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Deploey N, Van Moortel L, Rogatsky I, Peelman F, De Bosscher K. The Biologist's Guide to the Glucocorticoid Receptor's Structure. Cells 2023; 12:1636. [PMID: 37371105 PMCID: PMC10297449 DOI: 10.3390/cells12121636] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/09/2023] [Revised: 06/07/2023] [Accepted: 06/09/2023] [Indexed: 06/29/2023] Open
Abstract
The glucocorticoid receptor α (GRα) is a member of the nuclear receptor superfamily and functions as a glucocorticoid (GC)-responsive transcription factor. GR can halt inflammation and kill off cancer cells, thus explaining the widespread use of glucocorticoids in the clinic. However, side effects and therapy resistance limit GR's therapeutic potential, emphasizing the importance of resolving all of GR's context-specific action mechanisms. Fortunately, the understanding of GR structure, conformation, and stoichiometry in the different GR-controlled biological pathways is now gradually increasing. This information will be crucial to close knowledge gaps on GR function. In this review, we focus on the various domains and mechanisms of action of GR, all from a structural perspective.
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Affiliation(s)
- Nick Deploey
- VIB Center for Medical Biotechnology, VIB, 9052 Ghent, Belgium; (N.D.); (L.V.M.); (F.P.)
- Department of Biomolecular Medicine, Ghent University, 9000 Ghent, Belgium
- Translational Nuclear Receptor Research (TNRR) Laboratory, VIB, 9052 Ghent, Belgium
| | - Laura Van Moortel
- VIB Center for Medical Biotechnology, VIB, 9052 Ghent, Belgium; (N.D.); (L.V.M.); (F.P.)
- Department of Biomolecular Medicine, Ghent University, 9000 Ghent, Belgium
- Translational Nuclear Receptor Research (TNRR) Laboratory, VIB, 9052 Ghent, Belgium
| | - Inez Rogatsky
- Hospital for Special Surgery Research Institute, The David Z. Rosensweig Genomics Center, New York, NY 10021, USA;
- Graduate Program in Immunology and Microbial Pathogenesis, Weill Cornell Graduate School of Medical Sciences, New York, NY 10065, USA
| | - Frank Peelman
- VIB Center for Medical Biotechnology, VIB, 9052 Ghent, Belgium; (N.D.); (L.V.M.); (F.P.)
- Department of Biomolecular Medicine, Ghent University, 9000 Ghent, Belgium
| | - Karolien De Bosscher
- VIB Center for Medical Biotechnology, VIB, 9052 Ghent, Belgium; (N.D.); (L.V.M.); (F.P.)
- Department of Biomolecular Medicine, Ghent University, 9000 Ghent, Belgium
- Translational Nuclear Receptor Research (TNRR) Laboratory, VIB, 9052 Ghent, Belgium
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2
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Parsonnet NV, Lammer NC, Holmes ZE, Batey RT, Wuttke DS. The glucocorticoid receptor DNA-binding domain recognizes RNA hairpin structures with high affinity. Nucleic Acids Res 2019; 47:8180-8192. [PMID: 31147715 DOI: 10.1093/nar/gkz486] [Citation(s) in RCA: 26] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/13/2019] [Revised: 05/17/2019] [Accepted: 05/21/2019] [Indexed: 01/04/2023] Open
Abstract
The glucocorticoid receptor (GR) binds the noncoding RNA Gas5 via its DNA-binding domain (DBD) with functional implications in pro-apoptosis signaling. Here, we report a comprehensive in vitro binding study where we have determined that GR-DBD is a robust structure-specific RNA-binding domain. GR-DBD binds to a diverse range of RNA hairpin motifs, both synthetic and biologically derived, with apparent mid-nanomolar affinity while discriminating against uniform dsRNA. As opposed to dimeric recognition of dsDNA, GR-DBD binds to RNA as a monomer and confers high affinity primarily through electrostatic contacts. GR-DBD adopts a discrete RNA-bound state, as assessed by NMR, distinct from both free and DNA-bound. NMR and alanine mutagenesis suggest a heightened involvement of the C-terminal α-helix of the GR-DBD in RNA-binding. RNA competes for binding with dsDNA and occurs in a similar affinity range as dimer binding to the canonical DNA element. Given the prevalence of RNA hairpins within the transcriptome, our findings strongly suggest that many RNAs have potential to impact GR biology.
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Affiliation(s)
- Nicholas V Parsonnet
- Department of Biochemistry, University of Colorado at Boulder, Campus Box 596, Boulder, CO 80309-0596, USA
| | - Nickolaus C Lammer
- Department of Biochemistry, University of Colorado at Boulder, Campus Box 596, Boulder, CO 80309-0596, USA
| | - Zachariah E Holmes
- Department of Biochemistry, University of Colorado at Boulder, Campus Box 596, Boulder, CO 80309-0596, USA
| | - Robert T Batey
- Department of Biochemistry, University of Colorado at Boulder, Campus Box 596, Boulder, CO 80309-0596, USA
| | - Deborah S Wuttke
- Department of Biochemistry, University of Colorado at Boulder, Campus Box 596, Boulder, CO 80309-0596, USA
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3
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Escoter-Torres L, Caratti G, Mechtidou A, Tuckermann J, Uhlenhaut NH, Vettorazzi S. Fighting the Fire: Mechanisms of Inflammatory Gene Regulation by the Glucocorticoid Receptor. Front Immunol 2019; 10:1859. [PMID: 31440248 PMCID: PMC6693390 DOI: 10.3389/fimmu.2019.01859] [Citation(s) in RCA: 83] [Impact Index Per Article: 13.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/15/2019] [Accepted: 07/23/2019] [Indexed: 12/12/2022] Open
Abstract
For many decades, glucocorticoids have been widely used as the gold standard treatment for inflammatory conditions. Unfortunately, their clinical use is limited by severe adverse effects such as insulin resistance, cardiometabolic diseases, muscle and skin atrophies, osteoporosis, and depression. Glucocorticoids exert their effects by binding to the Glucocorticoid Receptor (GR), a ligand-activated transcription factor which both positively, and negatively regulates gene expression. Extensive research during the past several years has uncovered novel mechanisms by which the GR activates and represses its target genes. Genome-wide studies and mouse models have provided valuable insight into the molecular mechanisms of inflammatory gene regulation by GR. This review focusses on newly identified target genes and GR co-regulators that are important for its anti-inflammatory effects in innate immune cells, as well as mutations within the GR itself that shed light on its transcriptional activity. This research progress will hopefully serve as the basis for the development of safer immune suppressants with reduced side effect profiles.
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Affiliation(s)
- Laura Escoter-Torres
- Molecular Endocrinology, Helmholtz Zentrum München (HMGU), German Center for Diabetes Research (DZD), Institute for Diabetes and Cancer IDC, Munich, Germany
| | - Giorgio Caratti
- Department of Biology, Institute for Comparative Molecular Endocrinology, University of Ulm, Ulm, Germany
| | - Aikaterini Mechtidou
- Molecular Endocrinology, Helmholtz Zentrum München (HMGU), German Center for Diabetes Research (DZD), Institute for Diabetes and Cancer IDC, Munich, Germany
| | - Jan Tuckermann
- Department of Biology, Institute for Comparative Molecular Endocrinology, University of Ulm, Ulm, Germany
| | - Nina Henriette Uhlenhaut
- Molecular Endocrinology, Helmholtz Zentrum München (HMGU), German Center for Diabetes Research (DZD), Institute for Diabetes and Cancer IDC, Munich, Germany.,Gene Center, Ludwig-Maximilians-Universität (LMU), Munich, Germany
| | - Sabine Vettorazzi
- Department of Biology, Institute for Comparative Molecular Endocrinology, University of Ulm, Ulm, Germany
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4
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Desgeorges T, Caratti G, Mounier R, Tuckermann J, Chazaud B. Glucocorticoids Shape Macrophage Phenotype for Tissue Repair. Front Immunol 2019; 10:1591. [PMID: 31354730 PMCID: PMC6632423 DOI: 10.3389/fimmu.2019.01591] [Citation(s) in RCA: 71] [Impact Index Per Article: 11.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/16/2019] [Accepted: 06/25/2019] [Indexed: 12/16/2022] Open
Abstract
Inflammation is a complex process which is highly conserved among species. Inflammation occurs in response to injury, infection, and cancer, as an allostatic mechanism to return the tissue and to return the organism back to health and homeostasis. Excessive, or chronic inflammation is associated with numerous diseases, and thus strategies to combat run-away inflammation is required. Anti-inflammatory drugs were therefore developed to switch inflammation off. However, the inflammatory response may be beneficial for the organism, in particular in the case of sterile tissue injury. The inflammatory response can be divided into several parts. The first step is the mounting of the inflammatory reaction itself, characterized by the presence of pro-inflammatory cytokines, and the infiltration of immune cells into the injured area. The second step is the resolution phase, where immune cells move toward an anti-inflammatory phenotype and decrease the secretion of pro-inflammatory cytokines. The last stage of inflammation is the regeneration process, where the tissue is rebuilt. Innate immune cells are major actors in the inflammatory response, of which, macrophages play an important role. Macrophages are highly sensitive to a large number of environmental stimuli, and can adapt their phenotype and function on demand. This change in phenotype in response to the environment allow macrophages to be involved in all steps of inflammation, from the first mounting of the pro-inflammatory response to the post-damage tissue repair.
