Ligand-dependent dynamics of retinoic acid receptor binding during early neurogenesis
Ontology highlight
ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE19409 | GEO | 2010/12/10
SECONDARY ACCESSION(S): PRJNA123691
REPOSITORIES: GEO
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Project description:Among its many roles in development, retinoic acid determines the anterior-posterior identity of differentiating motor neurons by activating Retinoic Acid Receptor (RAR)-mediated transcription. RAR is thought to bind the genome constitutively, and only induce transcription in the presence of the retinoid ligand. However, little is known about where RAR binds to the genome or how it selects target sites. We tested the constitutive RAR binding model using the retinoic acid-driven differentiation of mouse embryonic stem cells into differentiated motor neurons. We find that retinoic acid treatment results in widespread changes in RAR genomic binding, including novel binding to genes directly responsible for anterior-posterior specification, as well as the subsequent recruitment of the basal polymerase machinery. Finally, we discovered that the binding of transcription factors at the embryonic stem cell stage can accurately predict where in the genome RAR binds after initial differentiation. We have characterized a ligand-dependent shift in RAR genomic occupancy at the initiation of neurogenesis. Our data also suggests that enhancers active in pluripotent embryonic stem cells may be preselecting regions that will be activated by RAR during neuronal differentiation. The differentiation of ventral motor neurons is induced by treating embryonic stem cell cultures with retinoic acid. Here, ChIP-seq is used to profile the genome-wide occupancy of RAR isofroms both immediately prior to and during exposure of the cells to retinoic acid. ChIP-seq is also used to profile the genomic occupancy of Pol2 with phosphorylated serine 5 (Pol2-S5P) and phosphorylated serine 2 (Pol2-S2P) after exposure to retinoic acid.
2010-12-10 | E-GEOD-19409 | biostudies-arrayexpress
Project description:Haffez2017 - RAR interaction with synthetic
analogues
This model is described in the article:
The molecular basis of the
interactions between synthetic retinoic acid analogues and the
retinoic acid receptors
Hesham Haffez, David R. Chisholm,
Roy Valentine, Ehmke Pohl, Christopher Redfern and Andrew
Whiting
MedChemComm
Abstract:
All-trans-retinoic acid (ATRA) and its synthetic analogues
EC23 and EC19 direct cellular differentiation by interacting as
ligands for the retinoic acid receptor (RARα,
β and γ) family of nuclear receptor
proteins. To date, a number of crystal structures of natural
and synthetic ligands complexed to their target proteins have
been solved, providing molecular level snap-shots of ligand
binding. However, a deeper understanding of receptor and ligand
flexibility and conformational freedom is required to develop
stable and effective ATRA analogues for clinical use.
Therefore, we have used molecular modelling techniques to
define RAR interactions with ATRA and two synthetic analogues,
EC19 and EC23, and compared their predicted biochemical
activities to experimental measurements of relative ligand
affinity and recruitment of coactivator proteins. A
comprehensive molecular docking approach that explored the
conformational space of the ligands indicated that ATRA is able
to bind the three RAR proteins in a number of conformations
with one extended structure being favoured. In contrast the
biologically-distinct isomer, 9-cis-retinoic acid (9CRA),
showed significantly less conformational flexibility in the RAR
binding pockets. These findings were used to inform docking
studies of the synthetic retinoids EC23 and EC19, and their
respective methyl esters. EC23 was found to be an excellent
mimic for ATRA, and occupied similar binding modes to ATRA in
all three target RAR proteins. In comparison, EC19 exhibited an
alternative binding mode which reduces the strength of key
polar interactions in RARα/γ but is
well-suited to the larger RARβ binding pocket. In
contrast, docking of the corresponding esters revealed the loss
of key polar interactions which may explain the much reduced
biological activity. Our computational results were
complemented using an in vitro binding assay based on FRET
measurements, which showed that EC23 was a strongly binding,
pan-agonist of the RARs, while EC19 exhibited specificity for
RARβ, as predicted by the docking studies. These
findings can account for the distinct behaviour of EC23 and
EC19 in cellular differentiation assays, and additionally, the
methods described herein can be further applied to the
understanding of the molecular basis for the selectivity of
different retinoids to RARα, β and
γ.
