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OCT4 and SOX2 Work as Transcriptional Activators in Reprogramming Human Fibroblasts

ABSTRACT: SOX2 and OCT4, in conjunction with KLF4 and cMYC, are sufficient to reprogram human fibroblasts to induced pluripotent stem cells (iPSCs), but it is unclear if they function as transcriptional activators or as repressors. We now show that, like OCT4, SOX2 functions as a transcriptional activator. We substituted SOX2-VP16 (a strong activator) for wild-type (WT) SOX2, and we saw an increase in the efficiency and rate of reprogramming, whereas the SOX2-HP1 fusion (a strong repressor) eliminated reprogramming. We report that, at an early stage of reprogramming, virtually all DNA-bound OCT4, SOX2, and SOX2-VP16 were embedded in putative enhancers, about half of which were created de novo. Those associated with SOX2-VP16 were, on average, stronger than those bearing WT SOX2. Many newly created putative enhancers were transient, and many transcription factor locations on DNA changed as reprogramming progressed. These results are consistent with the idea that, during reprogramming, there is an intermediate state that is distinct from both parental cells and iPSCs.

ORGANISM(S): Homo sapiens

PROVIDER: GSE81900 | GEO | 2017/08/30

SECONDARY ACCESSION(S): PRJNA324088

REPOSITORIES: GEO

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OCT4 and SOX2 Work as Transcriptional Activators in Reprogramming Human Fibroblasts

Project description:SOX2 and OCT4, in conjunction with KLF4 and cMYC, are sufficient to reprogram human fibroblasts to induced pluripotent stem cells (iPSCs), but it is unclear if they function as transcriptional activators or as repressors. We now show that, like OCT4, SOX2 functions as a transcriptional activator. We substituted SOX2-VP16 (a strong activator) for wild-type (WT) SOX2, and we saw an increase in the efficiency and rate of reprogramming, whereas the SOX2-HP1 fusion (a strong repressor) eliminated reprogramming. We report that, at an early stage of reprogramming, virtually all DNA-bound OCT4, SOX2, and SOX2-VP16 were embedded in putative enhancers, about half of which were created de novo. Those associated with SOX2-VP16 were, on average, stronger than those bearing WT SOX2. Many newly created putative enhancers were transient, and many transcription factor locations on DNA changed as reprogramming progressed. These results are consistent with the idea that, during reprogramming, there is an intermediate state that is distinct from both parental cells and iPSCs

2017-08-30 | GSE81899 | GEO

OCT4 and SOX2 Work as Transcriptional Activators in Reprogramming Human Fibroblasts

Project description:SOX2 and OCT4, in conjunction with KLF4 and cMYC, are sufficient to reprogram human fibroblasts to induced pluripotent stem cells (iPSCs), but it is unclear if they function as transcriptional activators or as repressors. We now show that, like OCT4, SOX2 functions as a transcriptional activator. We substituted SOX2-VP16 (a strong activator) for wild-type (WT) SOX2, and we saw an increase in the efficiency and rate of reprogramming, whereas the SOX2-HP1 fusion (a strong repressor) eliminated reprogramming. We report that, at an early stage of reprogramming, virtually all DNA-bound OCT4, SOX2, and SOX2-VP16 were embedded in puta- tive enhancers, about half of which were created de novo. Those associated with SOX2-VP16 were, on average, stronger than those bearing WT SOX2. Many newly created putative enhancers were tran- sient, and many transcription factor locations on DNA changed as reprogramming progressed. These results are consistent with the idea that, during re- programming, there is an intermediate state that is distinct from both parental cells and iPSCs.

2017-08-30 | GSE81891 | GEO

MEF_OSK_O_KS_T20

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

Excluding Oct4 from Yamanaka cocktail unleashes the developmental potential of iPSCs

Project description:Oct4 is widely considered the most important among the four Yamanaka reprogramming factors. Here we show that the combination of Sox2, Klf4, and cMyc (SKM) suffices for reprogramming mouse somatic cells to induced pluripotent stem cells (iPSCs). Simultaneous induction of Sox2 and cMyc in fibroblasts triggers immediate retroviral silencing, which explains the discrepancy with previous studies that attempted but failed to generate iPSCs without Oct4 using retroviral vectors. SKM induction could partially activate the pluripotency network even in Oct4-knockout fibroblasts. Importantly, reprogramming in the absence of exogenous Oct4 results in greatly improved developmental potential of iPSCs, determined by their ability to give rise to all-iPSC mice in the tetraploid complementation assay. Our data suggest that overexpression of Oct4 during reprogramming leads to off-target gene activation during reprogramming and epigenetic aberrations in resulting iPSCs, and thereby bear major implications for further development and application of iPSC technology.

