Expression data from the inducible expression of EWS-FLI1 (EF) in embryoid bodies
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE64686 | GEO | 2015/12/31
SECONDARY ACCESSION(S): PRJNA271644
REPOSITORIES: GEO
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Project description:Oncogenic transformation in Ewing sarcoma tumors is driven by the fusion oncogene EWS-FLI1. The inducible expression of EWS-FLI1 (EF) in embryoid bodies, or collections of differentiating stem cells, generates cells with properties of Ewing sarcoma tumors, including characteristics of transformation. These cell lines exhibit anchorage-independent growth, a lack of contact inhibition and a strong Ewing sarcoma gene expression signature. These cells also demonstrate a requirement for the persistent expression of EWS-FLI1 for cell survival and growth. We used microarrays to detail the effects of doxycycline-inducible expression of EWS-FLI1 on the gene expression signature of cells derived from differentiating stem cells, or embryoid bodies. Triplicate biological replicates were collected and analyzed.
2015-12-31 | E-GEOD-64686 | biostudies-arrayexpress
Project description:Ewing sarcoma is a highly aggressive tumor characterized by a translocation between members of the FET family of RNA binding proteins and one of several ETS transcription factors, with the most common translocation being EWS-FLI1. EWS-FLI1 leads to changes in gene expression through mechanisms that are not completely understood. We performed RNA sequencing analysis on primary pediatric human mesenchymal progenitor cells (pMPCs) expressing EWS-FLI1 in order to identify novel target genes. This analysis identified lnc277 as a previously uncharacterized long non-coding RNA upregulated by EWS-FLI1 in pMPCs. Inhibiting the expression of lnc277 diminished the ability of Ewing sarcoma cell lines to proliferate and form colonies in soft agar whereas inhibiting lnc277 had no effect on other cell types tested. By analyzing gene expression after shRNA knockdown, we found that both EWS-FLI1 and lnc277 repressed many more genes that they induced and that a significant fraction of EWS-FLI1 repressed targets were also repressed by lnc277. Analysis of primary human Ewing sarcoma RNA sequencing data further supports a role for lnc277 in mediating gene repression. We identified hnRNPK as an RNA binding protein that interacts directly with lnc277. We found a significant overlap in the genes repressed by hnRNPK and those repressed by both EWS-FLI1 and lnc277, suggesting that hnRNPK participates in lnc277 mediated gene repression. Thus, lnc277 is a previously uncharacterized long non-coding RNA downstream of EWS-FLI1 that facilitates the development of Ewing sarcoma via the repression of target genes. Our studies identify a novel mechanism of oncogenesis downstream of a chromosomal translocation and underscore the importance of lncRNA-mediated gene repression as a mechanism of EWS-FLI1 transcriptional regulation. RNA from primary human bone marrow derived mesenchymal cells either control or with inducible expression of EWS-FLI1 for 13 days was used to prepare PolyA selected cDNA libraries.
2014-11-01 | E-GEOD-60889 | biostudies-arrayexpress
Project description:Ewing sarcoma (EWS) is a malignant pediatric bone cancer. Most Ewing sarcomas are driven by EWS-FLI1 oncogenic transcription factor that plays roles in transcriptional regulation, DNA damage response, cell cycle checkpoint control, and alternative splicing. USP1, a deubiquitylase which regulates DNA damage and replication stress responses, is overexpressed at both the mRNA and protein levels in EWS cell lines compared to human mesenchymal stem cells, the EWS cell of origin. The functional significance of high USP1 expression in Ewing sarcoma is not known. Here, we identify USP1 as a transcriptional target of EWS-FLI1 and a key regulator of EWS cell survival. We show that EWS-FLI1 knockdown decreases USP1 mRNA and protein levels. ChIP and ChIP-seq analyses show EWS-FLI1 occupancy on the USP1 promoter. Importantly, USP1 knockdown or inhibition arrests EWS cell growth and induces cell death by apoptosis. We observe destabilization of Survivin (also known as BIRC5 or IAP4) and activation of caspases-3 and -7 following USP1 knockdown or inhibition in the absence of external DNA damage stimuli. Notably, EWS cells display hypersensitivity to combinatorial treatment of doxorubicin or etoposide, EWS standard of care drugs, and USP1 inhibitor compared to single agents alone. Together, our study demonstrates that USP1 is regulated by EWS-FLI1, the USP1-Survivin axis promotes EWS cell survival, and USP1 inhibition sensitizes EWS cells to standard of care chemotherapy.
