Expression data from shRFP, shATF1 in human embryonic stem cells (hESCs)
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE89397 | GEO | 2019/11/01
REPOSITORIES: GEO
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Project description:Mass spectrometry analysis was carried out to investigate the protein expression landscape of RNA-binding proteins (RBPs) during a time-course of spontaneously differentiating human embryonic stem cells (hESCs). Mass spectrometry was performed on whole-cell extracts of differentiating TE03 (I3) hESCs at four time-points (days 0, 23, 39, and 54), each with two biological replicates.
2023-05-05 | PXD025274 | Pride
Project description:Here, we performed RNA-interactome capture (RIC) on nuclear fractions from human embryonic stem cells (hESCs). The poly(A)+ RNA-bound proteome was determined by UV light-mediated cross-linking (CL) of RNAs to proteins in living cells, followed by nuclei isolation, oligo(dT) purification of poly(A)-RNA-protein complexes, and mass spectrometry analysis of captured proteins. As a control, we applied a similar strategy to non-cross-linked (non-CL) samples. RIC was performed in four independent biological replicates. This data accompanies the manuscript: "Uncovering the RNA-binding protein landscape in the pluripotency network of human embryonic stem cells". Abstract: "Embryonic stem cell (ESC) self-renewal and cell-fate decisions are driven by a broad array of molecular signals. While transcriptional regulators have been extensively studied in human ESCs (hESCs), the extent to which RNA-binding proteins (RBPs) contribute to human pluripotency remains unclear. Here, we carry out a proteome-wide screen and identify 810 proteins that directly bind RNA in hESCs. We reveal that RBPs are preferentially expressed in hESCs and dynamically regulated during exit from pluripotency and early lineage specification. Moreover, we show that nearly 200 RBPs are affected by knockdown of OCT4, a master regulator of pluripotency, several dozen of which are directly bound by this factor. Intriguingly, over 20 percent of the proteins detected in our study are putative DNA- and RNA-binding proteins (DRBPs), among them key transcription factors (TFs). Using fluorescently labeled RNA and seCLIP (single-end enhanced crosslinking and immunoprecipitation) experiments, we discover that the pluripotency-associated STAT3 and OCT4 TFs interact with RNA in hESCs and confirm the direct binding of STAT3 to the conserved NORAD long-noncoding RNA. Taken together, our findings indicate that RBPs have a more widespread role in human pluripotency than previously appreciated, reinforcing the importance of post-transcriptional regulation in stem cell biology".
2023-05-05 | PXD025262 | Pride
Project description:The proteome of undifferentiated human embryonic stem cells (hESCs) was profiled by deep mass spectrometry-based proteomics of whole-cell extracts from suspension cultures of TE03 cells, in four biological replicates. This data accompanies the manuscript: "Uncovering the RNA-binding protein landscape in the pluripotency network of human embryonic stem cells". Abstract: "Embryonic stem cell (ESC) self-renewal and cell-fate decisions are driven by a broad array of molecular signals. While transcriptional regulators have been extensively studied in human ESCs (hESCs), the extent to which RNA-binding proteins (RBPs) contribute to human pluripotency remains unclear. Here, we carry out a proteome-wide screen and identify 810 proteins that directly bind RNA in hESCs. We reveal that RBPs are preferentially expressed in hESCs and dynamically regulated during exit from pluripotency and early lineage specification. Moreover, we show that nearly 200 RBPs are affected by knockdown of OCT4, a master regulator of pluripotency, several dozen of which are directly bound by this factor. Intriguingly, over 20 percent of the proteins detected in our study are putative DNA- and RNA-binding proteins (DRBPs), among them key transcription factors (TFs). Using fluorescently labeled RNA and seCLIP (single-end enhanced crosslinking and immunoprecipitation) experiments, we discover that the pluripotency-associated STAT3 and OCT4 TFs interact with RNA in hESCs and confirm the direct binding of STAT3 to the conserved NORAD long-noncoding RNA. Taken together, our findings indicate that RBPs have a more widespread role in human pluripotency than previously appreciated, reinforcing the importance of post-transcriptional regulation in stem cell biology".
