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| Status |
Public on Dec 08, 2010 |
| Title |
Direct Reprogramming of Fibroblasts into Epiblast Stem Cells |
| Organism |
Mus musculus |
| Experiment type |
Expression profiling by array
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| Summary |
Embryonic stem cells (ESCs), which are derived from the primitive ectoderm of pre-implantation blastocysts, are pluripotent cells and can thus contribute to the formation of all somatic cell lineages in chimeric animals. Similarly, epiblast stem cells (EpiSCs), which are derived from epiblast tissue of post-implantation embryos, are also pluripotent and can give rise to derivatives of all three germ layers in teratoma assays. Introduction of the four transcription factors Oct4/Sox2/Klf4/c-Myc into somatic cells has been shown to generate induced pluripotent stem cells (iPSCs). iPSCs exhibit ESC-like properties and are virtually identical to ESCs with respect to a number of charactertistics. However, generation of EpiSC-like cells by direct reprogramming of somatic cells using these transcription factors has not been shown to date. Here we show that Yamanaka’s four transcription factors can be used to directly generate EpiSC-like iPS cells (ePSCs) under EpiSC culture conditions. These ePSCs are identical to EpiSCs with respect to morphology, gene expression pattern, epigenetic status, and chimera formation. This is the first study to demonstrate that the culture environment in transcription factor-mediated reprogramming determines the cell fate of the reprogrammed cell. We could therefore envision that we eventually are able to shape the identity of a directly reprogrammed cell at will simply by modulating the culture conditions.
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| Overall design |
RNA samples to be analyzed on microarrays were prepared using Qiagen RNeasy columns with on-column DNA digestion. 500 ng of total RNA per sample was used as input into a linear amplification protocol (Ambion), which involved synthesis of T7-linked double-stranded cDNA and 12 hrs of in-vitro transcription incorporating biotin-labelled nucleotides. Purified and labelled cRNA was then hybridized for 18 hrs onto MouseRef-8 v2 expression BeadChips (Illumina) according to the manufacturer's instructions. After washing, as recommended, chips were stained with streptavidin-Cy3 (GE Healthcare) and scanned using the iScan reader (Illumina) and accompanying software. Samples were hybridized as biological replicates.
24 samples were analyzed:
MEF: Female mouse embryonic fibroblast, 2 biological rep
ESC OG2: OG2 female embryonic stem cell, 2 biological rep
EpiSC OG2: OG2 female EpiSC grown as single cells with conditioned medium, 2 biological rep
EpiSC GOF18: GOF18 EpiSC grown as single cells with conditioned medium, 2 biological rep
EpiSC T9b: T9 EpiSC on MEFs in activin-containing CDM, harvested w/o feeders, subconfluent, 2 biological rep
ePSC L4: EpiSC-like iPS Cells L4, MEF reprogrammed to EpiSCs with 4F, line L4, grown with LIF and activin, 2 biological rep
ePSC L7: EpiSC-like iPS cells L7, MEF reprogrammed to EpiSCs with 4F, line L7, grown with LIF and activin, 2 biological rep
ePSC C1: EpiSC-like iPS Cells C1, MEF reprogrammed to EpiSCs with 4F, line C1, grown with LIF and activin, 2 biological rep
ePSC C3: EpiSC-like iPS Cells C3, MEF reprogrammed to EpiSCs with 4F, line C3, grown with LIF and activin, 2 biological rep
ePSC-R: EpiSC-like iPS Cells reverted, L4 ePSC cells reverted to an ESC-like state using Klf4 virus, 2 biological rep
ePSC-RC1: EpiSC-like iPS Cells reverted, L4 ePSR Cre1, Cre-treated L4 ePSR cells, line 1, 2 biological rep
ePSC-RC2: EpiSC-like iPS Cells reverted, L4 ePSR Cre2, Cre-treated L4 ePSR cells, line 2, 2 biological rep
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| Contributor(s) |
Han DW, Greber B, Wu G, Tapia N, Araúzo-Bravo MJ, Ko K, Stehling M, Schöler HR |
| Citation(s) |
21131959 |
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| Submission date |
Apr 26, 2010 |
| Last update date |
Jan 15, 2022 |
| Contact name |
Marcos J. Araúzo-Bravo |
| E-mail(s) |
[email protected]
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| Phone |
+34 943 00 6108
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| Organization name |
Max Planck Institute for Molecular Biomedicine
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| Department |
Cell and Developmental Biology
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| Lab |
Computational Biology and Bionformatics
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| Street address |
Rogentstrasse
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| City |
Muenster |
| ZIP/Postal code |
48149 |
| Country |
Germany |
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| Platforms (1) |
| GPL6885 |
Illumina MouseRef-8 v2.0 expression beadchip |
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| Samples (24)
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| GSM537498 |
ESC OG2, rep2 |
| GSM537499 |
EpiSC OG2, rep1 |
| GSM537500 |
EpiSC OG2, rep2 |
| GSM537501 |
EpiSC GOF18, rep1 |
| GSM537502 |
EpiSC GOF18, rep2 |
| GSM537503 |
EpiSC T9b, rep1 |
| GSM537504 |
EpiSC T9b, rep2 |
| GSM537505 |
ePSC L4, rep1 |
| GSM537506 |
ePSC L4, rep2 |
| GSM537507 |
ePSC L7, rep1 |
| GSM537508 |
ePSC L7, rep2 |
| GSM537509 |
ePSC C1, rep1 |
| GSM537510 |
ePSC C1, rep2 |
| GSM537511 |
ePSC C3, rep1 |
| GSM537512 |
ePSC C3, rep2 |
| GSM537513 |
ePSC-R, rep1 |
| GSM537514 |
ePSC-R, rep2 |
| GSM537515 |
ePSC-RC1, rep1 |
| GSM537516 |
ePSC-RC1, rep2 |
| GSM537517 |
ePSC-RC2, rep1 |
| GSM537518 |
ePSC-RC2, rep2 |
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| Relations |
| BioProject |
PRJNA125975 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE21516_RAW.tar |
3.1 Mb |
(http)(custom) |
TAR |
| GSE21516_non-normalized.txt.gz |
4.0 Mb |
(ftp)(http) |
TXT |
| Processed data included within Sample table |
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