Overlapping and distinct functions between ERα and ERβ homodimers and corresponding transcriptomes in the same cellular context
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE182431 | GEO | 2022/06/03
REPOSITORIES: GEO
Similar Datasets
Project description:Estrogen Receptor subtypes (ERα and ERβ) are transcription factors sharing similar structure, however, they often perform opposite roles in breast cancer’s cell proliferation and tumor progression. Besides the well-characterized genomic actions of ERs upon ligand binding, rapid non-genomic cytoplasmic changes together with the recently discovered ligand-free action of ERs are emerging as key regulators of tumorigenesis. The identification of cytoplasmic interaction partners of unliganded ERα and ERβ may help characterize the molecular basis of the extra-nuclear mechanism of action of these receptors, revealing novel mechanisms to explain their role in breast cancer response or resistance to endocrine therapy. To this aim, in this study, cytoplasmic extracts from stably expressing TAP-ERα and -ERβ MCF-7 cell clones were subjected to interaction proteomics in the absence of estrogen stimulation, leading to the identification of 84 and 142 proteins associated with unliganded ERα and ERβ, respectively. Functional analyses of ER subtype-specific interactomes revealed significant differences in the molecular pathways associated to each receptor in the cytoplasm. This work reports the first identification of the unliganded ERα and ERβ cytoplasmic interactomes in breast cancer cells, providing novel experimental evidence on the non-genomic effects of ERs in the absence of hormonal stimulus.
2015-03-11 | PXD001202 | Pride
Project description:The C4-12/Flag.ERβ cell line which stably expressed Flag.ERβ is used to study ERβ genomic functions without ERα interference. Mapping ERβ binding sites in these cells reveals ERβ unique distribution and motif enrichment patterns. Accompanying our mapping results, nascent RNA profiling is performed on cells at the same treatment time. The combined results allow the identification of ERβ target genes. Gene ontology analysis reveals that ERβ targets are enriched in differentiation, development and apoptosis. Concurrently, E2 treatment suppresses proliferation in these cells. Within ERβ binding sites, while the most prevalent binding motif is the canonical ERE, motifs of known ER interactors are also enriched in ERβ binding sites. Moreover, among enriched binding motifs are those of GFI, REST and EBF1, which are unique to ERβ binding sites in these cells. Further characterization confirms the association between EBF1 and the estrogen receptors, which favors the N-terminal region of the receptor. Furthermore, EBF1 negatively regulates ERs at the protein level. In summary, by studying ERβ genomic functions in our cell model, we confirm the anti-proliferative role of ERβ and discover the novel cross talk of ERβ with EBF1 which has various implications in normal physiology. C4-12/Flag.ERβ cells were treated with 10nM E2 (or ethanol as vehicle control) for 1 hour. Nuclei were extracted and processed with run-on assay. The resultant run-on RNA was reverse-transcribed to generate cDNA library which was subsequently sequenced by Illumina Genome Analyzer II or HiSeq2000. Two samples for each treatment were included in this experiment.
2013-06-21 | E-GEOD-48159 | biostudies-arrayexpress
Project description:The C4-12/Flag.ERβ cell line which stably expressed Flag.ERβ is used to study ERβ genomic functions without ERα interference. Mapping ERβ binding sites in these cells reveals ERβ unique distribution and motif enrichment patterns. Accompanying our mapping results, nascent RNA profiling is performed on cells at the same treatment time. The combined results allow the identification of ERβ target genes. Gene ontology analysis reveals that ERβ targets are enriched in differentiation, development and apoptosis. Concurrently, E2 treatment suppresses proliferation in these cells. Within ERβ binding sites, while the most prevalent binding motif is the canonical ERE, motifs of known ER interactors are also enriched in ERβ binding sites. Moreover, among enriched binding motifs are those of GFI, REST and EBF1, which are unique to ERβ binding sites in these cells. Further characterization confirms the association between EBF1 and the estrogen receptors, which favors the N-terminal region of the receptor. Furthermore, EBF1 negatively regulates ERs at the protein level. In summary, by studying ERβ genomic functions in our cell model, we confirm the anti-proliferative role of ERβ and discover the novel cross talk of ERβ with EBF1 which has various implications in normal physiology. C4-12/Flag.ERβ cells were treated with 10nM E2 for 1 hour then crosslinked with 1% formaldehyde; EtOH treatment was used as control. Crosslinked samples were processed for ChIP-seq and sequenced with Illumina Genome Analyzer II and aligned to hg18. QuEST was used as the peak-calling software, using default parameters recommended to analyze transcription factor ChIP-seq data. The entire ChIP-seq process was performed once on each sample (Vehicle or E2-treated).