2019-11-06 | GSE137001 | GEO

Proteomic analysis of mouse oocytes identifies PRMT7 as reprogramming factor that replaces SOX2 in the induction of pluripotent stem cells

Project description:The inclusion of oocyte factors together with Yamanaka’s previously identified reprogramming factors (OCT4, SOX2, KLF4 with or without cMYC; OSK(M)) may facilitate the reprogramming process that leads to induced pluripotent stem cells (iPSCs). We previously applied label-free LC-MS/MS analysis to search for such facilitators of reprogramming (reprogrammome), resulting in a catalog of 28 candidates that are (i) able to robustly access the cell nucleus, and (ii) shared between mature mouse oocytes and pluripotent embryonic stem (ES) cells. In the present study we hypothesized that our 28 reprogrammome candidates would also be (iii) abundant in mature mouse oocytes, (iv) depleted after oocyte-to-embryo transition, and (v) able to potentiate or replace the OSKM factors during iPSC reprogramming. Using LC-MS/MS and isotopic labeling methods we found that the abundance profiles of the 28 proteins was below that of known oocyte-specific and housekeeping proteins. Out of the 28 proteins only arginine methyltransferase 7 (PRMT7) presented a substantially changing profile during mouse embryogenesis and impacted on the conversion of mouse fibroblasts into iPSCs. PRMT7 indeed could very efficiently replace SOX2 in a factor-substitution assay yielding iPSCs. These findings show that proteomics can be used to prioritize the functional analysis of reprogrammome candidates.

2016-05-31 | PXD003093 | Pride

Transcriptional analysis of iPSCs during cellular reprogramming using RNA FACS

Project description:Mouse induced pluripotent stem cells (iPSCs) were derived from embryonic fibroblasts by overexpressing the Yamanaka factors Oct4, Sox2, Klf4 and c-Myc for 32 consecutive days. iPSCs were isolated by RNA FACS (endogenous Sox2) from the heterogeneous reprogramming culture and transcriptionally compared with (1) iPSCs stabilized in 2i medium (iPS-2i) and (2) iPSCs stabilized in co-culture with mouse embryonic fibroblasts (iPS-MEF) n = 2 replicates for each sample/condition

2014-03-20 | E-GEOD-55671 | biostudies-arrayexpress

Transcriptional analysis of iPSCs during cellular reprogramming using RNA FACS

Project description:Mouse induced pluripotent stem cells (iPSCs) were derived from embryonic fibroblasts by overexpressing the Yamanaka factors Oct4, Sox2, Klf4 and c-Myc for 32 consecutive days. iPSCs were isolated by RNA FACS (for endogenous Sox2) from a heterogeneous reprogramming culture and transcriptionally compared with (1) iPSCs stabilized in 2i medium (iPS-2i) and (2) iPSCs stabilized in co-culture with mouse embryonic fibroblasts (iPS-MEF)

2014-03-20 | GSE55671 | GEO

Induction of pluripotency in mouse somatic cells with novel factors (expression)

Project description:For years, reprogramming factors that induce pluripotency have been identified primarily from ESC-enriched factors, pluripotency-associated factors such as Oct4, Sox2, Klf4, c-Myc, Nanog, Esrrb, Sall4, Utf1, Lin28, PRDM14, and Tet2. Through genome-wide screen we identified novel regulators of reprogramming. Our study is the first proof-of-principle report to suggest a putative general mathematical model. This model provides novel mechanistic insight into the fundamental understanding of the establishment of cellular identity during programming and reprogramming. Through genome-wide screen to identify novel factors of reprogramming in addition to Oct4, Sox2, Klf4, cMyc Through genome-wide screen to identify novel factors of reprogramming in addition to Oct4, Sox2, Klf4, cMyc. Induced pluripotent stem cells versus mouse embryonic fibroblast.

2013-05-28 | E-GEOD-47441 | biostudies-arrayexpress

Nanog is Dispensable for the Generation of Induced Pluripotent Stem Cells

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

MEF_OSK_O_KS_T20

2015-09-17 | GSE61694 | GEO

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