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Affiliation(s)
- Thibaut Desgeorges
- Institut NeuroMyoGène, Université Claude Bernard Lyon 1, Univ Lyon, CNRS UMR 5310, INSERM U1217, Lyon, France
| | - Giorgio Caratti
- Institute of Comparative Molecular Endocrinology, University of Ulm, Ulm, Germany
| | - Rémi Mounier
- Institut NeuroMyoGène, Université Claude Bernard Lyon 1, Univ Lyon, CNRS UMR 5310, INSERM U1217, Lyon, France
| | - Jan Tuckermann
- Institute of Comparative Molecular Endocrinology, University of Ulm, Ulm, Germany
| | - Bénédicte Chazaud
- Institut NeuroMyoGène, Université Claude Bernard Lyon 1, Univ Lyon, CNRS UMR 5310, INSERM U1217, Lyon, France
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5
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Weikum ER, de Vera IMS, Nwachukwu JC, Hudson WH, Nettles KW, Kojetin DJ, Ortlund EA. Tethering not required: the glucocorticoid receptor binds directly to activator protein-1 recognition motifs to repress inflammatory genes. Nucleic Acids Res 2017; 45:8596-8608. [PMID: 28591827 PMCID: PMC5737878 DOI: 10.1093/nar/gkx509] [Citation(s) in RCA: 47] [Impact Index Per Article: 5.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/21/2017] [Accepted: 06/05/2017] [Indexed: 12/22/2022] Open
Abstract
The glucocorticoid receptor (GR) is a ligand-regulated transcription factor that controls the expression of extensive gene networks, driving both up- and down-regulation. GR utilizes multiple DNA-binding-dependent and -independent mechanisms to achieve context-specific transcriptional outcomes. The DNA-binding-independent mechanism involves tethering of GR to the pro-inflammatory transcription factor activator protein-1 (AP-1) through protein-protein interactions. This mechanism has served as the predominant model of GR-mediated transrepression of inflammatory genes. However, ChIP-seq data have consistently shown GR to occupy AP-1 response elements (TREs), even in the absence of AP-1. Therefore, the current model is insufficient to explain GR action at these sites. Here, we show that GR regulates a subset of inflammatory genes in a DNA-binding-dependent manner. Using structural biology and biochemical approaches, we show that GR binds directly to TREs via sequence-specific contacts to a GR-binding sequence (GBS) half-site found embedded within the TRE motif. Furthermore, we show that GR-mediated transrepression observed at TRE sites to be DNA-binding-dependent. This represents a paradigm shift in the field, showing that GR uses multiple mechanisms to suppress inflammatory gene expression. This work further expands our understanding of this complex multifaceted transcription factor.
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Affiliation(s)
- Emily R Weikum
- Department of Biochemistry, Emory University School of Medicine, Atlanta, GA 30322, USA
| | - Ian Mitchelle S de Vera
- Department of Integrative Structural and Computational Biology, The Scripps Research Institute, Jupiter, FL 33458, USA
| | - Jerome C Nwachukwu
- Department of Integrative Structural and Computational Biology, The Scripps Research Institute, Jupiter, FL 33458, USA
| | - William H Hudson
- Department of Biochemistry, Emory University School of Medicine, Atlanta, GA 30322, USA
| | - Kendall W Nettles
- Department of Integrative Structural and Computational Biology, The Scripps Research Institute, Jupiter, FL 33458, USA
| | - Douglas J Kojetin
- Department of Integrative Structural and Computational Biology, The Scripps Research Institute, Jupiter, FL 33458, USA.,Department of Molecular Medicine, The Scripps Research Institute, Jupiter, FL 33458, USA
| | - Eric A Ortlund
- Department of Biochemistry, Emory University School of Medicine, Atlanta, GA 30322, USA
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6
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Weikum ER, Knuesel MT, Ortlund EA, Yamamoto KR. Glucocorticoid receptor control of transcription: precision and plasticity via allostery. Nat Rev Mol Cell Biol 2017; 18:159-174. [PMID: 28053348 PMCID: PMC6257982 DOI: 10.1038/nrm.2016.152] [Citation(s) in RCA: 362] [Impact Index Per Article: 45.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022]
Abstract
The glucocorticoid receptor (GR) is a constitutively expressed transcriptional regulatory factor (TRF) that controls many distinct gene networks, each uniquely determined by particular cellular and physiological contexts. The precision of GR-mediated responses seems to depend on combinatorial, context-specific assembly of GR-nucleated transcription regulatory complexes at genomic response elements. In turn, evidence suggests that context-driven plasticity is conferred by the integration of multiple signals, each serving as an allosteric effector of GR conformation, a key determinant of regulatory complex composition and activity. This structural and mechanistic perspective on GR regulatory specificity is likely to extend to other eukaryotic TRFs.
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Affiliation(s)
- Emily R Weikum
- Department of Biochemistry, Emory University School of Medicine, 1510 Clifton Road, Atlanta, Georgia 30322, USA
| | - Matthew T Knuesel
- Department of Cellular and Molecular Pharmacology, University of California San Francisco School of Medicine, 600 16th Street, San Francisco, California 94143, USA
| | - Eric A Ortlund
- Department of Biochemistry, Emory University School of Medicine, 1510 Clifton Road, Atlanta, Georgia 30322, USA
| | - Keith R Yamamoto
- Department of Cellular and Molecular Pharmacology, University of California San Francisco School of Medicine, 600 16th Street, San Francisco, California 94143, USA
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Dougherty EJ, Elinoff JM, Ferreyra GA, Hou A, Cai R, Sun J, Blaine KP, Wang S, Danner RL. Mineralocorticoid Receptor (MR) trans-Activation of Inflammatory AP-1 Signaling: DEPENDENCE ON DNA SEQUENCE, MR CONFORMATION, AND AP-1 FAMILY MEMBER EXPRESSION. J Biol Chem 2016; 291:23628-23644. [PMID: 27650495 DOI: 10.1074/jbc.m116.732248] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/15/2016] [Indexed: 01/21/2023] Open
Abstract
Glucocorticoids are commonly used to treat inflammatory disorders. The glucocorticoid receptor (GR) can tether to inflammatory transcription factor complexes, such as NFκB and AP-1, and trans-repress the transcription of cytokines, chemokines, and adhesion molecules. In contrast, aldosterone and the mineralocorticoid receptor (MR) primarily promote cardiovascular inflammation by incompletely understood mechanisms. Although MR has been shown to weakly repress NFκB, its role in modulating AP-1 has not been established. Here, the effects of GR and MR on NFκB and AP-1 signaling were directly compared using a variety of ligands, two different AP-1 consensus sequences, GR and MR DNA-binding domain mutants, and siRNA knockdown or overexpression of core AP-1 family members. Both GR and MR repressed an NFκB reporter without influencing p65 or p50 binding to DNA. Likewise, neither GR nor MR affected AP-1 binding, but repression or activation of AP-1 reporters occurred in a ligand-, AP-1 consensus sequence-, and AP-1 family member-specific manner. Notably, aldosterone interactions with both GR and MR demonstrated a potential to activate AP-1. DNA-binding domain mutations that eliminated the ability of GR and MR to cis-activate a hormone response element-driven reporter variably affected the strength and polarity of these responses. Importantly, MR modulation of NFκB and AP-1 signaling was consistent with a trans-mechanism, and AP-1 effects were confirmed for specific gene targets in primary human cells. Steroid nuclear receptor trans-effects on inflammatory signaling are context-dependent and influenced by nuclear receptor conformation, DNA sequence, and the expression of heterologous binding partners. Aldosterone activation of AP-1 may contribute to its proinflammatory effects in the vasculature.
