Transcription profiling of mouse embryonic stem cell lines CGR8, E14TG2a and R1 to identify differences in expression
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ABSTRACT:
ORGANISM(S): Mus musculus
SUBMITTER: Oliver Hummel
PROVIDER: E-TABM-671 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:This experiment is part of the FunGenES project (FunGenES - Functional Genomics in Embryonic Stem Cells partially funded by the 6th Framework Programme of the European Union, http://www.fungenes.org). The experiment was conducted at the Inst. National de la Santé et de la Recherche Médicale - INS (INSERM U371, Bron, France) The goal of the experiment is to identify genes regulated by STAT3 and NANOG, which are known inhibitors of mouse ES cell differentiation.(I) E14-Cre-CTL cells deprived of LIF for 24 hours (n = 5); (II) E14-Cre-Tax cells as in (I) and restimulated with 100 nM tamoxifen for 2 hours (n = 5); (III) E14-STATA3-CTL cells deprived of LIF and 4âOHT for 24 hours (n = 4); (IV) E14-STAT3-LIF cells treated as in (III) and re-stimulated with 10,000 U/ml LIF for 1 hour to reactivate endogenous STAT3 (n = 5); (V) E14-STAT3-Tax cells as in (III) and restimulated with 100 nM 4âOHT for 2 hours to activate STAT3-ERT2 (n = 6);
2009-03-01 | E-TABM-562 | biostudies-arrayexpress
Project description:This experiment is part of the FunGenES project (FunGenES - Functional Genomics in Embryonic Stem Cells partially funded by the 6th Framework Programme of the European Union, http://www.fungenes.org). The experiment was conducted at the Institute for Molecular Biology and Biotechnology FORTH, Herakleion, Krete, Greece. Goal mouse ES cells grown in presence of LIF and treated with TSA (HDAC inhibitor). Materials and methods: CGR8 cells were treated by Trichostatin A (TSA) in presence of LIF. TSA doses ranging from 10 to 50nM were tested for time periods of 6, 12 and 24 hours.
2009-07-10 | E-TABM-670 | biostudies-arrayexpress
Project description:This experiment is part of the FunGenES project (FunGenES - Functional Genomics in Embryonic Stem Cells partially funded by the 6th Framework Programme of the European Union, http://www.fungenes.org). The experiment was conducted at University of Cologne, Cologne, Germany. Goal of the experiment was RNA expression profiling in the first 10 days of differentiation in CGR8 cells. Materials and methods: CGR8 cells were cultivated as EBs without addition of selective reagents in hanging drop cultures in a timecourse experiment. After 7 days, EBs were plated in gelatin-coated 6-well plates. RNA was prepared from ES cells (day 0), EBs (day 1 to 7) and plated EBs (day 10). Culture conditions: ES cells were differentiated via hanging drop protocol into embryoid bodies. RNA isolation method: The RNA isolation was carried out following standard methods (on column, RNeasy Mini kit, Qiagen)
2009-07-10 | E-TABM-672 | biostudies-arrayexpress
Project description:This experiment is part of the FunGenES project (FunGenES - Functional Genomics in Embryonic Stem Cells partially funded by the 6th Framework Programme of the European Union, http://www.fungenes.org). The experiment was conducted at the University Bordeaux 2 (CNRS UMR 5164). The purpose of the experiment was to identify the LIF-dependent genes and early differentiation markers in mouse ES cells. Leukemia Inhibitory Factor (LIF) is a member of the Interleukin-6 (IL-6) family of cytokines, which displays pleiotropic functions, depending on both cell maturity and cell type. LIF maintains ES cell pluripotency and is a critical cell survival factor in myocytes. Materials and methods: CGR8 cell line was used.6 cell conditions tested: + LIF, reinduced 25 min with serum + LIF reinduced 25 min with serum and LIF 500pM - LIF for 24h, reinduced 25 min with serum - LIF for 24h, reinduced 25 min with serum and LIF 500pM- LIF for 48h, reinduced 25 min with serum - LIF for 48h, reinduced 25 min with serum and LIF 500pM. Relationships between samples. 5 independent preparations of mRNA have been done on five different CGR8 passages. Culture conditions: Monolayer adherent cells. RNA isolation method: Qiagen column
2009-07-10 | E-TABM-667 | biostudies-arrayexpress
Project description:This experiment is part of the FunGenES project (FunGenES - Functional Genomics in Embryonic Stem Cells partially funded by the 6th Framework Programme of the European Union, http://www.fungenes.org). The experiment was conducted at Inserm U846, Bron, France. Aim: Kru_ppel-like factors (Klf) 4 and 5 are two closely related members of the Klf family, known to play key roles in somatic cell reprogramming and in self-renewal of pluripotent stem cells. In this study, we focused on the functional divergence between Klf4 and Klf5. We showed that Klf4 and Klf5 regulate the expression of distinct subsets of genes. Klf4 negatively regulates the expression of endodermal markers, some of which encode transcription factors involved in the commitment of pluripotent system cells to endoderm differentiation. In contrast, Klf5 negatively regulates the expression of mesodermal markers, some of which controls commitment to the mesoderm lineage. Functional studies with reporter cell lines indicate that knockdown of Klf4 enhances differentiation toward visceral endoderm, mesendoderm, and definitive endoderm, whereas knockdown of Klf5 specifically enhances differentiation toward mesoderm. Thus, additive functions of Klf4 and Klf5 secure pluripotent stem cell propagation by inhibiting endoderm and mesoderm differentiation.
