RNA-seq analysis of wild-type and csrp1KO mESCs and embryoid bodies of day 9
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE226212 | GEO | 2024/08/23
REPOSITORIES: GEO
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Project description:Mouse embryonic stem cells can spontaneously differentiate and assemble into a spherical embryoid body (EB) during suspension culture. The initial study aims to identify the up-regulated or down-regulated microRNAs during the differentiation process of pluripotent stem cells. From the microRNA profiling, we will focus on the microRNAs associated with mesodermal and endothelial differentiation of embryonic or induced pluripotent stem cells. In this study, the EBs were collected at 0 and 10-day differentiation of ESCs (ATCC: CRL-1934). The miRNA transcripts listed in Sanger miRBase Release 19.0 was detected by microarray.
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Project description:This experiment was specifically designed to measure neural targets of Shh signaling, we sought to profile the genes upregulated by Hh signaling in the ventral neural tube to obtain a valid dataset. To obtain ventral-specific markers, we generated retinoic acid-treated EBs grown in the presence or absence of HH-Ag. We did not observe induction of ventral Hh markers in RA-treated constitutive Gli1FLAG EBs and used these for the control, baseline set. The presence of FoxA2, Nkx2.9 and Nkx6.1 amongst the top 10 genes based on expression levels suggests that profiling significantly enriches for Hh-dependent cell types. As expected, the benchmark standard Gli1 was not up-regulated in our array, since it is constitutively expressed in the control as well. Keywords: neural progenitors, embryoid bodies, differentiation, Hedgehog, retinoic acid
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Project description:Embryonic stem (ES) cells, when grown in suspension culture without feeders, spontaneously form round structures known as embryoid bodies. Given the appropriate conditions, these cells can differentiate over time into precursors of all three germ layers. Embryoid bodies, in a disorganized way, mimic early embryonic development to a certain extent and can be used as a synchronously differentiating large scale source of tissue for the study of biological determinants of early differentiation. Embryoid bodies have been used as a source for most early protocols that derive specific differentiated cell types from undifferentiated ES cells, although some protocols, notably those that derive neurons from ES cells, have moved on from EBs as a result of varying replicability and yield. We have decided to look at the transcriptomic profiles of embryoid bodies during the initial stages of embryoid body formation and differentiation, in order to pinpoint novel determinants of key developmental stages. Keywords: time course
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Project description:To unravel the molecular mechanism by which HOXB4 promotes the expansion of early hematopoietic progenitors within differentiating ES cells, we analzed the gene expression profiles of embryoid bodies (EBs) in which transcription of HOXB4 had been induced or not induced. A substantial number of the identified HOXB4 target genes are involved in signaling pathways important for controlling self-renewal, maintenance and differentiation of stem cells. Furthermore, we demonstrate that HOXB4 activity and FGF-signaling are intertwined. HOXB4-mediated expansion of ES cell-derived early progenitors was enhanced by specific and complete inhibition of FGF-receptors. In contrast, the expanding activity of HOXB4 on hematopoietic progenitors in day4-6 embryoid bodies was blunted in the presence of basic FGF (FGF2) indicating a dominant negative effect of FGF-signaling on the earliest hematopoietic cells. Taken together, we show that modulation of FGF signaling is an essential feature of HOXB4 activity in the context of embryonic hematopoiesis. Experiment Overall Design: The Hoxb4i ES cell line (Kyba et al. 2002, Cell 109:29-37) contains an integrated âtet-onâ cassette that allows induction of HOXB4 expression upon treatment with doxycycline. These ES cells can be used to produce hematopoietic cells through the formation of embryoid bodies (EBs). Hematopoiesis starts in these EBs at day 4 and the differentiation into hematopoietic fates can be quantified by colony assays on methyl-cellulose using cells dissociated from EBs at day 6 of incubation. The induction of HOXB4 by incubation with doxycycline increases the production of hematopoietic progenitors within EBs by day 6. Using this specific ES cell line, we compared the transcriptome between embryoid bodies (EBs) in which transcription of HOXB4 had been induced or not induced from day 4 to day 6 (48hours). Experiment Overall Design: Biological replicates: 3
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