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Affiliation(s)
- Edward J Dougherty
- From the Critical Care Medicine Department, Clinical Center, National Institutes of Health, Bethesda, Maryland 20892
| | - Jason M Elinoff
- From the Critical Care Medicine Department, Clinical Center, National Institutes of Health, Bethesda, Maryland 20892
| | - Gabriela A Ferreyra
- From the Critical Care Medicine Department, Clinical Center, National Institutes of Health, Bethesda, Maryland 20892
| | - Angela Hou
- From the Critical Care Medicine Department, Clinical Center, National Institutes of Health, Bethesda, Maryland 20892
| | - Rongman Cai
- From the Critical Care Medicine Department, Clinical Center, National Institutes of Health, Bethesda, Maryland 20892
| | - Junfeng Sun
- From the Critical Care Medicine Department, Clinical Center, National Institutes of Health, Bethesda, Maryland 20892
| | - Kevin P Blaine
- From the Critical Care Medicine Department, Clinical Center, National Institutes of Health, Bethesda, Maryland 20892
| | - Shuibang Wang
- From the Critical Care Medicine Department, Clinical Center, National Institutes of Health, Bethesda, Maryland 20892
| | - Robert L Danner
- From the Critical Care Medicine Department, Clinical Center, National Institutes of Health, Bethesda, Maryland 20892
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8
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Quetglas EG, Mujagic Z, Wigge S, Keszthelyi D, Wachten S, Masclee A, Reinisch W. Update on pathogenesis and predictors of response of therapeutic strategies used in inflammatory bowel disease. World J Gastroenterol 2015; 21:12519-12543. [PMID: 26640330 PMCID: PMC4658608 DOI: 10.3748/wjg.v21.i44.12519] [Citation(s) in RCA: 21] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/06/2015] [Accepted: 09/14/2015] [Indexed: 02/06/2023] Open
Abstract
The search for biomarkers that characterize specific aspects of inflammatory bowel disease (IBD), has received substantial interest in the past years and is moving forward rapidly with the help of modern technologies. Nevertheless, there is a direct demand to identify adequate biomarkers for predicting and evaluating therapeutic response to different therapies. In this subset, pharmacogenetics deserves more attention as part of the endeavor to provide personalized medicine. The ultimate goal in this area is the adjustment of medication for a patient’s specific genetic background and thereby to improve drug efficacy and safety rates. The aim of the following review is to utilize the latest knowledge on immunopathogenesis of IBD and update the findings on the field of Immunology and Genetics, to evaluate the response to the different therapies. In the present article, more than 400 publications were reviewed but finally 287 included based on design, reproducibility (or expectancy to be reproducible and translationable into humans) or already measured in humans. A few tests have shown clinical applicability. Other, i.e., genetic associations for the different therapies in IBD have not yet shown consistent or robust results. In the close future it is anticipated that this, cellular and genetic material, as well as the determination of biomarkers will be implemented in an integrated molecular diagnostic and prognostic approach to manage IBD patients.
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Frenkel B, White W, Tuckermann J. Glucocorticoid-Induced Osteoporosis. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 2015. [PMID: 26215995 DOI: 10.1007/978-1-4939-2895-8_8] [Citation(s) in RCA: 112] [Impact Index Per Article: 11.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/16/2022]
Abstract
Osteoporosis is among the most devastating side effects of glucocorticoid (GC) therapy for the management of inflammatory and auto-immune diseases. Evidence from both humans and mice indicate deleterious skeletal effects within weeks of pharmacological GC administration, both related and unrelated to a decrease in bone mineral density (BMD). Osteoclast numbers and bone resorption are also rapidly increased, and together with osteoblast inactivation and decreased bone formation, these changes lead the fastest loss in BMD during the initial disease phase. Bone resorption then decreases to sub-physiological levels, but persistent and severe inhibition of bone formation leads to further bone loss and progressively increased fracture risk, up to an order of magnitude higher than that observed in untreated individuals. Bone forming osteoblasts are thus considered the main culprits in GC-induced osteoporosis (GIO). Accordingly, we focus this review primarily on deleterious effects on osteoblasts: inhibition of cell replication and function and acceleration of apoptosis. Mediating these adverse effects, GCs target pivotal regulatory mechanisms that govern osteoblast growth, differentiation and survival. Specifically, GCs inhibit growth factor pathways, including Insulin Growth Factors, Growth Hormone, Hepatocyte Growth/Scatter Factor and IL6-type cytokines. They also inhibit downstream kinases, including PI3-kinase and the MAP kinase ERK, the latter attributable in part to direct transcriptional stimulation of MAP kinase phosphatase 1. Most importantly, however, GCs inhibit the Wnt signaling pathway, which plays a pivotal role in osteoblast replication, function and survival. They transcriptionally stimulate expression of Wnt inhibitors of both the Dkk and Sfrp families, and they induce reactive oxygen species (ROS), which result in loss of ß-catenin to ROS-activated FoxO transcription factors. Identification of dissociated GCs, which would suppress the immune system without causing osteoporosis, is proving more challenging than initially thought, and GIO is currently managed by co-treatment with bisphosphonates or PTH. These drugs, however, are not ideally suited for GIO. Future therapeutic approaches may aim at GC targets such as those mentioned above, or newly identified targets including the Notch pathway, the AP-1/Il11 axis and the osteoblast master regulator RUNX2.
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Affiliation(s)
- Baruch Frenkel
- Department of Orthopaedic Surgery, Keck School of Medicine, Institute for Genetic Medicine, University of Southern California, 2250 Alcazar Street, CSC-240, Los Angeles, CA, 90033, USA,
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Cuzzoni E, De Iudicibus S, Franca R, Stocco G, Lucafò M, Pelin M, Favretto D, Pasini A, Montini G, Decorti G. Glucocorticoid pharmacogenetics in pediatric idiopathic nephrotic syndrome. Pharmacogenomics 2015; 16:1631-1648. [PMID: 26419298 DOI: 10.2217/pgs.15.101] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/16/2022] Open
Abstract
Idiopathic nephrotic syndrome represents the most common type of primary glomerular disease in children: glucocorticoids (GCs) are the first-line therapy, even if considerable interindividual differences in their efficacy and side effects have been reported. Immunosuppressive and anti-inflammatory effects of these drugs are mainly due to the GC-mediated transcription regulation of pro- and anti-inflammatory genes. This mechanism of action is the result of a complex multistep pathway that involves the glucocorticoid receptor and several other proteins, encoded by polymorphic genes. Aim of this review is to highlight the current knowledge on genetic variants that could affect GC response, particularly focusing on children with idiopathic nephrotic syndrome.