2014-04-28 | E-MTAB-1838 | biostudies-arrayexpress
Project description:This experiment is a follow-up experiment of the FunGenES project (FunGenES - Functional Genomics in Embryonic Stem Cells partially funded by the 6th Framework Programme of the European Union, http://www.fungenes.org). The experiment was conducted at Inserm U846, Bron, France. Aim: Kru_ppel-like factors (Klf) 4 and 5 are two closely related members of the Klf family, known to play key roles in somatic cell reprogramming and in self-renewal of pluripotent stem cells. In this study, we focused on the functional divergence between Klf4 and Klf5. We showed that Klf4 and Klf5 regulate the expression of distinct subsets of genes. Klf4 negatively regulates the expression of endodermal markers, some of which encode transcription factors involved in the commitment of pluripotent system cells to endoderm differentiation. In contrast, Klf5 negatively regulates the expression of mesodermal markers, some of which controls commitment to the mesoderm lineage. Functional studies with reporter cell lines indicate that knockdown of Klf4 enhances differentiation toward visceral endoderm, mesendoderm, and definitive endoderm, whereas knockdown of Klf5 specifically enhances differentiation toward mesoderm. Thus, additive functions of Klf4 and Klf5 secure pluripotent stem cell propagation by inhibiting endoderm and mesoderm differentiation.
2014-04-28 | E-MTAB-1839 | biostudies-arrayexpress
Project description:This experiment is part of the FunGenES project (FunGenES - Functional Genomics in Embryonic Stem Cells partially funded by the 6th Framework Programme of the European Union, http://www.fungenes.org). The experiment was conducted at University of Cologne, Cologne, Germany. Goal of the experiment was an analysis the transcriptome of the ES-cell derived pure T-brachyury expressing CGR8 cells. Materials and methods: On day 0, EBs were made from T-brachyury ES (CGR8) cell line by hanging drop protocol. On second day, the hanging drop EBs were transferred into suspension. On third day, the EBs were treated or untreated with puromycin at a concentration of 5ug/ml for 2 days. On 5th day, the medium was changed and added medium with or without puromycin at a concentration of 2 ug/ml for 1 day. On day 6th , RNA was isolated from EBs treated or untreated with puromycin for affymetrix analysis.
2010-05-12 | E-TABM-716 | biostudies-arrayexpress
Project description:This experiment is part of the FunGenES project (FunGenES - "Functional Genomics in Embryonic Stem Cells" partially funded by the 6th Framework Programme of the European Union, http://www.fungenes.org). The experiment was conducted at the Leibniz Institute of Plant Genetics and Crop Plant Research (IPK, Gatersleben, Germany) The goal of the experiment is to optimize existing differentiation protocols for endoderm differentiation of ES cells to efficiently generate optimal levels of pancreatic cell types. To characterize potential pancreatic progenitors during the process of induced pancreatic differentiation, wild type R1 cells and R1 cells stably expressing the pancreatic control gene Pax4, RNA samples of undifferentiated cells, pancreatic precursor and differentiated cells were prepared.
