Project description:Glucocorticoids are primary stress hormones and their synthetic derivatives are widely used clinically. The therapeutic efficacy of these steroids is limited by severe side effects and glucocorticoid resistance. Multiple glucocorticoid receptor (GR) isoforms are produced by alternative translation initiation; however, the role individual isoforms play in controlling tissue-specific responses to glucocorticoids in vivo is unknown. We have generated knockin mice that exclusively express the most active receptor isoform, GR-C3. The GR-C3 knockin mice die at birth due to respiratory distress but can be completely rescued by antenatal glucocorticoid administration. To evaluate the GR-C3 transcriptome, we prepared mouse embryonic fibroblasts (MEFs) from E12.5 wild-type (WT) and GR-C3 knockin embryos and treated the cells with vehicle or the synthetic glucocorticoid Dexamethasone (Dex) for 6 hours. The GR-C3 isoform was found to have a markedly different gene-regulatory profile than GR in WT MEFs.
Project description:Glucocorticoids are primary stress hormones and their synthetic derivatives are widely used clinically. The therapeutic efficacy of these steroids is limited by severe side effects and glucocorticoid resistance. Multiple glucocorticoid receptor (GR) isoforms are produced by alternative translation initiation; however, the role individual isoforms play in controlling tissue-specific responses to glucocorticoids in vivo is unknown. We have generated knockin mice that exclusively express the most active receptor isoform, GR-C3. The GR-C3 knockin mice die at birth due to respiratory distress but can be completely rescued by antenatal glucocorticoid administration. Rescued GR-C3 mice exhibited alterations in circadian rhythm in a sexually dimorphic manner and in sensitivity to lipopolysaccharide (LPS)-induced endotoxemia. To evaluate the ability of glucocorticoids to protect against LPS-induced inflammation, we measured gene expression in spleens from WT and rescued GR-C3 knockin mice that had been treated with vehicle or LPS for 3 and 24 hours. The GR-C3 isoform was found to be deficient in its ability to repress a large cohort of immune and inflammatory genes.
Project description:We performed RNA sequencing in Mouse Embryonic Fibroblasts (MEFs) of GR wild-type and GR Zn. Cells were treated with: vehicle (EtOH), Dexamethasone (Dex), Lipopolysaccharide (LPS) or Dex+LPS. We show that GR Zn does not regulate any GR target gene
Project description:Transcriptional profiling of mouse primary embryonic fibroblasts (MEFs) from wild type (WT) and knockin littermates expressing STAT1F77A (KI). For both genotypes, untreated control cells were compared to cells treated with IFN-alpha or IFN-gamma.
Project description:We performed ChIP-Sequencing for GR in GR wild-type, GR Zn mutant and GR Knock-Out Mouse Embryonic Fibroblasts (MEFs). We show that GR Zn mutant chromatin binding is mostly lost except for some tethered binding sites. Together with the corresponding RNA-Seq data, our results illustrate how GR requires direct DNA binding for activation and repression of target genes.
Project description:The Glucocorticoid Receptor (GR) co-ordinates metabolic and behavioural responses to stressors. We hypothesised that GR influences behaviour by modulating epigenetic and transcriptional processes in the brain. Using the zebrafish as a model organism, the brain methylomes of wild-type and grs357 mutant adults were analysed and GR-sensitive, differentially methylated regions (GR-DMRs) were identified. DMRs linked to genes with biologically salient functions were selected for confirmatory deep sequencing analysis in wild-type and GR mutant 5dpf larval heads, using the BisPCR2 technique (Bernstein et al., 2015). Our results demonstrate that the genes fkbp5 and aplp1 exemplify two distinct modes of GR-regulated, glucocorticoid-responsive DNA methylation in the brain.
Project description:To identify the sequences responsible for recruitment of Glucocorticoid receptor (GR) to individual loci, we performed ChIP-seq in four cell lines : A549 (ATTC:CCL-185), Nalm-6 (ATCC:CRL-1567), immortalized mouse embryonic fibroblasts (MEFs)(PMID 21131905), and immortalized PCAF-/-; GCN5flox/ MEFs (PMID 21131905) upon glucocorticoid treatment (1.5 hrs, 1M dexamethasone).
Project description:The objective of this study was to dertemine GR binding patterns within wild type murine embryonic fibroblasts in comparison to embryonic fibroblasts derived from mice containing a null mutation for Klf15.
Project description:Transcriptional profiling of mouse primary embryonic fibroblasts (MEFs) from wild type (WT) and knockin littermates expressing STAT1F77A (KI). For both genotypes, untreated control cells were compared to cells treated with IFN-alpha or IFN-gamma. Two condition experiments; untreated versus IFN treated
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.