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Affiliation(s)
- Eva Cuzzoni
- Graduate School in Reproduction & Developmental Sciences, University of Trieste, I-34127 Trieste, Italy
| | - Sara De Iudicibus
- Institute for Maternal & Child Health IRCCS Burlo Garofolo, I-34137 Trieste, Italy
| | - Raffaella Franca
- Institute for Maternal & Child Health IRCCS Burlo Garofolo, I-34137 Trieste, Italy
| | - Gabriele Stocco
- Department of Life Sciences, University of Trieste, I-34127 Trieste, Italy
| | - Marianna Lucafò
- Department of Medical, Surgical and Health Sciences, University of Trieste, I-34127 Trieste, Italy
| | - Marco Pelin
- Department of Life Sciences, University of Trieste, I-34127 Trieste, Italy
| | - Diego Favretto
- Institute for Maternal & Child Health IRCCS Burlo Garofolo, I-34137 Trieste, Italy
| | - Andrea Pasini
- Nephrology and Dialysis Unit, Department of Pediatrics, Azienda Ospedaliera Universitaria Sant'Orsola-Malpighi, I-40138 Bologna, Italy
| | - Giovanni Montini
- Pediatric Nephrology and Dialysis Unit, Department of Clinical Sciences and Community Health, University of Milan, Fondazione IRCCS Cà Granda, Ospedale Maggiore Policlinico, I-20122 Milano, Italy
| | - Giuliana Decorti
- Department of Life Sciences, University of Trieste, I-34127 Trieste, Italy
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11
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Yu Y, Deng Y, Lu BM, Liu YX, Li J, Bao JK. Green tea catechins: a fresh flavor to anticancer therapy. Apoptosis 2013; 19:1-18. [DOI: 10.1007/s10495-013-0908-5] [Citation(s) in RCA: 49] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
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12
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Roohk DJ, Mascharak S, Khambatta C, Leung H, Hellerstein M, Harris C. Dexamethasone-mediated changes in adipose triacylglycerol metabolism are exaggerated, not diminished, in the absence of a functional GR dimerization domain. Endocrinology 2013; 154:1528-39. [PMID: 23493372 PMCID: PMC3602623 DOI: 10.1210/en.2011-1047] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/06/2023]
Abstract
The glucocorticoid (GC) receptor (GR) has multiple effector mechanisms, including dimerization-mediated transactivation of target genes via DNA binding and transcriptional repression mediated by protein-protein interactions. Much attention has been focused on developing selective GR modulators that would dissociate adverse effects from therapeutic anti-inflammatory effects. The GR(dim/dim) mouse has a mutation in the dimerization domain of GR and has been shown to have attenuated transactivation with intact repression. To understand the role of GR dimerization-dependent targets in multiple tissues, we measured metabolic fluxes through several disease-relevant GC target pathways using heavy water labeling and mass spectrometry in wild-type and GR(dim/dim) mice administered the potent GC dexamethasone (DEX). Absolute triglyceride synthesis was increased in both wild-type and GR(dim/dim) mice by DEX in the inguinal and epididymal fat depots. GR(dim/dim) mice showed an exaggerated response to DEX in both depots. De novo lipogenesis was also greatly increased in both depots in response to DEX in GR(dim/dim), but not wild-type mice. In contrast, the inhibitory effect of DEX on bone and skin collagen synthesis rates was greater in wild-type compared with GR(dim/dim) mice. Wild-type mice were more sensitive to DEX-dependent decreases in insulin sensitivity than GR(dim/dim) mice. Wild-type and GR(dim/dim) mice were equally sensitive to DEX-dependent decreases in muscle protein synthesis. Chronic elevation of GCs in GR(dim/dim) mice results in severe runting and lethality. In conclusion, some metabolic effects of GC treatment are exaggerated in adipose tissue of GR(dim/dim) mice, suggesting that selective GR modulators based on dissociating GR transactivation from repression should be evaluated carefully.
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Affiliation(s)
- Donald J Roohk
- Department of Nutritional Science and Toxicology, University of California Berkeley, Berkeley, California 94720, USA
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13
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Chen DWC, Saha V, Liu JZ, Schwartz JM, Krstic-Demonacos M. Erg and AP-1 as determinants of glucocorticoid response in acute lymphoblastic leukemia. Oncogene 2012; 32:3039-48. [PMID: 22869147 DOI: 10.1038/onc.2012.321] [Citation(s) in RCA: 25] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/18/2022]
Abstract
Glucocorticoids (GCs) are among the most widely prescribed medications in clinical practice. The beneficial effects of GCs in acute lymphoblastic leukemia (ALL) are based on their ability to induce apoptosis, but the underlying transcriptional mechanisms remain poorly defined. Computational modeling has enormous potential in the understanding of biological processes such as apoptosis and the discovery of novel regulatory mechanisms. We here present an integrated analysis of gene expression kinetic profiles using microarrays from GC sensitive and resistant ALL cell lines and patients, including newly generated and previously published data sets available from the Gene Expression Omnibus. By applying time-series clustering analysis in the sensitive ALL CEM-C7-14 cells, we identified 358 differentially regulated genes that we classified into 15 kinetic profiles. We identified GC response element (GRE) sequences in 33 of the upregulated known or potential GC receptor (GR) targets. Comparative study of sensitive and resistant ALL showed distinct gene expression patterns and indicated unexpected similarities between sensitivity-restored and resistant ALL. We found that activator protein 1 (AP-1), Ets related gene (Erg) and GR pathways were differentially regulated in sensitive and resistant ALL. Erg protein levels were substantially higher in CEM-C1-15-resistant cells, c-Jun was significantly induced in sensitive cells, whereas c-Fos was expressed at low levels in both. c-Jun was recruited on the AP-1 site on the Bim promoter, whereas a transient Erg occupancy on the GR promoter was detected. Inhibition of Erg and activation of GR lead to increased apoptosis in both sensitive and resistant ALL. These novel findings significantly advance our understanding of GC sensitivity and can be used to improve therapy of leukemia.
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Affiliation(s)
- D W-C Chen
- Faculty of Life Sciences, The University of Manchester, Manchester, UK
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14
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Pawlak M, Lefebvre P, Staels B. General molecular biology and architecture of nuclear receptors. Curr Top Med Chem 2012; 12:486-504. [PMID: 22242852 PMCID: PMC3637177 DOI: 10.2174/156802612799436641] [Citation(s) in RCA: 103] [Impact Index Per Article: 7.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/30/2011] [Accepted: 11/22/2011] [Indexed: 12/12/2022]
Abstract
Nuclear receptors (NRs) regulate and coordinate multiple processes by integrating internal and external signals, thereby maintaining homeostasis in front of nutritional, behavioral and environmental challenges. NRs exhibit strong similarities in their structure and mode of action: by selective transcriptional activation or repression of cognate target genes, which can either be controlled through a direct, DNA binding-dependent mechanism or through crosstalk with other transcriptional regulators, NRs modulate the expression of gene clusters thus achieving coordinated tissue responses. Additionally, non genomic effects of NR ligands appear mediated by ill-defined mechanisms at the plasma membrane. These effects mediate potential therapeutic effects as small lipophilic molecule targets, and many efforts have been put in elucidating their precise mechanism of action and pathophysiological roles. Currently, numerous nuclear receptor ligand analogs are used in therapy or are tested in clinical trials against various diseases such as hypertriglyceridemia, atherosclerosis, diabetes, allergies and cancer and others.
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Affiliation(s)
- Michal Pawlak
- Récepteurs nucléaires, maladies cardiovasculaires et diabète
INSERM : U1011Institut Pasteur de LilleUniversité Lille II - Droit et santé1 rue du Prof Calmette 59019 Lille Cedex,FR
| | - Philippe Lefebvre
- Récepteurs nucléaires, maladies cardiovasculaires et diabète
INSERM : U1011Institut Pasteur de LilleUniversité Lille II - Droit et santé1 rue du Prof Calmette 59019 Lille Cedex,FR
| | - Bart Staels
- Récepteurs nucléaires, maladies cardiovasculaires et diabète
INSERM : U1011Institut Pasteur de LilleUniversité Lille II - Droit et santé1 rue du Prof Calmette 59019 Lille Cedex,FR
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15
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Iudicibus SD, Franca R, Martelossi S, Ventura A, Decorti G. Molecular mechanism of glucocorticoid resistance in inflammatory bowel disease. World J Gastroenterol 2011; 17:1095-1108. [PMID: 21448414 PMCID: PMC3063901 DOI: 10.3748/wjg.v17.i9.1095] [Citation(s) in RCA: 85] [Impact Index Per Article: 6.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/10/2010] [Revised: 12/21/2010] [Accepted: 12/28/2010] [Indexed: 02/06/2023] Open
Abstract
Natural and synthetic glucocorticoids (GCs) are widely employed in a number of inflammatory, autoimmune and neoplastic diseases, and, despite the introduction of novel therapies, remain the first-line treatment for inducing remission in moderate to severe active Crohn's disease and ulcerative colitis. Despite their extensive therapeutic use and the proven effectiveness, considerable clinical evidence of wide inter-individual differences in GC efficacy among patients has been reported, in particular when these agents are used in inflammatory diseases. In recent years, a detailed knowledge of the GC mechanism of action and of the genetic variants affecting GC activity at the molecular level has arisen from several studies. GCs interact with their cytoplasmic receptor, and are able to repress inflammatory gene expression through several distinct mechanisms. The glucocorticoid receptor (GR) is therefore crucial for the effects of these agents: mutations in the GR gene (NR3C1, nuclear receptor subfamily 3, group C, member 1) are the primary cause of a rare, inherited form of GC resistance; in addition, several polymorphisms of this gene have been described and associated with GC response and toxicity. However, the GR is not self-standing in the cell and the receptor-mediated functions are the result of a complex interplay of GR and many other cellular partners. The latter comprise several chaperonins of the large cooperative hetero-oligomeric complex that binds the hormone-free GR in the cytosol, and several factors involved in the transcriptional machinery and chromatin remodeling, that are critical for the hormonal control of target genes transcription in the nucleus. Furthermore, variants in the principal effectors of GCs (e.g. cytokines and their regulators) have also to be taken into account for a comprehensive evaluation of the variability in GC response. Polymorphisms in genes involved in the transport and/or metabolism of these hormones have also been suggested as other possible candidates of interest that could play a role in the observed inter-individual differences in efficacy and toxicity. The best-characterized example is the drug efflux pump P-glycoprotein, a membrane transporter that extrudes GCs from cells, thereby lowering their intracellular concentration. This protein is encoded by the ABCB1/MDR1 gene; this gene presents different known polymorphic sites that can influence its expression and function. This editorial reviews the current knowledge on this topic and underlines the role of genetics in predicting GC clinical response. The ambitious goal of pharmacogenomic studies is to adapt therapies to a patient's specific genetic background, thus improving on efficacy and safety rates.
