Associations between promoter usage and alternative splicing of the glucocorticoid receptor gene (original) (raw)
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The Human Glucocorticoid Receptor Isoform: EXPRESSION, BIOCHEMICAL PROPERTIES, AND PUTATIVE FUNCTION
Carolina Digital Repository (University of North Carolina at Chapel Hill), 1996
Alternative splicing of the human glucocorticoid receptor (hGR) primary transcript produces two receptor isoforms, hGR␣ and hGR, which differ at their carboxyl termini. The hGR␣ isoform conveys endocrine information to target tissues by altering patterns of gene expression in a hormone-dependent fashion. In contrast to hGR␣, very little is known about the hGR splice variant. Using hGR␣and hGR-specific riboprobes on human multiple tissue Northern blots, we show that the hGR message has a widespread tissue distribution. We also prove by reverse transcriptase-polymerase chain reaction that the alternative splicing event underlying the formation of the hGR message occurs in these tissues. Because the hGR protein differs from hGR␣ at the extreme COOH terminus, we investigated several of the biochemical properties of hGR expressed in transfected cells. hGR does not bind the glucocorticoid agonist dexamethasone nor the glucocorticoid antagonist RU38486 in vivo. Moreover, in contrast to hGR␣, hGR is located primarily in the nucleus of transfected cells independent of hormone administration. Finally, in the absence of hGR␣, hGR is transcriptionally inactive on a glucocorticoid-responsive enhancer. However, when both isoforms are expressed in the same cell, hGR inhibits the hormone-induced, hGR␣-mediated stimulation of gene expression. Thus, hGR potentially functions as a dominant negative inhibitor of hGR␣ activity.
Expression of glucocorticoid receptor α- and β-isoforms in human cells and tissues
American Journal of Physiology-Cell Physiology, 2002
Alternative splicing of the human glucocorticoid receptor (GR) primary transcript generates two protein isoforms: GR-α and GR-β. We investigated the expression of both GR isoforms in healthy human cells and tissues. GR-α mRNA abundance (×106cDNA copies/μg total RNA) was as follows: brain (3.83 ± 0.80) > skeletal muscle > macrophages > lung > kidney > liver > heart > eosinophils > peripheral blood mononuclear cells (PBMCs) > nasal mucosa > neutrophils > colon (0.33 ± 0.04). GR-β mRNA was much less expressed than GR-α mRNA. Its abundance (×103cDNA copies/μg total RNA) was as follows: eosinophils (1.55 ± 0.58) > PBMCs > liver ≥ skeletal muscle > kidney > macrophages > lung > neutrophils > brain ≥ nasal mucosa > heart (0.15 ± 0.08). GR-β mRNA was not found in colon. While GR-α protein was detected in all cells and tissues, GR-β was not detected in any specimen. Our results suggest that, in physiological conditions, the default spl...
Glucocorticoid receptor isoforms generate transcription specificity
Trends in Cell Biology, 2006
Glucocorticoids are necessary for life and are essential in all aspects of health and disease as they regulate processes from mitosis to apoptosis, from metabolism to growth and development. However, responses to glucocorticoids vary among individuals, cells and tissues. Recent evidence indicates that multiple glucocorticoid receptor (GR) isoforms are generated from one single GR gene by alternative splicing and alternative translation initiation. These isoforms all have unique tissue distribution patterns and transcriptional regulatory profiles. Furthermore, each is subject to various post-translational modifications that affect receptor function. Thus, increasing evidence suggests that unique GR isoform compositions within cells could determine the cell-specific response to glucocorticoids. Here, we discuss a new molecular model potentially underlying tissue-specific glucocorticoid resistance and selectivity.
Characterization of the Human Glucocorticoid Receptor Promoter
Biochemistry, 1995
To elucidate the functional elements that are involved in the regulation of the human glucocorticoid receptor (hGR) gene, transient expression, DNase I footprinting, and gel mobility shift analyses were conducted. We found that the hGR promoter region between-700 and +38 bp contained 1 1 footprinted sites. Deletion of the-374 to-183 bp region, which is highly conserved between human and mouse (93%), induced a 5-24-fold reduction in promoter activity in HeLa, NIH3T3, CV1, and HepG2 cells. Three footprints, FP5, FP6, and FP7, were shown to map to this region. In particular, the FP7 site was found to be within the-374 to-347 bp region. Deletion of this region triggered a significant decline in promoter activity in HeLa and NIH3T3 cells but not in HepG2 cells. AP2 was found to bind FP7. In HepG2 cells AP2 elicited transactivation of the hGR promoter activity. Transfection data revealed that the upstream GC box-rich fragment between-700 and-375 bp induced a 4-7-fold activation of the heterologous tk promoter in an orientation-independent manner. Our studies demonstrate that several transcription factors are involved in regulating GR expression and that AP2 could function as an important positive regulator of GR promoter activity.