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2013-01-31 | BIOMD0000000629 | BioModels
Project description:In mouse embryonic cells, a retinoic acid (RA) stimulation triggers a massive change of gene expression leading the pluripotent, proliferating cells to a lineage-specific differentiation process. The retinoic acid receptor (RAR) plays a key role in this response by inhibiting pluripotency-maintaining genes and simultaneously activating some major actors of cell differentiation. To investigate the mechanism underlying this dual regulation, we performed joint RAR/RXR ChIP-seq and mRNA-seq time series during the first 48 hours of the RA-induced Primitive Endoderm differentiation process in F9 embryonic carcinoma cells. We detected significantly more RAR/RXR binding regions than previous studies and identified among them a handful of typical binding intensity patterns during differentiation. We demonstrate that these patterns are correlated with the coincidental binding of essential transcription factors (TFs) for pluripotency maintenance or PrE differentiation of embryonic stem (ES) cells, as well as the presence of variants of RAR binding motifs. Most importantly, early-bound regions coincide with pluripotency-associated transcription factor binding in ES (like Pou5f1, Sox2, Esrrb and Nr5a2) and display an increased frequency of the DR0 type RAR binding motifs; late-bound sites are associated to the PrE marker Sox17 and are enriched in the canonical DR5 binding motif. Our data offer an unprecedently detailed view on the action of RA in triggering pluripotent cell differentiation. Altogether, this work sheds light on the relocation of RAR/RXR binding sites throughout differentiation, and shows how RAR/RXR progressively shift from DR0 enriched regions, which were specifically identified in undifferentiated models, to canonical RAR binding sites containing loci. Time course (0, 2, 6, 12, 24, 48h) after stimulation of F9 cultured cells by retinoic acid: PanRAR and PanRXR ChIP-seq at 0, 2, 24 and 48h (no replicate); WCE-seq at 0h (no replicate); mRNA-seq at 0, 6, 12, 24, 48h (2 replicates except for time 0h, 4 replicates). Additionally, a control time course (culture in DMSO) sampled at 24 and 48h (no replicates).
2015-04-17 | E-GEOD-56893 | biostudies-arrayexpress
Project description:In mouse embryonic cells, a retinoic acid (RA) stimulation triggers a massive change of gene expression leading the pluripotent, proliferating cells to a lineage-specific differentiation process. The retinoic acid receptor (RAR) plays a key role in this response by inhibiting pluripotency-maintaining genes and simultaneously activating some major actors of cell differentiation. To investigate the mechanism underlying this dual regulation, we performed joint RAR/RXR ChIP-seq and mRNA-seq time series during the first 48 hours of the RA-induced Primitive Endoderm differentiation process in F9 embryonic carcinoma cells. We detected significantly more RAR/RXR binding regions than previous studies and identified among them a handful of typical binding intensity patterns during differentiation. We demonstrate that these patterns are correlated with the coincidental binding of essential transcription factors (TFs) for pluripotency maintenance or PrE differentiation of embryonic stem (ES) cells, as well as the presence of variants of RAR binding motifs. Most importantly, early-bound regions coincide with pluripotency-associated transcription factor binding in ES (like Pou5f1, Sox2, Esrrb and Nr5a2) and display an increased frequency of the DR0 type RAR binding motifs; late-bound sites are associated to the PrE marker Sox17 and are enriched in the canonical DR5 binding motif. Our data offer an unprecedently detailed view on the action of RA in triggering pluripotent cell differentiation. Altogether, this work sheds light on the relocation of RAR/RXR binding sites throughout differentiation, and shows how RAR/RXR progressively shift from DR0 enriched regions, which were specifically identified in undifferentiated models, to canonical RAR binding sites containing loci.
2015-04-17 | GSE56893 | GEO
Project description:The aim of this study is to profile gene expression dynamics during the in vitro differentiation of embryonic stem cells into ventral motor neurons. Expression levels were profiled using Affymetrix microarrays at six timepoints during in vitro differentiation: ES cells (Day 0), embryoid bodies (Day 2), retinoid induction of neurogenesis (Day 2 +8hours of exposure to retinoic acid), neural precursors (Day 3), progenitor motor neurons (Day 4), postmitotic motor neurons (Day 7). The differentiation of ventral motor neurons is induced by treating embryonic stem cell cultures with retinoic acid and hedgehog agonist. Here, gene expression patterns are profiled at various defined stages during the differentiation process using Affymetrix expression arrays.
2010-12-10 | E-GEOD-19372 | biostudies-arrayexpress
Project description:Retinoic acid receptors (RARs) ?, ?, and ? heterodimerize with Retinoid X receptors (RXR) ?, ?, and ? and bind the cis-acting response elements known as RAREs to execute the biological functions of retinoic acid during mammalian development. RAR? mediates the anti-proliferative and apoptotic effects of retinoids in certain tissues and cancer cells, such as melanoma and neuroblastoma cells. Furthermore, ablation of RAR? enhanced the tumor incidence of Ras transformed keratinocytes and was associated with resistance to retinoid mediated growth arrest and apoptosis. We used microarray analysis to identify genes, which upon 8 or 24 hr of treatment with all-trans retinoic acid display differential expression in RAR? knockout (RAR?KO) murine embryonic stem cells relative to CCE WT cells. We demonstrate that following RA treatment the majority of inducible transcripts are present at lower levels in RAR?KO ES cells compared to WT ES cells. Murine embryonic stem cells (WT and RAR?KO) were treated with either all-trans retinoic acid (up to 24 hr) or with vehicle control (EtOH).