2023-05-05 | PXD025278 | Pride
Project description:Embryonic stem cell (ESC) self-renewal and cell-fate decisions are driven by a broad array of molecular signals. While transcriptional regulators have been extensively studied in human ESCs (hESCs), the extent to which RNA-binding proteins (RBPs) contribute to human pluripotency remains unclear. Here, we carried out a proteome-wide screen and identified 810 proteins that directly bind RNA in hESCs. We determined the RBP catalog by using RNA-interactome capture (RIC), a method based on UV light-mediated cross-linking (CL) of RNAs to proteins in living cells, followed by oligo(dT) purification of poly(A)-RNA-protein complexes and mass spectrometry analysis of captured proteins. As control, we applied a similar strategy to non-cross-linked (non-CL) samples. To uncover the identity of the eluted proteins, we performed in-solution tryptic digestion of CL and non-CL eluates and analyzed their contents by a high-resolution mass spectrometer (Q-Exactive Plus). We then performed differential proteome analysis between CL and non-CL eluates, resulting in a set of 810 high-confidence protein groups, defined as the hESC RNA-interactome. RIC was carried out in four independent biological replicates. This data accompanies the manuscript: "Uncovering the RNA-binding protein landscape in the pluripotency network of human embryonic stem cells".
2023-05-05 | PXD025194 | Pride
Project description:To detect the effect of early culture on hESCs, almost all initial-passaged hESCs have normal expression of DLK1-DIO3 gene cluster, the low-passaged hESCs lost the expression of DLK1-DIO3 gene cluster.
2014-06-30 | GSE38662 | GEO
Project description:To compare transcriptomic profiles between UCSF4 hESCs and their neural derivatives, neural precursor cells (NPCs), we performed microarray analysis using the Affymetrix Human Gene 2.0 ST array platform.
2016-03-01 | GSE74378 | GEO
Project description:To detect the effect of early culture on hESCs, almost all initial-passaged hESCs have normal expression of DLK1-DIO3 gene cluster, the low-passaged hESCs lost the expression of DLK1-DIO3 gene cluster. hESCs were extracted for RNA with TriZOL and hybridization on Affymetrix arrays
2014-06-30 | E-GEOD-38662 | biostudies-arrayexpress
Project description:To compare transcriptomic profiles between UCSF4 hESCs and their neural derivatives, neural precursor cells (NPCs), we performed microarray analysis using the Affymetrix Human Gene 2.0 ST array platform. Six biological samples, three biological replicates for each developmental cell stage (in vitro)
2016-03-01 | E-GEOD-74378 | biostudies-arrayexpress
Project description:Rapamycin inhibits mTORC1 complex in a variety of cells. We demonstrated that hESCs are more sensitive to rapamycin treatment as compared to somatic cells (BJ Fibroblasts) and also certain cancer cells (KBM7 cells). To study the molecular mechanisms of this disparity, we performed a transcriptome analysis (RNAseq) of rapamycin-treated hESCs and human fibroblasts for two time points (2 and 4 days of treatment).
2018-04-30 | GSE107965 | GEO
Project description:We used microarray analysis to study the expression differences between controls and hESCs expressing RB7LP for 3 days, and controls and hESCs expressing T121 for 3 days. RB7LP is the large pocket fragment of the retinoblastoma protein (RB) fused to GFP, in which seven phosphorylation sites for Cdk have been mutated (Angus, SP et al. Mol Cell Biol 23, 8172 (Nov, 2003). T121 is a truncated form of the SV40 Large T (LT) viral oncoprotein that inactivates the function of the three members of the retinoblastoma (RB) protein family (RB, p107, p130). We plated hESCs that had been infected with inducible lentiviruses to express either RB7LP or T121. The day after plating, one well from each virus was treated with doxycyline to induce expression of either RB7LP or T121 and after 3 days were harvested for microrarray analysis.
2012-06-28 | E-GEOD-29783 | biostudies-arrayexpress