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16
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Lisse TS, Hewison M, Adams JS. Hormone response element binding proteins: novel regulators of vitamin D and estrogen signaling. Steroids 2011; 76:331-9. [PMID: 21236284 PMCID: PMC3042887 DOI: 10.1016/j.steroids.2011.01.002] [Citation(s) in RCA: 31] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/01/2010] [Revised: 01/04/2011] [Accepted: 01/05/2011] [Indexed: 01/11/2023]
Abstract
Insights from vitamin D-resistant New World primates and their human homologues as models of natural and pathological insensitivity to sterol/steroid action have uncovered a family of novel intracellular vitamin D and estrogen regulatory proteins involved in hormone action. The proteins, known as "vitamin D or estrogen response element-binding proteins", behave as potent cis-acting, transdominant regulators to inhibit steroid receptor binding to DNA response elements and is responsible for vitamin D and estrogen resistances. This set of interactors belongs to the heterogeneous nuclear ribonucleoprotein (hnRNP) family of previously known pre-mRNA-interacting proteins. This review provides new insights into the mechanism by which these novel regulators of signaling and metabolism can act to regulate responses to vitamin D and estrogen. In addition the review also describes other molecules that are known to influence nuclear receptor signaling through interaction with hormone response elements.
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Affiliation(s)
- Thomas S Lisse
- Department of Orthopaedic Surgery and Molecular Biology Institute, David Geffen School of Medicine at UCLA, 615 Charles E. Young Drive South, Los Angeles, CA 90095, USA.
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17
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Patel R, Patel M, Tsai R, Lin V, Bookout AL, Zhang Y, Magomedova L, Li T, Chan JF, Budd C, Mangelsdorf DJ, Cummins CL. LXRβ is required for glucocorticoid-induced hyperglycemia and hepatosteatosis in mice. J Clin Invest 2010; 121:431-41. [PMID: 21123945 DOI: 10.1172/jci41681] [Citation(s) in RCA: 92] [Impact Index Per Article: 6.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/05/2009] [Accepted: 10/13/2010] [Indexed: 12/16/2022] Open
Abstract
Although widely prescribed for their potent antiinflammatory actions, glucocorticoid drugs (e.g., dexamethasone) cause undesirable side effects that are features of the metabolic syndrome, including hyperglycemia, fatty liver, insulin resistance, and type II diabetes. Liver x receptors (LXRs) are nuclear receptors that respond to cholesterol metabolites and regulate the expression of a subset of glucocorticoid target genes. Here, we show LXRβ is required to mediate many of the negative side effects of glucocorticoids. Mice lacking LXRβ (but not LXRα) were resistant to dexamethasone-induced hyperglycemia, hyperinsulinemia, and hepatic steatosis, but remained sensitive to dexamethasone-dependent repression of the immune system. In vivo, LXRα/β knockout mice demonstrated reduced dexamethasone-induced expression of the key hepatic gluconeogenic gene, phosphoenolpyruvate carboxykinase (PEPCK). In perfused liver and primary mouse hepatocytes, LXRβ was required for glucocorticoid-induced recruitment of the glucocorticoid receptor to the PEPCK promoter. These findings suggest a new avenue for the design of safer glucocorticoid drugs through a mechanism of selective glucocorticoid receptor transactivation.
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Affiliation(s)
- Rucha Patel
- Department of Pharmaceutical Sciences, University of Toronto, Toronto, Ontario, Canada
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18
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Jacques E, Semlali A, Boulet LP, Chakir J. AP-1 overexpression impairs corticosteroid inhibition of collagen production by fibroblasts isolated from asthmatic subjects. Am J Physiol Lung Cell Mol Physiol 2010; 299:L281-7. [PMID: 20543003 DOI: 10.1152/ajplung.00360.2009] [Citation(s) in RCA: 14] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022] Open
Abstract
Asthma is characterized by airway remodeling associated with an increase in the deposition of ECM proteins such as type I collagen. These components are mainly produced by fibroblasts. Inhaled corticosteroids are considered the cornerstone of asthma therapy. Despite substantial evidence as to the anti-inflammatory action of corticosteroids, their effect on controlling ECM protein deposition in the airways is not completely understood. This study determined the effect of dexamethasone (Dex) on collagen production by bronchial fibroblasts derived from asthmatic and healthy subjects. Expression of procollagen mRNA in fibroblasts from asthmatics and normal controls was determined by quantitative PCR. Regulation of the procollagen-alpha(1)I promoter was evaluated by transient transfections. Transforming growth factor-beta (TGF-beta) protein expression was determined by ELISA. Protein expression of glucocorticoid receptor (GR) and interaction with activator protein-1 (AP-1), a collagen regulatory transcription factor, was assessed by Western blots, coimmunoprecipitations, and EMSA. AP-1 overexpression was performed by transient transfection using c-Fos/c-Jun expression plasmids. Dex significantly downregulated procollagen production and promoter activity in normal fibroblasts but had no effect on asthmatic fibroblasts. AP-1 and GR interaction increased after Dex stimulation in asthmatic fibroblasts. AP-1 overexpression in control fibroblasts abrogated collagen gene response to Dex. These results show that Dex failed to reduce collagen production in fibroblasts from asthmatic subjects. This impaired response may be related to AP-1 overexpression in these cells.
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Affiliation(s)
- Eric Jacques
- Centre de Recherche, Institut Universitaire de Cardiologie et de Pneumologie, Sainte-Foy, Québec, Canada
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19
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Kim JJ, Sefton EC, Bulun SE. Progesterone receptor action in leiomyoma and endometrial cancer. PROGRESS IN MOLECULAR BIOLOGY AND TRANSLATIONAL SCIENCE 2009; 87:53-85. [PMID: 20374701 DOI: 10.1016/s1877-1173(09)87002-6] [Citation(s) in RCA: 15] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/14/2022]
Abstract
Progesterone is a key hormone in the regulation of uterine function. In the normal physiological context, progesterone is primarily involved in remodeling of the endometrium and maintaining a quiescent myometrium. When pathologies of the uterus develop, specifically, endometrial cancer and uterine leiomyoma, response to progesterone is usually altered. Progesterone acts through mainly two isoforms of the progesterone receptor (PR), PRA and PRB which have been reported to exhibit different transcriptional activities. Studies examining the expression and function of the PRs in the normal endometrium and myometrium as well as in endometrial cancer and uterine leiomyoma are summarized here. The clinical use of progestins and the transcriptional activity of the PR on genes specific to endometrial cancer and leiomyoma are described. An increased understanding of the differential expression of PRs and response to progesterone in these two diseases is critical in order to develop more efficient and targeted therapies.