2012-12-31 | E-GEOD-43221 | biostudies-arrayexpress
Project description:All-trans retinoic acid (ATRA) alters gene expression in CCE WT embryonic stem cells, but has no effect on gene expression in RAR-deficient TKO cells (devoid of Retinoic Acid Receptors α, β, and γ)
2018-05-25 | GSE112412 | GEO
Project description:Purpose: In acute myeloid leukemia (AML) without retinoic acid receptor (RAR) rearrangement the effect of all-trans retinoic acid (ATRA) is still poorly understood despite an association of NPM1 mutation and ATRA response. Recently, PRAME (preferentially expressed antigen in melanoma) has been shown to be a dominant repressor of RAR-signaling. Experimental design: Thus, we further investigated ATRA response mechanisms, especially the impact of PRAME expression on ATRA-responsiveness by profiling gene expression in K562 cell lines. Results: Our data revealed a PRAME-expression associated gene pattern to be significantly enriched for genes involved in the retinoic acid metabolic process. In leukemia cell line models we could demonstrate that retinoic acid-regulated cell proliferation and differentiation are impacted by PRAME expression. Conclusions: PRAME seems to impair differentiation and to increase proliferation likely via blocking RAR-signaling, which might be reversed by ATRA.
2013-03-10 | GSE43258 | GEO
Project description:We defined the RARα interactome in the MDA-MB453 breast cancer cell line genetically engineered to over-express an N-terminally tagged version of the nuclear retinoic acid receptor. Twenty eight nuclear proteins which interact with RARα and whose interaction is stimulated or reduced by the pan-RAR ligand, all-trans retinoic acid (ATRA) were identified. Given the potential significance of the S100A3 calcium-binding protein in the control of tumor progression, we focused our attention on this factor. Using the two models represented by the ATRA-sensitive SKBR3 and MCF7 breast cancer cell lines characterized by constitutive expression of S100A3 and RARα, we demonstrate that the endogenous forms of S100A3 and RARα interact in physiological conditions. The interaction of S100A3 with RARα is cell context independent and it is observed not only in breast cancer but also in acute promyelocytioc leukemia (APL) cells, characterized by expression of the RARα-derived PML-RARα oncogene, which is the product of the t(15:17) chromosomal translocation. S100A3 interacts directly and specifically with RARα and PML-RARα, being unable to bind other members of the RAR/RXR family of retinoid nuclear receptors. The interaction surface maps to the carboxyl-terminal region of the RARα ligand binding domain. Binding of S100A3 to RARα and PML-RARα controls the constitutive and ATRA-dependent degradation of the two receptors. Silencing of the S100A3 gene decreases the amounts of RARα in breast SK-BR-3 and lung A549 cancer cells, rendering them more refractory to the anti-proliferative action of ATRA. In SK-BR-3 cells, this effect is accompanied by a decrease in the lactogenic/differentiating action of ATRA. In APL-derived NB4 cells, S100A3 knock-down reduces the amounts of both RARα and PML-RARα. Contemporaneous down-regulation of the two receptors is associated with an increase in the basal and ATRA-induced expression of many granulocytic differentiation markers. Opposite on RARα and PML-RARα levels as well as ATRA induced differentiation markers are observed upon over-expression of S100A3 in NB4 cells.
2019-12-18 | PXD008761 | Pride
Project description:Retinoic acid (RA) functions as a ligand for the nuclear RA receptors (RARs), which regulate the expression of target genes by binding to RA response elements. RA signaling is required for multiple processes during chordate embryonic development, such as body axis extension, hindbrain antero-posterior patterning and forelimb bud initiation. Although some RA target genes have been identified, little is known about the genome-wide effects of RA signaling during in vivo embryonic development. Here we stimulate the RA pathway during development by treating zebrafish embryos with all-trans-RA (atRA), the most abundant form of RA, and use a combination of RNA-seq, ATAC-seq, ChIP-seq and HiChIP to gain insight into the molecular mechanisms by which RA signaling control target gene expression. We find that RA signaling is involved in anterior/posterior patterning and development of the central nervous system, participating in the transition from pluripotency to differentiation. atRA treatment also induces alterations in chromatin accessibility during early development and promotes chromatin binding of RARaa and the RA targets Hoxb1b, Meis2b and Sox3, which cooperate in central nervous system development. Finally, we show that RA induces a rewiring of chromatin architecture, with alterations in chromatin 3D interactions that are consistent with target gene expression. This is illustrated by the specific induction of anterior HoxB genes by RARs, among other examples. Altogether, our findings identify genome-wide targets of RA signaling during embryonic development and provide a molecular mechanism by which developmental signaling pathways regulate the expression of target genes by altering chromatin topology.
2023-06-13 | GSE233695 | GEO