2024-01-26 | PXD041033 | Pride
Project description:Ewing sarcoma is a highly aggressive tumor characterized by a translocation between members of the FET family of RNA binding proteins and one of several ETS transcription factors, with the most common translocation being EWS-FLI1. EWS-FLI1 leads to changes in gene expression through mechanisms that are not completely understood. We performed RNA sequencing analysis on primary pediatric human mesenchymal progenitor cells (pMPCs) expressing EWS-FLI1 in order to identify novel target genes. This analysis identified lnc277 as a previously uncharacterized long non-coding RNA upregulated by EWS-FLI1 in pMPCs. Inhibiting the expression of lnc277 diminished the ability of Ewing sarcoma cell lines to proliferate and form colonies in soft agar whereas inhibiting lnc277 had no effect on other cell types tested. By analyzing gene expression after shRNA knockdown, we found that both EWS-FLI1 and lnc277 repressed many more genes that they induced and that a significant fraction of EWS-FLI1 repressed targets were also repressed by lnc277. Analysis of primary human Ewing sarcoma RNA sequencing data further supports a role for lnc277 in mediating gene repression. We identified hnRNPK as an RNA binding protein that interacts directly with lnc277. We found a significant overlap in the genes repressed by hnRNPK and those repressed by both EWS-FLI1 and lnc277, suggesting that hnRNPK participates in lnc277 mediated gene repression. Thus, lnc277 is a previously uncharacterized long non-coding RNA downstream of EWS-FLI1 that facilitates the development of Ewing sarcoma via the repression of target genes. Our studies identify a novel mechanism of oncogenesis downstream of a chromosomal translocation and underscore the importance of lncRNA-mediated gene repression as a mechanism of EWS-FLI1 transcriptional regulation.
2014-11-01 | GSE60889 | GEO
Project description:EWS-FLI1 is the driver oncogene in >85% of Ewing sarcoma tumors. We used siRNA to knock down EWS-FLI1 for 48h hours in two Ewing sarcoma cell lines (EW8, TC71) to identify downstream genes and pathways affected by loss of EWS-FLI1 activity. RNAseq analysis was performed after 48 hours of treatment with a nontargeting siControl or siEWS-FLI1.
2024-09-30 | GSE263504 | GEO
Project description:We have performed a high-throughput RNA interference screen to identify targets inhibiting EWS-FLI1 driven cell proliferation in Ewing sarcoma cells. EWS-FLI1 expressing A673 Ewing sarcoma cells were screened both in presence and absence of EWS-FLI1 shRNA induction with druggable siRNA library. Leucine rich repeats and WD repeat Domain containing 1 (LRWD1) targeting siRNA pool was the strongest anti-proliferative hit identified only in presence of EWS-FLI1. Validation experiments confirmed the anti-proliferative effect of LRWD1 depletion especially in EWS-FLI1 expressing cells. Functional analysis of differentially expressed genes in LRWD1 depleted Ewing sarcoma cells showed over-representation of connective tissue development, cell projection morphogenesis and neuronal processes.
2018-09-16 | GSE73092 | GEO
Project description:Posterior homeobox D genes, in particular HOXD13, are over-expressed by Ewing sarcoma, a tumor driven by the oncogenic fusion protein EWS-FLI1. Here, we have found that EWS-FLI1 maintains HOXD13 expression through a GGAA microsatellite enhancer in the developmental posterior HOXD regulatory domain. Activation of this enhancer is EWS-FLI1 dependent and epigenomic silencing of this region leads to loss of HOXD13 expression in Ewing sarcoma cells, but not in unrelated cells. To determine the function of HOXD13 activation in Ewing sarcoma we performed nascent RNA sequencing or RNA sequencing upon HOXD13 knockdown.
2022-06-28 | GSE148494 | GEO
Project description:We identified global DNA binding properties of EWS-FLI1 in mouse Ewing sarcoma. GGAA microsatellites were found as binding sites of EWS-FLI1 but with less frequency than that in human Ewing sarcoma, and genomic distribution is not conserved between human and mouse.
2018-07-25 | GSE108631 | GEO
Project description:Ewing sarcoma is driven by fusion proteins containing a low complexity (LC) domain that is intrinsically disordered and a powerful transcriptional regulator. The most common fusion protein found in Ewing sarcoma, EWS-FLI1, takes its LC domain from the RNA-binding protein EWSR1 (Ewing Sarcoma RNA-binding protein 1) and a DNA-binding domain from the transcription factor FLI1 (Friend Leukemia Virus Integration 1). The LC domain in EWS-FLI1 can bind RNA polymerase II (RNA Pol II) and can self-assemble through a process known as phase separation. The ability of EWSR1 and related RNA-binding proteins to assemble into ribonucleoprotein granules in cells has been intensely studied but the role of phase separation in EWS-FLI1 activity is less understood. We investigated the overlapping functions of EWSR1 and EWS-FLI1 in controlling gene expression and tumorigenic cell growth in Ewing sarcoma, and our results suggested that these proteins function closely together. We then studied the nature of interactions among EWS-FLI1, EWSR1, and RNA Pol II. We observed EWSR1 and RNA Pol II to be present in protein granules in cells. We then identified protein granules in cells associated with the fusion protein, EWS-FLI1. The tyrosine residues in the LC domain are required for the abilities of EWS-FLI1 to bind its partners, EWSR1 and RNA Pol II, and to incorporate into protein granules. These data suggest that interactions among EWS-FLI1, RNA Pol II, and EWSR1 in Ewing sarcoma can occur in the context of a molecular scaffold found within protein granules in the cell.
2020-07-23 | GSE154944 | GEO
Project description:Comparison of gene expression profile of Ewing sarcoma cells which have an exchange of the endogenous EWS/FLI1 to either wild-type or a turnover-deficient mutant EWS/FLI1. Most target genes are saturated as only a few target genes are soly driven by increasing protein amount. The effect of a stable EWS/FLI1 mutant on global gene expression was evaluated in A673 Ewing sarcoma cells.
2016-11-16 | GSE81018 | GEO