2013-06-21 | E-GEOD-48096 | biostudies-arrayexpress
Project description:The closely related transcription factors (TFs), estrogen receptors ERα and ERβ, regulate divergent gene expression programs and proliferative outcomes in breast cancer. Utilizing MCF-7 breast cancer cells with ERα, ERβ, or both receptors as a model system to define the basis of differing response specification by related TFs, we show that these TFs and their key coregulators, SRC3 and RIP140, generate overlapping as well as unique chromatin-binding and transcription-regulating modules. Cistrome and transcriptome analyses and use of clustering algorithms delineated 11 clusters representing different chromatin-bound receptor and coregulator assemblies that could be functionally associated through enrichment analysis with distinct patterns of gene regulation and preferential coregulator usage, RIP140 with ERβ and SRC3 with ERα. The receptors modified each other’s transcriptional effect, and ERβ countered the proliferative drive of ERα through several novel mechanisms associated with specific binding site clusters. Our findings delineate distinct TF-coregulator assemblies that function as control nodes specifying precise patterns of gene regulation, proliferation, and metabolism, as exemplified by two of the most important nuclear hormone receptors in human breast cancer. MCF-7 cells expressing endogenous ERalpha were infected with adenovirus carrying either estrogen receptor beta (AdERb) or no insert (Ad) at multiplicity of infection (moi) of 50. ERβ only cells were generated from these cells by knockdown of ERα in parental cells using the following siERα sequences from Dharmacon: forward, 5’-UCAUCGCAUUCCUUGCAAAdTdT-3’, and reverse, 5’-UUUGCAAGGAAUGCGAUGAdTdT-3’. siRNA experiments were performed as previously described, and resulted in knocknown of ERα mRNA and protein by greater than 95% (GSE4006, PMID 16809442). Briefly, cells were transfected with 20 nM siCtrl [GSE4006] or siERα for 48 h after infection. Then cells were treated with 0.1% EtOH (Veh) or 10 nM E2 (Sigma-Aldrich) for 24h. All experiments were conducted with two replicates. key words; Adenovirus infection,siRNA knock-down, ligand treatment
2013-07-05 | E-GEOD-42347 | biostudies-arrayexpress
Project description:Estrogens play an important role in breast cancer (BC) development and progression, where the two isoforms of the estrogen receptor (ERα and ERβ) are generally co-expressed and mediate the effects of these hormones in cancer cells. ERβ has been suggested to exert an antagonist role toward the oncogenic activities of ERα, and for this reason it is considered an oncosuppressor. As clinical evidence regarding a prognostic role for this receptor subtype in hormone-responsive BC is still limited and conflicting, more knowledge is required on the biological functions of ERβ in cancer cells. We described previously the ERβ and ERα interactomes of BC cells, identifying specific and distinct patterns of protein interactions for the two receptors. In particular, we identified factors involved in mRNA splicing and maturation as important components of both ERα and ERβ pathways. Guided by these findings, we investigated here in depth the differences in the early transcriptional events and RNA splicing patterns induced in ERα vs ERα+ERβ cells, by expressing ERβ in ERα+ human BC MCF-7 cells. High-throughput mRNA sequencing was then performed in both cell lines after stimulation with 17b-estradiol, and the results obtained were compared.