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Affiliation(s)
- J Julie Kim
- Division of Reproductive Biology Research, Department of Obstetrics and Gynecology, Robert H. Lurie Comprehensive Cancer Center, Chicago, Illinois 60611, USA
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20
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Gronemeyer H, Bourguet W. Allosteric effects govern nuclear receptor action: DNA appears as a player. Sci Signal 2009; 2:pe34. [PMID: 19491383 DOI: 10.1126/scisignal.273pe34] [Citation(s) in RCA: 36] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/02/2022]
Abstract
Nuclear receptors (NRs) are a family of transcription factors that regulate cognate gene networks, resulting in profound physiological and pathophysiological changes. Dysfunctional NR signaling leads to proliferative, reproductive, and metabolic diseases such as cancer, infertility, obesity, or diabetes. Indeed, NR-based pharmaceuticals are among the most commonly used drugs. NRs function by communicating with the intracellular and extracellular environment, thereby both sensing and modulating the status of cells. They respond to incoming signals by orchestrating transcriptional as well as nongenomic effects. They do so through an ability to respond to various effectors, such as the cognate ligand, by allosteric structural alterations that are the basis of further signal propagation. A mechanism has now been revealed by which DNA could act as an allosteric effector to modulate glucocorticoid receptor activity. This is a new regulatory paradigm for NR action that may help to explain how a receptor fine-tunes its target gene network.
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Affiliation(s)
- Hinrich Gronemeyer
- Department of Cancer Biology, Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), BP 10142, 67404 Illkirch Cedex, CU de Strasbourg, France.
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21
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Rani CSS, Elango N, Wang SS, Kobayashi K, Strong R. Identification of an activator protein-1-like sequence as the glucocorticoid response element in the rat tyrosine hydroxylase gene. Mol Pharmacol 2009; 75:589-98. [PMID: 19060113 PMCID: PMC2645927 DOI: 10.1124/mol.108.051219] [Citation(s) in RCA: 25] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/07/2008] [Accepted: 12/04/2008] [Indexed: 12/21/2022] Open
Abstract
Glucocorticoids (GCs) generally stimulate gene transcription via consensus glucocorticoid response elements (GREs) located in the promoter region. To identify the GRE in the rat tyrosine hydroxylase (TH) gene promoter, we transiently transfected PC12 cells with a 9-kilobase (kb) TH promoter-luciferase (Luc) construct. Dexamethasone (Dex) stimulated Luc activity, which was abolished by mifepristone (RU486). Serial deletion mutations revealed a Dex-responsive 7-base pair (bp) sequence, TGACTAA, located at -5734 to -5728. Deletion of just these seven nucleotides from the 9-kb promoter completely abolished the Dex response and partially reduced the response to phorbol ester but not to forskolin. The Dex response was fully retained in a construct in which most of the 9-kb promoter was deleted, except for 100 bp around the -5.7-kb region, clearly identifying this 7-bp sequence as solely responsible for GC responsiveness. Conversely, deletion of the proximal cAMP-response element (-45/-38) or activator protein-1 (AP-1) (-207/-201) sites in the 9-kb promoter did not affect Dex and phorbol ester responses. A radiolabeled 25-bp promoter fragment bearing the 7-bp TH-GRE/AP-1 showed specific binding to PC12 nuclear proteins. Using antibodies against the glucocorticoid receptors and AP-1 family of proteins and primers for the TH-GRE/AP-1 region, we detected a specific DNA amplicon in a chromatin immunoprecipitation assay. This 7-bp TH-GRE/AP-1 sequence (TGACTAA) does not bear similarity to any known GRE but closely resembles the consensus AP-1 binding site, TGACTCA. Our studies describe for the first time a novel GRE/AP-1 site present in the TH gene promoter that is critical for glucocorticoid regulation of the TH gene.
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Affiliation(s)
- C S Sheela Rani
- Department of Pharmacology,Barshop Institute for Longevity and Aging Studies, University of Texas Health Science Center at San Antonio, San Antonio, Texas 78229, USA
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22
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Diefenbacher M, Sekula S, Heilbock C, Maier JV, Litfin M, van Dam H, Castellazzi M, Herrlich P, Kassel O. Restriction to Fos family members of Trip6-dependent coactivation and glucocorticoid receptor-dependent trans-repression of activator protein-1. Mol Endocrinol 2008; 22:1767-80. [PMID: 18535250 DOI: 10.1210/me.2007-0574] [Citation(s) in RCA: 49] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/14/2023] Open
Abstract
The term activator protein (AP)-1 describes homodimeric and heterodimeric transcription factors composed of members of the Jun, Fos, and cAMP response element-binding protein (CREB)/activating transcription factor (ATF) families of proteins. Distinct AP-1 dimers, for instance the prototypical c-Jun:c-Fos and c-Jun:ATF2 dimers, are differentially regulated by signaling pathways and bind related yet distinct response elements in the regulatory regions of AP-1 target genes. Little is known about the dimer-specific regulation of AP-1 activity at the promoter of its target genes. We have previously shown that nTrip6, the nuclear isoform of the LIM domain protein Trip6, acts as an AP-1 coactivator. Moreover, nTrip6 is an essential component of glucocorticoid receptor (GR)-mediated trans-repression of AP-1, in that it mediates the tethering of GR to the promoter-bound AP-1. We have now discovered a striking specificity of nTrip6 actions determined by the binding preference of its LIM domains. We show that nTrip6 interacts only with Fos family members. Consequently, nTrip6 is a selective coactivator for AP-1 dimers containing Fos. nTrip6 also assembles activated GR to c-Jun:c-Fos-driven promoters. Neither nTrip6 nor GR are recruited to a promoter occupied by c-Jun:ATF2. Thus, only Fos-containing dimers are trans-repressed by GR. Thus, the dimer composition of AP-1 determines the mechanism of both the positive and negative regulation of AP-1 transcriptional activity. Interestingly, on a second level of action, GR represses the increase in transcriptional activity of c-Jun:ATF2 induced by c-Jun N-terminal kinase (JNK)-dependent phosphorylation. This repression depends on GR-mediated induction of MAPK phosphatase 1 (MKP-1) expression, which results in c-Jun N-terminal kinase inactivation.
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Affiliation(s)
- Markus Diefenbacher
- Institut für Toxikologie und Genetik, Forschungszentrum Karlsruhe, Hermann-von-Helmholtz Platz 1, D- 76344 Eggenstein-Leopoldshafen, Germany
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23
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Chinenov Y, Rogatsky I. Glucocorticoids and the innate immune system: crosstalk with the toll-like receptor signaling network. Mol Cell Endocrinol 2007; 275:30-42. [PMID: 17576036 DOI: 10.1016/j.mce.2007.04.014] [Citation(s) in RCA: 83] [Impact Index Per Article: 4.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/15/2007] [Accepted: 04/28/2007] [Indexed: 02/07/2023]
Abstract
Toll-like receptors (TLRs) are responsible for the recognition of a variety of microbial pathogens and the initial induction of immune and inflammatory responses. These responses are normally restricted by the adrenally produced glucocorticoid hormones which provide a feedback mechanism to curb unabated inflammation. Glucocorticoids act through a ligand-dependent transcription factor-the glucocorticoid receptor (GR), which engages in a complex network of protein:protein and protein:DNA interactions ultimately activating or repressing target gene transcription. Not surprisingly, multiple mechanisms account for the glucocorticoid interference with TLR signaling including enhanced expression of the natural inhibitors of TLR pathways, direct repression of TLR-activated transcriptional regulators and cross-utilization of cofactors essential for both GR and TLR signaling. Here we discuss recent and unexpected examples of crosstalk between the two transcriptional networks and the emerging role of GR in the regulation of innate immunity.
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Affiliation(s)
- Yurii Chinenov
- Hospital for Special Surgery, Department of Microbiology & Immunology, Weill Medical College of Cornell University, 535 E70th Street, Research Building Room 425, New York, NY 10021,USA
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24
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Yao M, Denver RJ. Regulation of vertebrate corticotropin-releasing factor genes. Gen Comp Endocrinol 2007; 153:200-16. [PMID: 17382944 DOI: 10.1016/j.ygcen.2007.01.046] [Citation(s) in RCA: 65] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/28/2006] [Accepted: 01/21/2007] [Indexed: 11/17/2022]
Abstract
Developmental, physiological, and behavioral adjustments in response to environmental change are crucial for animal survival. In vertebrates, the neuroendocrine stress system, comprised of the hypothalamus, pituitary, and adrenal/interrenal glands (HPA/HPI axis) plays a central role in adaptive stress responses. Corticotropin-releasing factor (CRF) is the primary hypothalamic neurohormone regulating the HPA/HPI axis. CRF also functions as a neurotransmitter/neuromodulator in the limbic system and brain stem to coordinate endocrine, behavioral, and autonomic responses to stressors. Glucocorticoids, the end products of the HPA/HPI axis, cause feedback regulation at multiple levels of the stress axis, exerting direct and indirect actions on CRF neurons. The spatial expression patterns of CRF, and stressor-dependent CRF gene activation in the central nervous system (CNS) are evolutionarily conserved. This suggests conservation of the gene regulatory mechanisms that underlie tissue-specific and stressor-dependent CRF expression. Comparative genomic analysis showed that the proximal promoter regions of vertebrate CRF genes are highly conserved. Several cis regulatory elements and trans acting factors have been implicated in stressor-dependent CRF gene activation, including cyclic AMP response element binding protein (CREB), activator protein 1 (AP-1/Fos/Jun), and nerve growth factor induced gene B (NGFI-B). Glucocorticoids, acting through the glucocorticoid and mineralocorticoid receptors, either repress or promote CRF expression depending on physiological state and CNS region. In this review, we take a comparative/evolutionary approach to understand the physiological regulation of CRF gene expression. We also discuss evolutionarily conserved molecular mechanisms that operate at the level of CRF gene transcription.
