ABSTRACT: Mammalian lung development during the saccular and alveolar stages is dependent upon antagonistic molecular signalling by endogenous retinoic acid (RA) and glucocorticoids (GCs) which regulate gene expression via the retinoic acid receptor (RAR) family and the glucocorticoid receptor (GR), respectively. The genomic mechanism of this antagonism was investigated with in vitro distal lung explant cultures from E18.5 GR-null (GR-/-) mice treated with all-trans-RA (atRA) for 2h . Whole mouse genome microarray analysis from lung explant tissue identified a small number of gene targets which were not only significantly induced by atRA in the wildtype lung, but also significantly stimulated to levels greater than atRA-treated wildtype lungs in GR-/- lungs.
Project description:The aim of this study is to identify novel glucocorticoid receptor (GR) regulated hepatic target genes. In order to do so, we used microarray technology to compare the gene expression of wild type and GR-null mice after 3 hrs of dexamethasone treatment.
Project description:Development of eukaryotic organisms is controlled by transcription factors that trigger specific and global changes in gene expression programmes. In plants, MADS-domain transcription factors act as master regulators of developmental switches and organ specification. However, the mechanisms by which these factors dynamically regulate the expression of their target genes at different developmental stages are still poorly understood. Here, we characterize the dynamic relationship of chromatin accessibility, gene expression and DNA-binding of two MADS-domain proteins during Arabidopsis flower development. The developmental dynamics of DNA-binding of APETALA1 and SEPALLATA3 is largely independent of chromatin accessibility, and our findings suggest that AP1 acts as 'pioneer factor' that modulates chromatin accessibility, thereby facilitating access of other transcriptional regulators to their target genes. Our data provide a primer to the idea that cellular differentiation in plants can be associated to dynamic changes in chromatin accessibility, as consequence of the action of master transcription factors. We used the AP1-GR system to conduct chromatin immunoprecipitation experiments with SEP3-specific antibodies and GR atibodies followed by deep-sequencing (ChIP-Seq) in order to determine SEP3 and AP1 binding sites on a genome-wide scale. Samples were generated from tissue in which the AP1-GR protein was induced using a treatment of 1 uM DEX to the shoot apex. The material was collect 2, 4 and 8 days after treatment. As control, we performed ChIP experiments using pre-immune serum at the different time points. Experiments were done in two biological replicates for 4 days and 8 days time-points while one biological replicate was done for control samples and 2 days time-point. The GSE47981 includes expression data that are complementary to the data in the GSE46986 and GSE46894.
Project description:Phenotypic plasticity, the ability of one genotype to express different phenotypes in response to changing environmental conditions, is one of the most common phenomena characterising the living world and is not only relevant for the ecology but also for the evolution of species. Daphnia, the waterflea, is a textbook example for predator induced phenotypic plastic defences including changes in life-history, behaviour and morphology. However, the analysis of molecular mechanisms underlying these inducible defences is still in its early stages.<br><br>We exposed Daphnia magna to chemical cues of the predator Triops cancriformis to identify key processes underlying plastic defensive trait formation. D. magna is known to develop an array of morphological changes in the presence of T. cancriformis including changes of carapace morphology and cuticle hardening. To get a more comprehensive idea of this phenomenon, we studied four different genotypes originating from habitats with different predation history, reaching from predator-free to temporary habitats containing T. cancriformis.<br><br>We analysed the morphologies as well as proteomes of predator-exposed and control animals. Three genotypes showed morphological changes when the predator was present. Using a high-throughput proteomics approach, we found 294 proteins which were significantly altered in their abundance after predator exposure in a general or genotype dependant manner. Proteins connected to genotype dependant responses were related to the cuticle, protein synthesis and calcium binding whereas the yolk protein vitellogenin increased in abundance in all genotypes, indicating their involvement in a more general response. Furthermore, genotype dependant responses at the proteome level correlated well with local adaptation to Triops predation.<br><br>Altogether, our study provides new insights concerning genotype dependant and general molecular processes involved in predator-induced phenotypic plasticity in D. magna.
Project description:Exogenous glucocorticoids are frequently used to treat inflammatory disorders and as adjuncts in solid cancers. However, their use is associated with severe side effects and therapy resistance. Novel glucocorticoid receptor (GR) ligands with a patient-validated reduced side effect profile have not yet reached the clinic. GR is a member of the nuclear receptor family of transcription factors and heavily relies on interactions with coregulator proteins for its transcriptional activity. To elucidate the role of the GR interactome in the altered transcriptional activity of GR following treatment with agonists, antagonists or lead selective GR agonists and modulators (SEGRAMs), we generated comprehensive interactome maps by high-confidence proximity proteomics in lung epithelial carcinoma cells. We found that the GR antagonist RU486 and the SEGRAM Dagrocorat both reduced GR interaction with CREB-binding protein (CBP)/p300 and the mediator complex compared to the full agonist Dexamethasone. Our data offer new insights into the role differential coregulator protein recruitment in shaping specific GR-mediated transcriptional responses.
Project description:How steroid hormone receptors (SHRs) orchestrate transcriptional activity remains only partly understood. Upon activation, SHRs bind the genome and recruit their co-regulators, crucial to induce gene expression. However, it remains unknown which components of the SHR-recruited coregulator complex are essential to drive transcription following hormonal stimuli. Through a FACS-based genome-wide CRISPR screen, we comprehensively dissected the Glucocorticoid Receptor (GR) co-regulatory complex involved in gene-target regulation. We describe a novel functional cross-talk between PAXIP1 and the cohesin subunit STAG2 that is critical for regulation of gene expression by GR. Without altering the GR cistrome, PAXIP1 and STAG2 depletion alter the GR transcriptome, by impairing the recruitment of 3D-genome organization proteins to the GR complex. Importantly, we demonstrate that PAXIP1 is required for stability of cohesin on the genome, its localization to GR-occupied sites, and maintenance of enhancer-promoter interactions. Moreover, in lung cancer, where GR acts as tumor suppressor, PAXIP1/STAG2 loss enhances GR-mediated tumor suppressor activity by modifying local chromatin interactions. All together, we introduce PAXIP1 and STAG2 as novel co-regulators of GR, required to maintain 3D-genome architecture and drive the GR transcriptional programme following hormonal stimuli.
Project description:Glucocorticoid receptor (GR) is an essential transcription factor (TF), controlling metabolism, development and immune responses. SUMOylation regulates chromatin occupancy and target gene expression of GR in a locus-selective manner, but the mechanism of regulation has remained elusive. Here, we show using selective isolation of chromatin-associated proteins that the protein network around chromatin-bound GR is affected by SUMOylation, with several nuclear receptor coregulators and chromatin modifiers being more avidly associated with SUMOylation-deficient than SUMOylation competent GR. This difference is reflected in our chromatin accessibility and gene expression data, showing that the SUMOylation-deficient GR is more potent in opening chromatin at glucocorticoid-regulated enhancers and inducing expression of their target loci. Our results thus show that SUMOylation determines GR specificity by regulating the chromatin protein network and accessibility at GR-driven enhancers. We speculate that a similar mechanism is utilized by many other SUMOylated TFs.
Project description:Pro35SLBD16:GR or Pro35SLBD18:GR transgenic seedlings that overexpress LBD16 or LBD18 fused to glucocorticoidsteroid hormone binding domain(GR) under CaMV35S promoter were grown for 12 days under long-day conditions (16h light/ 8h dark).