2015-09-01 | GSE64590 | GEO
Project description:The human ERβ variant, ERβ2, is expressed at higher levels than ERβ1 in many breast cancer cell lines and breast tumours and increasing evidence supports the hypothesis that ERβ2, in contrast to ERβ1, is associated with aggressive phenotypes of various cancers. We identified a breast cancer cell line that expresses endogenous ERβ2, but not ERα or ERβ1, allowing studies of ERβ2 in the absence of other ER isoforms. Importantly, we show that knockdown of endogenous ERβ2 inhibited cell proliferation and cell invasion. To gain insights into the molecular mechanism responsible for these cellular phenotypes, we assayed ERβ2 dependent global gene expression profiles. A total of 263 genes were identified as potentially ERβ2-upregulated genes and 662 as ERβ2-downregulated genes. ERβ2-regulated genes were involved in cell morphology, DNA replication and repair, cell death and survival. Our findings demonstrate that ERβ2 alone has a key role in enhancing cell proliferation and invasion, beyond modulation of ERα and ERβ1 signalling. BT549 cells grown in antibiotic free medium were transfected with a control siRNA, siRNA targeting ERβ2, ERβ2 cDNA, empty vector control, and then global gene expression profiles were assessed. Three biological replicates were used for each group.
2015-05-01 | E-GEOD-57379 | biostudies-arrayexpress
Project description:The C4-12/Flag.ERβ cell line which stably expressed Flag.ERβ is used to study ERβ genomic functions without ERα interference. Mapping ERβ binding sites in these cells reveals ERβ unique distribution and motif enrichment patterns. Accompanying our mapping results, nascent RNA profiling is performed on cells at the same treatment time. The combined results allow the identification of ERβ target genes. Gene ontology analysis reveals that ERβ targets are enriched in differentiation, development and apoptosis. Concurrently, E2 treatment suppresses proliferation in these cells. Within ERβ binding sites, while the most prevalent binding motif is the canonical ERE, motifs of known ER interactors are also enriched in ERβ binding sites. Moreover, among enriched binding motifs are those of GFI, REST and EBF1, which are unique to ERβ binding sites in these cells. Further characterization confirms the association between EBF1 and the estrogen receptors, which favors the N-terminal region of the receptor. Furthermore, EBF1 negatively regulates ERs at the protein level. In summary, by studying ERβ genomic functions in our cell model, we confirm the anti-proliferative role of ERβ and discover the novel cross talk of ERβ with EBF1 which has various implications in normal physiology.
2013-06-21 | GSE48096 | GEO
Project description:The C4-12/Flag.ERβ cell line which stably expressed Flag.ERβ is used to study ERβ genomic functions without ERα interference. Mapping ERβ binding sites in these cells reveals ERβ unique distribution and motif enrichment patterns. Accompanying our mapping results, nascent RNA profiling is performed on cells at the same treatment time. The combined results allow the identification of ERβ target genes. Gene ontology analysis reveals that ERβ targets are enriched in differentiation, development and apoptosis. Concurrently, E2 treatment suppresses proliferation in these cells. Within ERβ binding sites, while the most prevalent binding motif is the canonical ERE, motifs of known ER interactors are also enriched in ERβ binding sites. Moreover, among enriched binding motifs are those of GFI, REST and EBF1, which are unique to ERβ binding sites in these cells. Further characterization confirms the association between EBF1 and the estrogen receptors, which favors the N-terminal region of the receptor. Furthermore, EBF1 negatively regulates ERs at the protein level. In summary, by studying ERβ genomic functions in our cell model, we confirm the anti-proliferative role of ERβ and discover the novel cross talk of ERβ with EBF1 which has various implications in normal physiology.
