pluripotency, yielded iPSCs and iNSCs carrying the Oct4-CreER-derived lineage label. Together, these data suggest that iNSC generation using OKSM and related reprogramming factors requires passage through a transient iPSC state. 5 samples were anlyzed in total, 2 induced pluripotent stem cells (iPSCs), 1 neural stem cells (NSCs) and 2 induced NSCs (iNSCs)
2015-04-30 | E-GEOD-65951 | biostudies-arrayexpress
Project description:Brief expression of pluripotency-associated factors such as Oct4, Klf4, Sox2 and c-Myc (OKSM), in combination with differentiation-inducing signals, has been reported to trigger transdifferentiation of fibroblasts into other cell types. Here we show that OKSM expression in mouse fibroblasts gives rise to both induced pluripotent stem cells (iPSCs) and induced neural stem cells (iNSCs) under conditions previously shown to induce only iNSCs. Fibroblast-derived iNSC colonies silenced retroviral transgenes and reactivated silenced X chromosomes, both hallmarks of pluripotent stem cells. Moreover, lineage tracing with an Oct4-CreER labeling system demonstrated that virtually all iNSC colonies originated from cells transiently expressing Oct4, whereas ablation of Oct4+ cells prevented iNSC formation. Lastly, an alternative transdifferentiation cocktail that lacks Oct4 and was reportedly unable to support induced pluripotency yielded iPSCs and iNSCs carrying the Oct4-CreER-derived lineage label. Together, these data suggest that iNSC generation from fibroblasts using OKSM and other pluripotency-related reprogramming factors requires passage through a transient iPSC state.
2015-04-30 | GSE65951 | GEO
Project description:Chickarmane2008 - Stem cell lineage - NANOG GATA-6 switch
In this work, a dynamical model of lineage
determination based upon a minimal circuit, as discussed in PMID: 17215298
, which contains the Oct4/Sox2/Nanog core as well its interaction
with a few other key genes is discussed.
This model is described in the article:
A computational model for understanding stem cell, trophectoderm and endoderm lineage determination.
Chickarmane V, Peterson C
PloS one. 2008, 3(10):e3478
Abstract:
BACKGROUND: Recent studies have associated the transcription factors, Oct4, Sox2 and Nanog as parts of a self-regulating network which is responsible for maintaining embryonic stem cell properties: self renewal and pluripotency. In addition, mutual antagonism between two of these and other master regulators have been shown to regulate lineage determination. In particular, an excess of Cdx2 over Oct4 determines the trophectoderm lineage whereas an excess of Gata-6 over Nanog determines differentiation into the endoderm lineage. Also, under/over-expression studies of the master regulator Oct4 have revealed that some self-renewal/pluripotency as well as differentiation genes are expressed in a biphasic manner with respect to the concentration of Oct4. METHODOLOGY/
PRINCIPAL FINDINGS: We construct a dynamical model of a minimalistic network, extracted from ChIP-on-chip and microarray data as well as literature studies. The model is based upon differential equations and makes two plausible assumptions; activation of Gata-6 by Oct4 and repression of Nanog by an Oct4-Gata-6 heterodimer. With these assumptions, the results of simulations successfully describe the biphasic behavior as well as lineage commitment. The model also predicts that reprogramming the network from a differentiated state, in particular the endoderm state, into a stem cell state, is best achieved by over-expressing Nanog, rather than by suppression of differentiation genes such as Gata-6.
CONCLUSIONS: The computational model provides a mechanistic understanding of how different lineages arise from the dynamics of the underlying regulatory network. It provides a framework to explore strategies of reprogramming a cell from a differentiated state to a stem cell state through directed perturbations. Such an approach is highly relevant to regenerative medicine since it allows for a rapid search over the host of possibilities for reprogramming to a stem cell state.
This model is hosted on BioModels Database
and identified
by: MODEL8389825246
.
To cite BioModels Database, please use: BioModels Database: An enhanced, curated and annotated resource
for published quantitative kinetic models
.
To the extent possible under law, all copyright and related or
neighbouring rights to this encoded model have been dedicated to the public
domain worldwide. Please refer to CC0 Public Domain
Dedication
for more information.
2024-09-02 | BIOMD0000000210 | BioModels
Project description:Chickarmane2008 - Stem cell lineage determination
In this work, a dynamical model of lineage
determination based upon a minimal circuit, as discussed in PMID: 17215298
, which contains the Oct4/Sox2/Nanog core as well its interaction
with a few other key genes is discussed.
This model is described in the article:
A computational model for understanding stem cell, trophectoderm and endoderm lineage determination.