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Affiliation(s)
- Meng Yao
- Department of Molecular, Cellular and Developmental Biology, 3065C Kraus Natural Science Building, The University of Michigan, Ann Arbor, MI 48109-1048, USA
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25
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Wu J, Bresnick EH. Glucocorticoid and growth factor synergism requirement for Notch4 chromatin domain activation. Mol Cell Biol 2007; 27:2411-22. [PMID: 17220278 PMCID: PMC1820485 DOI: 10.1128/mcb.02152-06] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
The Notch signaling pathway modulates cell fate in diverse contexts, including vascular development. Notch4 is selectively expressed in vascular endothelium and regulates vascular remodeling. The signal-dependent transcription factor activator protein 1 (AP-1) activates Notch4 transcription in endothelial cells, but other factors/signals that regulate Notch4 are largely unknown. We demonstrate that, unlike the established transrepression mechanism in which the glucocorticoid receptor (GR) antagonizes AP-1, AP-1 and GR synergistically activated Notch4 transcription in endothelial cells. Fibroblast growth factor 2 (FGF-2) and cortisol induced AP-1 and GR occupancy, respectively, at a Notch4 promoter composite response element consisting of an imperfect half-glucocorticoid response element and an AP-1 motif, which mediated signal-dependent activation. Analysis of Notch4 promoter complex assembly provided evidence that GR and AP-1 independently occupy the composite response element, but AP-1 stabilizes GR occupancy. In multipotent 10T1/2 cells, FGF-2 and cortisol induced a histone modification pattern at the Notch4 locus mimicking that present in endothelial cells and reprogrammed Notch4 from a repressed to an active state. These results establish the molecular basis for a novel AP-1/GR-Notch4 axis in vascular endothelium.
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Affiliation(s)
- Jing Wu
- Department of Pharmacology, University of Wisconsin School of Medicine, Madison, WI 53706, USA
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26
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Abstract
The therapeutic and prophylactic use of glucocorticoids is widespread due to their powerful anti-inflammatory, antiproliferative and immunomodulatory activity. However, long-term use of these drugs can result in severe dose-limiting side effects. One of the most critical and debilitating side effects is osteoporosis, which leads to increased risk of fractures. Glucocorticoids damage bone through several different mechanisms. The search for novel glucocorticoids that have reduced side effects in bone and other tissues is being driven by the identification of new mechanisms of action of the glucocorticoid receptor. This may facilitate the detection of new, safer therapies with efficacies equivalent to currently prescribed steroids.
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Affiliation(s)
- Jeffrey N Miner
- Department of Molecular and Cell Biology, Ligand Pharmaceuticals, San Diego, CA 92121, USA
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27
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Tung L, Abdel-Hafiz H, Shen T, Harvell DME, Nitao LK, Richer JK, Sartorius CA, Takimoto GS, Horwitz KB. Progesterone receptors (PR)-B and -A regulate transcription by different mechanisms: AF-3 exerts regulatory control over coactivator binding to PR-B. Mol Endocrinol 2006; 20:2656-70. [PMID: 16762974 DOI: 10.1210/me.2006-0105] [Citation(s) in RCA: 74] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/19/2022] Open
Abstract
The two, nearly identical, isoforms of human progesterone receptors (PR), PR-B and -A, share activation functions (AF) 1 and 2, yet they possess markedly different transcriptional profiles, with PR-B being much stronger transactivators. Their differences map to a unique AF3 in the B-upstream segment (BUS), at the far N terminus of PR-B, which is missing in PR-A. Combined mutation of two LXXLL motifs plus tryptophan 140 in BUS, to yield PR-BdL140, completely destroys PR-B activity, because strong AF3 synergism with downstream AF1 and AF2 is eliminated. This synergism involves cooperative interactions among receptor multimers bound at tandem hormone response elements and is transferable to AFs of other nuclear receptors. Other PR-B functions-N-/C-terminal interactions, steroid receptor coactivator-1 coactivation, ligand-dependent down-regulation-also require an intact BUS. All three are autonomous in PR-A, and map to N-terminal regions common to both PR. This suggests that the N-terminal structure adopted by the two PR is different, and that for PR-B, this is controlled by BUS. Indeed, gene expression profiling of breast cancer cells stably expressing PR-B, PR-BdL140, or PR-A shows that mutation of AF3 destroys PR-B-dependent gene transcription without converting PR-B into PR-A. In sum, AF3 in BUS plays a critical modulatory role in PR-B, and in doing so, defines a mechanism for PR-B function that is fundamentally distinct from that of PR-A.
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Affiliation(s)
- Lin Tung
- Department of Medicine, RC1 South, 12801 East 17th Avenue, P.O. Box 6511, Aurora, Colorado 80045, USA
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28
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Geserick C, Meyer HA, Haendler B. The role of DNA response elements as allosteric modulators of steroid receptor function. Mol Cell Endocrinol 2005; 236:1-7. [PMID: 15876478 DOI: 10.1016/j.mce.2005.03.007] [Citation(s) in RCA: 72] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/19/2005] [Accepted: 03/23/2005] [Indexed: 11/25/2022]
Abstract
Steroid receptors are ligand-activated transcription factors which control the expression of their target genes by binding to specific DNA elements. Consensus response elements have been delineated for the glucocorticoid, androgen, progesterone and mineralocorticoid receptors on one hand (steroid response element, SRE) and for the estrogen receptor on the other hand (estrogen response element, ERE). Small variations in these sequences not only affect the binding but may also have a dramatic impact on the transcriptional activity of steroid receptors. It has now become obvious that DNA response elements do not merely tether regulatory proteins to control regions of target genes but may additionally impart conformational changes onto the DNA-binding domain as well as to neighbouring domains of steroid receptors. This in turn will create unique platforms for selective recruitment of cofactors and possibly for induction of modifications in local chromatin architecture. An additional level of complexity is added by the frequent presence of multiple response elements in gene promoter regions. The allosteric effects of DNA response elements on steroid receptors may be essential for differential gene expression and this offers interesting perspectives for the identification of selective modulators.
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Abstract
Steroidal glucocorticoids are commonly used due to their powerful antiinflammatory activity. However, despite their excellent efficacy, severe side effects frequently limit the use of these drugs. The search for novel glucocorticoids with reduced side effects has been intensified by the discovery of new molecular details regarding the function of the glucocorticoid receptor. These new insights may pave the way for novel, safer therapies that retain the efficacy of currently prescribed steroids.
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Affiliation(s)
- Jonathan Rosen
- Department of Molecular and Cell Biology, Ligand Pharmaceuticals, 10275 Science Center Drive, San Diego, California 92121, USA
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30
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Schoneveld OJLM, Gaemers IC, Lamers WH. Mechanisms of glucocorticoid signalling. ACTA ACUST UNITED AC 2004; 1680:114-28. [PMID: 15488991 DOI: 10.1016/j.bbaexp.2004.09.004] [Citation(s) in RCA: 215] [Impact Index Per Article: 10.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/02/2004] [Revised: 09/10/2004] [Accepted: 09/13/2004] [Indexed: 10/26/2022]
Abstract
It has become increasingly clear that glucocorticoid signalling not only comprises the binding of the glucocorticoid receptor (GR) to its response element (GRE), but also involves indirect regulation glucocorticoid-responsive genes by regulating or interacting with other transcription factors. In addition, they can directly regulate gene expression by binding to negative glucocorticoid response elements (nGREs), to simple GREs, to GREs, or to GREs and GRE half sites (GRE1/2s) that are part of a regulatory unit. A response unit allows a higher level of glucocorticoid induction than simple GREs and, in addition, allows the integration of tissue-specific information with the glucocorticoid response. Presumably, the complexity of such a glucocorticoid response unit (GRU) depends on the number of pathways that integrate at this unit. Because GRUs are often located at distant sites relative to the transcription-start site, the GRU has to find a way to communicate with the basal-transcription machinery. We propose that the activating signal of a distal enhancer can be relayed onto the transcription-initiation complex by coupling elements located proximal to the promoter.