Steroids, 2013
Glucocorticoids are important regulators of metabolism and immune function. Synthetic glucocorticoids are extensively used for immunosuppression/anti-inflammatory therapy. Since the glucocorticoid receptor (GR) is central to most hormone effects; its in vivo regulation will influence hormone/drug action. An alternative splice variant, GRb, is present in humans and may function as a dominant negative regulator of GR transcriptional activity. Recently, a similar splice variant was reported in mouse, although the mechanism of alternative splicing differs from that in humans. We present evidence that a splice variant of GR with an alternative C-terminus also occurs in the rat by a mechanism of intron inclusion. A highly quantitative qRT-PCR assay for the simultaneous measurement of both splice variants in a single sample was developed in order to accurately measure their regulation. We used this assay to assess the tissue specific expression of both mRNAs, and demonstrate that GRa is predominant in all tissues. In addition, the regulation of both GRa and GRb mRNA by various physiological factors in rat liver was assessed. GRa showed a robust circadian rhythm, which was entrained with the circadian oscillation of the endogenous hormone. Time series experiments showed that both corticosteroids and LPS but not insulin dosing resulted in the transient down-regulation of GRa mRNA. LPS treatment also resulted in down-regulation of GRb expression. A modest up-regulation in GRb expression was observed only in animals having chronically elevated plasma insulin concentrations. However the expression of GRb was significantly lower than that of GRa in all cases.
The human glucocorticoid receptor: One gene, multiple proteins and diverse responses
Steroids, 2005
Glucocorticoids are a vital class of endogenous steroid hormones that regulate essential biological processes including growth, development, metabolism, behavior and apoptosis. Most, if not all, of these actions are thought to be mediated through the glucocorticoid receptor. The exact mechanisms of how one hormone, via one receptor, modulates such diverse biological functions are largely unknown. However, recent studies from our lab and others have suggested that a contribution for the diversity results from multiple isoforms of the glucocorticoid receptor that result from alternative RNA splicing and translation initiation of the glucocorticoid receptor mRNA. Additionally, each isoform is subject to several post-translational modifications, including phosphorylation, ubiquitination and sumoylation, which have been shown to modulate the receptor protein stability and/or function. Together these data provide potentially diverse mechanisms to establish cell type specific regulation of gene expression by a single transcription factor. Here, we summarize the recent advances and processes that generate these receptor isoforms and these post-translational modifications. We speculate that the composition and proportion of individual isoforms expressed in particular cellular contexts account for the diverse effects of glucocorticoid hormones.
The Human Glucocorticoid Receptor β Isoform
Journal of Biological Chemistry, 1996
Alternative splicing of the human glucocorticoid receptor (hGR) primary transcript produces two receptor isoforms, hGR␣ and hGR, which differ at their carboxyl termini. The hGR␣ isoform conveys endocrine information to target tissues by altering patterns of gene expression in a hormone-dependent fashion. In contrast to hGR␣, very little is known about the hGR splice variant. Using hGR␣and hGR-specific riboprobes on human multiple tissue Northern blots, we show that the hGR message has a widespread tissue distribution. We also prove by reverse transcriptase-polymerase chain reaction that the alternative splicing event underlying the formation of the hGR message occurs in these tissues. Because the hGR protein differs from hGR␣ at the extreme COOH terminus, we investigated several of the biochemical properties of hGR expressed in transfected cells. hGR does not bind the glucocorticoid agonist dexamethasone nor the glucocorticoid antagonist RU38486 in vivo. Moreover, in contrast to hGR␣, hGR is located primarily in the nucleus of transfected cells independent of hormone administration. Finally, in the absence of hGR␣, hGR is transcriptionally inactive on a glucocorticoid-responsive enhancer. However, when both isoforms are expressed in the same cell, hGR inhibits the hormone-induced, hGR␣-mediated stimulation of gene expression. Thus, hGR potentially functions as a dominant negative inhibitor of hGR␣ activity.
Human glucocorticoid receptor gene promotor—homologous down regulation
The Journal of Steroid Biochemistry and Molecular Biology, 1991
To study the regulation of the human glucocorticoid receptor (hGR), we characterized the promoter region by primer extension, Sl nuclease mapping and by DNA sequencing. We found that the promoter is extremely G + C rich (72% GC content) and contains a "TAATA" and a "CAT" box, eight "GGGCGG", three "CCGCCC" and two "CACCC" motifs and a motif similar to the glucocorticoid responsive element (GRE) which included two interchanged nucleotides "TCTTGT'. In contrast to other steroid receptor genes, exon I or GHGR contains the major part of the 5' non-coding sequences of hGR mRNA while exon II contains coding sequences for the first 394 amino acid residues of the A/B region of hGR. The major transcriptional start site was found to be 134 bp upstream of the ATG initiation codon. Transfection of HeLa cells with plasmids containing various deletions of GHGR promoter fused to a promoterless CAT vector suggested the region between-470 and-1030, at the 5' end of the mRNA start site, to contain sequences required for down regulation by hormone.