Chickarmane V, Peterson C
PloS one. 2008, 3(10):e3478
Abstract:
BACKGROUND: Recent studies have associated the transcription factors, Oct4, Sox2 and Nanog as parts of a self-regulating network which is responsible for maintaining embryonic stem cell properties: self renewal and pluripotency. In addition, mutual antagonism between two of these and other master regulators have been shown to regulate lineage determination. In particular, an excess of Cdx2 over Oct4 determines the trophectoderm lineage whereas an excess of Gata-6 over Nanog determines differentiation into the endoderm lineage. Also, under/over-expression studies of the master regulator Oct4 have revealed that some self-renewal/pluripotency as well as differentiation genes are expressed in a biphasic manner with respect to the concentration of Oct4. METHODOLOGY/
PRINCIPAL FINDINGS: We construct a dynamical model of a minimalistic network, extracted from ChIP-on-chip and microarray data as well as literature studies. The model is based upon differential equations and makes two plausible assumptions; activation of Gata-6 by Oct4 and repression of Nanog by an Oct4-Gata-6 heterodimer. With these assumptions, the results of simulations successfully describe the biphasic behavior as well as lineage commitment. The model also predicts that reprogramming the network from a differentiated state, in particular the endoderm state, into a stem cell state, is best achieved by over-expressing Nanog, rather than by suppression of differentiation genes such as Gata-6.
CONCLUSIONS: The computational model provides a mechanistic understanding of how different lineages arise from the dynamics of the underlying regulatory network. It provides a framework to explore strategies of reprogramming a cell from a differentiated state to a stem cell state through directed perturbations. Such an approach is highly relevant to regenerative medicine since it allows for a rapid search over the host of possibilities for reprogramming to a stem cell state.
This model is hosted on BioModels Database
and identified
by: MODEL8390025091
.
To cite BioModels Database, please use: BioModels Database: An enhanced, curated and annotated resource
for published quantitative kinetic models
.
To the extent possible under law, all copyright and related or
neighbouring rights to this encoded model have been dedicated to the public
domain worldwide. Please refer to CC0 Public Domain
Dedication
for more information.
2024-09-02 | BIOMD0000000209 | BioModels
Project description:Chavez2009 - a core regulatory network of OCT4 in human embryonic stem cells
A core OCT4-regulated network has been identified as a test case, to analyase stem cell characteristics and cellular differentiation.
This model is described in the article:
In silico identification of a core regulatory network of OCT4 in human embryonic stem cells using an integrated approach.
Chavez L, Bais AS, Vingron M, Lehrach H, Adjaye J, Herwig R
BMC Genomics, 2009, 10:314
Abstract:
BACKGROUND: The transcription factor OCT4 is highly expressed in pluripotent embryonic stem cells which are derived from the inner cell mass of mammalian blastocysts. Pluripotency and self renewal are controlled by a transcription regulatory network governed by the transcription factors OCT4, SOX2 and NANOG. Recent studies on reprogramming somatic cells to induced pluripotent stem cells highlight OCT4 as a key regulator of pluripotency.
RESULTS: We have carried out an integrated analysis of high-throughput data (ChIP-on-chip and RNAi experiments along with promoter sequence analysis of putative target genes) and identified a core OCT4 regulatory network in human embryonic stem cells consisting of 33 target genes. Enrichment analysis with these target genes revealed that this integrative analysis increases the functional information content by factors of 1.3 - 4.7 compared to the individual studies. In order to identify potential regulatory co-factors of OCT4, we performed a de novo motif analysis. In addition to known validated OCT4 motifs we obtained binding sites similar to motifs recognized by further regulators of pluripotency and development; e.g. the heterodimer of the transcription factors C-MYC and MAX, a prerequisite for C-MYC transcriptional activity that leads to cell growth and proliferation.
CONCLUSION: Our analysis shows how heterogeneous functional information can be integrated in order to reconstruct gene regulatory networks. As a test case we identified a core OCT4-regulated network that is important for the analysis of stem cell characteristics and cellular differentiation. Functional information is largely enriched using different experimental results. The de novo motif discovery identified well-known regulators closely connected to the OCT4 network as well as potential new regulators of pluripotency and differentiation. These results provide the basis for further targeted functional studies.
This model is hosted on BioModels Database
and identified
by: MODEL1305010000
.
To cite BioModels Database, please use: BioModels Database: An enhanced, curated and annotated resource
for published quantitative kinetic models
.
To the extent possible under law, all copyright and related or
neighbouring rights to this encoded model have been dedicated to the public
domain worldwide. Please refer to CC0 Public Domain
Dedication
for more information.