2013-06-21 | GSE48159 | GEO
Project description:We explored the combinatorial interactions between p53, estrogen receptors and NFkB using the breast adenocarcinoma-derived MCF7 cells. Recently, we have established that p53 can modulate transcription also through non-canonical response elements (REs), consisting of half-sites and ¾ sites. In particular, we identified a half-site p53 response element in the promoter of FLT1/VEGFR-I gene that was required but not sufficient for the p53 responsiveness of the promoter. The transcriptional control required also estrogen receptor (ER) half-sites (EREs) located in close proximity to the p53 RE. Further studies led us to establish that p53-mediated transcription from the FLT1 half site RE is dependent on ER binding at the EREs and strongly influenced by the level of ER proteins, and the specific nature of the p53-inducing stress as well as ER ligand. In another study we found an enrichment of NFkB REs nearby p53 REs in p53 target genes, suggesting an additional mode of functional interactions between these two transcription factors. To identify at a genome scale the transcriptional network that selectively responds to the concomitant activation of p53, ER and/or NFkB, we measured gene expression upon treatment with specific chemotherapeutic agents combined with 17-β estradiol (E2) and/or the inflammatory cytokine TNFα. Firstly we measured using MCF7 cells growing in estrogen-depleted media the transcriptome changes induced by doxorubicin (doxo) and 5-fluorouracil (5FU), two different drugs both able to stabilize and activate the p53 protein. We next obtained the expression profiles in the presence of E2 with or without doxo or 5FU. We also measured transcriptome changes in response to TNFαalone or in combination with doxo and/or E2. All these analyses were conducted using the same batch of MCF7 cells (wild type for p53, ERα, ERβ -weakly-, and NFkB positive) that we had previously used to characterize the cis-mediated transcriptional cooperation between p53 and ER at the FLT1 gene. Keywords: transcription factor, transcriptional networks, p53, ER, NFkB, p53-ER-NFkB crosstalk, doxorubicin, 5-fluorouracil, 17-β estradiol, TNFα, combined genotoxic, estrogenic and inflammatory responses, composite DNA binding site signatures, MCF7. The cooperation at the transcriptional level among p53, ERs and NFkB was analyzed by transcriptome analysis in response to different stimuli (doxo or 5FU, E2, TNFα). To identify mRNA molecules that would be modulated by the coordinated activity of the three TFs, gene expression signals derived from the combinatorial treatment were compared by microarrays analyses to those obtained from each drug when individually administered. Total RNA was isolated from MCF7 cells grown in estrogen-depleted medium and treated so as to stimulate the activity of the three different transcription factors (p53, ERs and NFkB). Cells lysates were collected after 10 hours from the treatments. All experiments were run from triplicate to septuples.
2011-06-01 | E-GEOD-24065 | biostudies-arrayexpress
Project description:We explored the combinatorial interactions between p53, estrogen receptors and NFkB using the breast adenocarcinoma-derived MCF7 cells. Recently, we have established that p53 can modulate transcription also through non-canonical response elements (REs), consisting of half-sites and ¾ sites. In particular, we identified a half-site p53 response element in the promoter of FLT1/VEGFR-I gene that was required but not sufficient for the p53 responsiveness of the promoter. The transcriptional control required also estrogen receptor (ER) half-sites (EREs) located in close proximity to the p53 RE. Further studies led us to establish that p53-mediated transcription from the FLT1 half site RE is dependent on ER binding at the EREs and strongly influenced by the level of ER proteins, and the specific nature of the p53-inducing stress as well as ER ligand. In another study we found an enrichment of NFkB REs nearby p53 REs in p53 target genes, suggesting an additional mode of functional interactions between these two transcription factors. To identify at a genome scale the transcriptional network that selectively responds to the concomitant activation of p53, ER and/or NFkB, we measured gene expression upon treatment with specific chemotherapeutic agents combined with 17-β estradiol (E2) and/or the inflammatory cytokine TNFα. Firstly we measured using MCF7 cells growing in estrogen-depleted media the transcriptome changes induced by doxorubicin (doxo) and 5-fluorouracil (5FU), two different drugs both able to stabilize and activate the p53 protein. We next obtained the expression profiles in the presence of E2 with or without doxo or 5FU. We also measured transcriptome changes in response to TNFα alone or in combination with doxo and/or E2. All these analyses were conducted using the same batch of MCF7 cells (wild type for p53, ERα, ERβ -weakly-, and NFkB positive) that we had previously used to characterize the cis-mediated transcriptional cooperation between p53 and ER at the FLT1 gene. Keywords: transcription factor, transcriptional networks, p53, ER, NFkB, p53-ER-NFkB crosstalk, doxorubicin, 5-fluorouracil, 17-β estradiol, TNFα, combined genotoxic, estrogenic and inflammatory responses, composite DNA binding site signatures, MCF7.
2011-06-01 | GSE24065 | GEO