Dissecting pluripotency through the different OCT4 and SOX2 assembly configurations
Ontology highlight
ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE65345 | GEO | 2017/02/13
SECONDARY ACCESSION(S): PRJNA273740
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: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.
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2005-01-01 | MODEL1305010000 | BioModels
Project description:Induced pluripotent stem cells (iPSCs) can be derived from somatic cells by the introduction of the transcription factors Oct4, Sox2, Klf4 and cMyc using various methods. Here, we describe a new approach for the derivation of murine iPSCs using a polycistronic non-viral inducible vector integrated into pseudo attP sites via the C31 integrase-mediated site-specific recombination and subsequent vector excision by Cre recombinase. The pluripotency of the derived iPSCs was proved by in vitro and in vivo tests. The derived transgene-free iPSCs reactivated the endogenous pluripotency genes like e.g. Oct4, Sox2 and Nanog and the global gene expression profiles of iPSCs lines are highly similar to ESCs and distinct from parental murine fibroblasts. We demonstrated the differentiation potential of iPSCs by generation cells of the three germ layers as well as we successfully created germline chimeric mice from transgene-free iPSCs. In this study, we presented an efficient method for the generation of transgene-free iPSCs using dual-recombinase technology. expression data of iPSCs/ESCs/MEFs
2014-03-29 | E-GEOD-51605 | biostudies-arrayexpress
Project description:In this study, we set out to identify those molecular features of the POU transcription factor Oct4 that are responsible for inducing pluripotency in somatic cells. Oct4 is known to have a strong preference to cooperate with Sox2 on heterodimeric SoxOct elements predominantly found in enhancers of genes expressed in embryonic stem cells (ESCs). To test whether this partnership is specific to Oct4, we compared its DNA recognition and reprogramming activities to the paralogous transcription factor Oct6, which cannot induce and maintain pluripotency in mouse cells. By analyzing ChIP-Seq data and performing quantitative dimerization assays, we found that in somatic cells, instead of heterodimerzing with Sox-factors, Oct6 more potently homodimerizes on OctOct elements. We identified that a single amino acid is crucial in directing binding to the respective composite DNA element. As a consequence, just changing this one amino acid hampers Oct4 in generating induced pluripotent stem cells (iPSCs). In contrast, the reverse mutation in Oct6 did not augment its reprogramming activity. This was achieved with at least two additional exchanges. In summary, we demonstrate that cell-type specific POU factor function is determined by a limited set of residues that affect DNA and partner factor interactions. Such relatively minor changes lead to a pronounced impact on regulatory function and reprogramming activity.
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Project description:Next Generation Sequencing with differdent pluripotent transcript factor overexpression in MEF Somatic cells can be reprogrammed into induced pluripotent stem cells (iPSCs) by defined factors. The low efficiency of reprogramming limited the potential application of iPSCs. Here we found that knockdown LSD1 , which demethylates histone H3 Lys 4 or 9 , could increase iPSCs generation. It has been reported that LSD1 interaction with Oct4 which is the core factor of reprogramming. So we try to find out the different of overexpression Oct4 or Kllf4/Sox2 in MEF and LSD1 inhibitor. Retrovirus-mediated different pluripotent transcript factor overexpression in MEF cells after 4 days collect mRNA profiles processing with Illumina MiSeq; MEF, Mouse Embryonic Fibroblast O, MEF infected with Oct4 KS,MEF infected with Klf4 and Sox2 OSK, MEF infected with Oct4 Klf4 and Sox2 T20, MEF infected with Oct4 Klf4 and Sox2 and treated with 20nm LSD1 inhibitor Tranylcypromine
2015-09-17 | E-GEOD-61694 | biostudies-arrayexpress
Project description:Our understanding of pluripotency remains limited: iPSC generation has only been established for a few model species, pluripotent stem cell lines exhibit inconsistent developmental potential, and germline transmission demonstrated only for mice and rats. By swapping structural elements between Sox2 and Sox17, we built a chimeric super-SOX factor, Sox2-17, that enhanced iPSC generation in five tested species: mouse, human, cynomolgus monkey, cow, and pig. A swap of alanine to valine at the interface between Sox2 and Oct4 delivered a remarkable gain-of-function by stabilizing Sox2/Oct4 dimerization on DNA, enabling generation of high-grade OSKM iPSCs capable of supporting the development of healthy all-iPSC mice. Sox2/Oct4 dimerization emerged as the core driver of naïve pluripotency with its levels diminished upon priming. Transient overexpression of Sox2-17 and Klf4 (S*K cocktail) restored the dimerization and boosted the developmental potential of pluripotent stem cells across species, providing a universal method for naïve reset in mammals.
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Project description:Our understanding of pluripotency remains limited: iPSC generation has only been established for a few model species, pluripotent stem cell lines exhibit inconsistent developmental potential, and germline transmission demonstrated only for mice and rats. By swapping structural elements between Sox2 and Sox17, we built a chimeric super-SOX factor, Sox2-17, that enhanced iPSC generation in five tested species: mouse, human, cynomolgus monkey, cow, and pig. A swap of alanine to valine at the interface between Sox2 and Oct4 delivered a remarkable gain-of-function by stabilizing Sox2/Oct4 dimerization on DNA, enabling generation of high-grade OSKM iPSCs capable of supporting the development of healthy all-iPSC mice. Sox2/Oct4 dimerization emerged as the core driver of naïve pluripotency with its levels diminished upon priming. Transient overexpression of Sox2-17 and Klf4 (S*K cocktail) restored the dimerization and boosted the developmental potential of pluripotent stem cells across species, providing a universal method for naïve reset in mammals.
2023-11-27 | GSE247049 | GEO
Project description:Our understanding of pluripotency remains limited: iPSC generation has only been established for a few model species, pluripotent stem cell lines exhibit inconsistent developmental potential, and germline transmission demonstrated only for mice and rats. By swapping structural elements between Sox2 and Sox17, we built a chimeric super-SOX factor, Sox2-17, that enhanced iPSC generation in five tested species: mouse, human, cynomolgus monkey, cow, and pig. A swap of alanine to valine at the interface between Sox2 and Oct4 delivered a remarkable gain-of-function by stabilizing Sox2/Oct4 dimerization on DNA, enabling generation of high-grade OSKM iPSCs capable of supporting the development of healthy all-iPSC mice. Sox2/Oct4 dimerization emerged as the core driver of naïve pluripotency with its levels diminished upon priming. Transient overexpression of Sox2-17 and Klf4 (S*K cocktail) restored the dimerization and boosted the developmental potential of pluripotent stem cells across species, providing a universal method for naïve reset in mammals.
2023-11-27 | GSE247048 | GEO
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.
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