2005-01-01 | MODEL1305010000 | BioModels
Project description:The generation of induced pluripotent stem cells (iPSCs) from differentiated cells following forced expression of Oct4, Klf4, Sox2 and c-Myc (OKSM) is slow and inefficient, suggesting that transcription factors have to overcome somatic barriers that resist cell fate change. Here, we performed an ubiased serial shRNA enrichment screen to identify novel repressors of somatic cell reprogramming into iPSCs. This effort uncovered the sumoylation effector protein Sumo2 as one of the strongest roadblocks to iPSC formation. Depletion of Sumo2 both enhances and accelerates reprogramming, yielding transgene-independent, chimera-competent iPSCs after as little as 36 hours of OKSM expression. We further show that the Sumo2 pathway acts independently of exogenous c-Myc expression and in parallel with small molecule enhancers of reprogramming. Critically, suppression of SUMO2 also promotes the generation of human iPSCs. Together, our results reveal sumoylation as a crucial post-transcriptional mechanism that resists the acquisition of pluripotency from fibroblasts using defined factors. Microarray analysis was performed during reprogramming or of iPSC lines derived upon Sumo2 knockdown Total RNA was isolated from day 6 reprogramming fibroblasts with or without Sumo2 knockdown; as well as stable iPSC clones derived from Sumo2 knockdown fibroblasts.
2016-03-03 | E-GEOD-76699 | biostudies-arrayexpress
Project description:The generation of induced pluripotent stem cells (iPSCs) from differentiated cells following forced expression of Oct4, Klf4, Sox2 and c-Myc (OKSM) is slow and inefficient, suggesting that transcription factors have to overcome somatic barriers that resist cell fate change. Here, we performed an ubiased serial shRNA enrichment screen to identify novel repressors of somatic cell reprogramming into iPSCs. This effort uncovered the sumoylation effector protein Sumo2 as one of the strongest roadblocks to iPSC formation. Depletion of Sumo2 both enhances and accelerates reprogramming, yielding transgene-independent, chimera-competent iPSCs after as little as 36 hours of OKSM expression. We further show that the Sumo2 pathway acts independently of exogenous c-Myc expression and in parallel with small molecule enhancers of reprogramming. Critically, suppression of SUMO2 also promotes the generation of human iPSCs. Together, our results reveal sumoylation as a crucial post-transcriptional mechanism that resists the acquisition of pluripotency from fibroblasts using defined factors. Microarray analysis was performed during reprogramming or of iPSC lines derived upon Sumo2 knockdown
2016-03-03 | GSE76699 | GEO
Project description:The reprogramming of somatic cells into induced pluripotent stem cells (iPSCs) upon overexpression of OCT4, KLF4, SOX2 and c-MYC (OKSM) provides a powerful system to interrogate basic mechanisms of cell fate change. However, iPSC formation with standard methods is typically protracted and inefficient, resulting in heterogeneous cell populations. We show that exposure of OKSM-expressing cells to both ascorbic acid and a GSK3-β inhibitor (AGi) facilitates more synchronous and rapid iPSC formation from several mouse cell types. AGi treatment restored the ability of refractory cell populations to yield iPSC colonies, and it attenuated the activation of developmental regulators commonly observed during the reprogramming process. Moreover, AGi supplementation gave rise to chimera-competent iPSCs after as little as 48 h of OKSM expression. Our results offer a simple modification to the reprogramming protocol, facilitating iPSC induction at unparalleled efficiencies and enabling dissection of the underlying mechanisms in more homogeneous cell populations. 18 samples were analyzed in total, samples represent bulk cultures of reprogrammable-MEFs (Rep-MEFs) that express OKSM and were supplemented with ascorbic acid and GSK3i during iPS cell induction. Control samples represent similar bulk cultures of reprogrammable-MEFs that express OKSM but were not supplemented with ascorbic acid and GSK3i during iPS cell induction. GMP-iPSCs were generated using 48 hours of OKSM+AGi induction. Fibroblast-iPSCs were generated using 96 hours of OKSM+AGi induction.
2014-09-23 | E-GEOD-57774 | biostudies-arrayexpress
Project description:Recent reports have proposed a new paradigm for obtaining mature somatic cell types from fibroblasts without going through a pluripotent state, by briefly expressing canonical iPSC reprogramming factors Oct4, Sox2, Klf4 and c-Myc (abbreviated as OSKM), in cells expanded in lineage differentiation promoting conditions. Here we apply genetic lineage tracing for endogenous Nanog, Oct4 and X chromosome reactivation during OSKM induced trans-differentiation, as these molecular events mark final stages for acquisition of induced pluripotency. Remarkably, the vast majority of reprogrammed cardiomyocytes or neural stem cells derived from mouse fibroblasts via OSKM mediated trans-differentiation were attained after transient acquisition of pluripotency, and followed by rapid differentiation. Our findings underscore a molecular and functional coupling between inducing pluripotency and obtaining “trans-differentiated” somatic cells via OSKM induction, and have implications on defining molecular trajectories assumed during different cell reprogramming methods. poly RNA-Seq and Chromatin accesibility (ATAC-seq) were measured during conversion of mouse embryonic fibroblasts to neural stem cells using OSKM trans-differentiation method, as well as in mouse emrbyonic fibroblasts, iPSCs and mouse ESCs.
2015-04-30 | E-GEOD-67298 | biostudies-arrayexpress