2009-03-01 | E-TABM-493 | biostudies-arrayexpress
Project description:This experiment is part of the FunGenES project (FunGenES - Functional Genomics in Embryonic Stem Cells partially funded by the 6th Framework Programme of the European Union, http://www.fungenes.org).The experiment was conducted at the Unyversity of Bath, Bath, UK. Aim of the experiment is the identification of genes regulated by PI3K-dependent signaling in undifferentiated ES cells. Materials and methods: To identify target genes of PI3-K signalling, the PI3-K signalling inhibitor LY294002 was used at a concentration which was shown to perturb self-renewal in the presence of LIF and which knocks down PI3K signals. CGR8 cells cultivated in serum-free conditions in the presence of LIF were starved for 24h in media lacking LIF and then pre-inubated for 30 minutes with 5 mM LY294002 in DMSO or with DMSO alone. Cells were then stimulated with LIF for 2h, prior to preparation of RNA samples. Relationships between samples: Samples were either maintained in LIF or LIF plus LY294002 (PI3K inhibitor) for the required treatment times. Treatments: Treatment times with PI3K inhibitor LY294002 at 5 uM 2h (n=5), 3h (n=5).
2009-05-07 | E-TABM-673 | biostudies-arrayexpress
Project description:This experiment is part of the FunGenES project (FunGenES - Functional Genomics in Embryonic Stem Cells partially funded by the 6th Framework Programme of the European Union, http://www.fungenes.org). The experiment was conducted at the University Nice Sophia-Antipolis, Nice, France. The aim of this project is to identify new regulators, and new markers of the early steps of adipocyte commitment.Addition of RA on early-differentiating mES cells (Hanging drops, Day 3) orientates their commitment to the adipocyte lineage, while inhibiting myogenesis. The effect of RA on adipogenesis is mediated by retinoic acid receptor beta (RARb), since it can be mimiked by the addition of CD2314 compound, a specific agonist of this receptor isotype. In contrast, mES cell adipogenesis can be bloked by artificial activation of the Wnt pathway using GSK3 inhibitors such as lithium, or Bio.To decipher the genetic pathways that control the early steps of adipogenesis, we have compared the transcriptome of mES cells stimulated to differentiate using adipogenesis-enhancing (CD2314) or -inhibitory (BIO, or CD2314+BIO) treatments (treatment between day 3 and day 6). We have analysed the transcriptome of these cells at day 3 (before treatment), at day 6 (just after stimulation or inhibition of the early steps of adipocyte commitment) and at day 11 (to investigate later regulators of adipogenesis). Materials and methods: CGR8 at different time point during adipogenic differentiation. - Induction of differentiation by removal of LIF at day 0, hanging drops from day 0 to 3. - At day 3, harvesting of the hanging drops, and start adipogenesis-enhancing (CD2314) or -inhibitory (BIO, or CD2314+BIO) treatments (treatment from day 3 to 6, EBs in suspension on petri dishes).- At day 6, harvesting of the EBs and plating on gelatin-treated TC plates in normal culture medium ( GMEM+10%serum).- At Day 7, treatment with final inducers of adipogenic differentiation (Insuline, T3, BRL). Same medium until end point of differentiation (Day 21). Relationships between samples: day03= CGR8, day 3 of differentiation, hanging drops from day 0 (removal of LIF) and 3. day06_0= day 6 of differentiation, control condition from day 3 to 6 (GMEM+serum)day06_bio= day 6 of differentiation, treatment with Bio from day 3 to day 6. day06_cd= day 6 of differentiation,treatment with CD2314 from day 3 to day 6.day06_cd_bio= day 6 of differentiation,treatment with CD2314 plus Bio from day 3 to day 6.Day11_0= day 11 of differentiation, cells in control condition from day 3 to 6 (GMEM+serum).Day11_bio= day 11 of differentiation, cells treated with Bio from day 3 to day 6.Day11_cd= day 11 of differentiation,cells treated with CD2314 from day 3 to day 6.Day11_cd_bio= day 11 of differentiation,cells treated with CD2314 plus Bio from day 3 to day 6.Treatments: - At day 3, addition of adipogenesis-enhancing (CD2314) or -inhibitory (BIO, or CD2314+BIO) treatments (treatment from day 3 to 6).- At Day 7, treatment with final inducers of adipogenic differentiation (Insuline, T3, BRL). O= No treatment CD= treatment w Culture conditions; - Hanging drops from day 0 to Day 3.- Suspension culture between day 3 to 6, Petri dishes.- Plating on gelatine from day 6 to end point.
2009-07-10 | E-TABM-668 | biostudies-arrayexpress