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Affiliation(s)
- Onard J L M Schoneveld
- AMC Liver Center, Academic Medical Center, University of Amsterdam, Meibergdreef 69-71, 1105 BK, Amsterdam, The Netherlands
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31
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De Bosscher K, Vanden Berghe W, Haegeman G. The interplay between the glucocorticoid receptor and nuclear factor-kappaB or activator protein-1: molecular mechanisms for gene repression. Endocr Rev 2003; 24:488-522. [PMID: 12920152 DOI: 10.1210/er.2002-0006] [Citation(s) in RCA: 635] [Impact Index Per Article: 28.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/23/2022]
Abstract
The inflammatory response is a highly regulated physiological process that is critically important for homeostasis. A precise physiological control of inflammation allows a timely reaction to invading pathogens or to other insults without causing overreaction liable to damage the host. The cellular signaling pathways identified as important regulators of inflammation are the signal transduction cascades mediated by the nuclear factor-kappaB and the activator protein-1, which can both be modulated by glucocorticoids. Their use in the clinic includes treatment of rheumatoid arthritis, asthma, allograft rejection, and allergic skin diseases. Although glucocorticoids have been widely used since the late 1940s, the molecular mechanisms responsible for their antiinflammatory activity are still under investigation. The various molecular pathways proposed so far are discussed in more detail.
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Affiliation(s)
- Karolien De Bosscher
- Department of Molecular Biology, Ghent University, K. L. Ledeganckstraat 35, 9000 Gent, Belgium
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32
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Olswang Y, Blum B, Cassuto H, Cohen H, Biberman Y, Hanson RW, Reshef L. Glucocorticoids repress transcription of phosphoenolpyruvate carboxykinase (GTP) gene in adipocytes by inhibiting its C/EBP-mediated activation. J Biol Chem 2003; 278:12929-36. [PMID: 12560325 DOI: 10.1074/jbc.m300263200] [Citation(s) in RCA: 30] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/07/2023] Open
Abstract
The cytosolic form of the phosphoenolpyruvate carboxykinase (PEPCK-C) gene is selectively expressed in several tissues, primarily in the liver, kidney, and adipose tissue. The transcription of the gene is reciprocally regulated by glucocorticoids in these tissues. It is induced in the liver and kidney but repressed in the white adipose tissue. To elucidate which adipocyte-specific transcription factors participate in the repression of the gene, DNase I footprinting analyses of nuclear proteins from 3T3-F442A adipocytes and transient transfection experiments in NIH3T3 cells were utilized. Glucocorticoid treatment slightly reduced the nuclear C/EBP alpha concentration but prominently diminished the binding of adipocyte-derived nuclear proteins to CCAAT/enhancer-binding protein (C/EBP) recognition sites, without affecting the binding to nuclear receptor sites in the PEPCK-C gene promoter. Of members of the C/EBP family of transcription factors, C/EBP alpha was the strongest trans-activator of the PEPCK-C gene promoter in the NIH3T3 cell line. The glucocorticoid receptor (GR), in the presence of its hormone ligand, inhibited the activation of the PEPCK-C gene promoter by C/EBP alpha or C/EBP beta but not by the adipocyte-specific peroxisome proliferator-activated receptor gamma 2. This inhibition effect was similar using the wild type or mutant GR and did not depend on GR binding to the DNA. The glucocorticoid response unit (GRU) in the PEPCK-C gene promoter (-2000 to +73) restrained C/EBP alpha-mediated trans-activation, because mutation of each single GRU element increased this activation by 3-4-fold. This series of GRU mutations were repressed by wild type GR to the same percent as was the nonmutated PEPCK-C gene promoter. In contrast, the repression by mutant GR depended on the intact AF1 site in the gene promoter, whereby mutation of the AF1 element abolished the repression.
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Affiliation(s)
- Yael Olswang
- Department of Developmental Biochemistry, Hebrew University-Hadassah Medical School, Jerusalem, Israel 91120, USA
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33
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Abstract
Glucocorticoids (GCs) are the most common group of medications used in the treatment of allergic and autoimmune disorders. They produce potent anti-inflammatory effects by inducing or repressing the expression of target genes. Although most patients with allergic diseases and autoimmune disorders respond to GC therapy, a small subset of patients demonstrate persistent tissue inflammation despite treatment with high doses of GCs. This condition results from an interaction between susceptibility genes, the host's environment, and immunologic factors. The treatment of these patients requires a systematic approach to rule out underlying conditions that lead to steroid resistance or treatment failure, as well as the use of alternative strategies to inhibit tissue inflammation.
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Affiliation(s)
- Donald Y M Leung
- Division of Pediatric Allergy/Immunology, National Jewish Medical and Research Center, Denver, CO 80206, USA
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34
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Periyasamy S, Sánchez ER. Antagonism of glucocorticoid receptor transactivity and cell growth inhibition by transforming growth factor-beta through AP-1-mediated transcriptional repression. Int J Biochem Cell Biol 2002; 34:1571-85. [PMID: 12379279 DOI: 10.1016/s1357-2725(02)00057-2] [Citation(s) in RCA: 19] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/16/2022]
Abstract
We have examined the interaction of the glucocorticoid receptor (GR) and transforming growth factor-beta (TGF-beta) signal pathways because of their mutual involvement in the regulation of cell growth, development and differentiation. Most studies of this cross-talk event have focused on the effects of glucocorticoids (GCs) on TGF-beta responses. In this work, we show that TGF-beta can antagonize dexamethasone (Dex)-mediated growth suppression in mouse fibrosarcoma L929 cells. TGF-beta also repressed GR-mediated reporter (pMMTV-CAT) gene expression in a concentration-dependent manner, with an IC(50) of 5 ng/ml of TGF-beta. Maximal inhibition (76%) was observed at 10 ng/ml of TGF-beta. Conversely, Dex inhibited TGF-beta-mediated promoter (p3TP-Lux) activity in these same cells. As TGF-beta inhibition of GR-mediated gene expression occurred after Dex-mediated nuclear translocation of GR, we conclude that TGF-beta inhibition of GR signaling occurs at the level of GR-mediated transcription activity. However, TGF-beta did not repress GR-mediated gene expression using the pGRE(2)E1B-CAT minimal promoter construct, suggesting that TGF-beta did not inhibit intrinsic GR activity but, rather, required DNA-binding factor(s) distinct from GR. As the MMTV promoter contains several putative AP-1 binding sites, we hypothesized that AP-1, a transcription factor composed of c-jun and c-fos proteins, might be involved in the TGF-beta inhibition of GR functions. Curcumin, a potent inhibitor of AP-1 expression, completely abolished the inhibitory effect of TGF-beta on GR-mediated gene expression without affecting GR activity in the absence of TGF-beta, and this drug blocked TGF-beta-induced binding of AP-1 to a response element derived from the MMTV sequence. Furthermore, curcumin abolished TGF-beta inhibition of Dex-induced growth suppression. Taken as a whole, our data suggest that TGF-beta can antagonize the growth inhibitory properties of GR by blocking GR transactivity at complex promoters through a mechanism involving transcriptional repression by DNA-bound AP-1.
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Affiliation(s)
- Sumudra Periyasamy
- Department of Pharmacology, Medical College of Ohio, 3035 Arlington Avenue, Toledo, OH 43614, USA.
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35
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Ayala JE, Streeper RS, Svitek CA, Goldman JK, Oeser JK, O'Brien RM. Accessory elements, flanking DNA sequence, and promoter context play key roles in determining the efficacy of insulin and phorbol ester signaling through the malic enzyme and collagenase-1 AP-1 motifs. J Biol Chem 2002; 277:27935-44. [PMID: 12032154 DOI: 10.1074/jbc.m203682200] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/19/2023] Open
Abstract
Insulin stimulates malic enzyme (ME)-chloramphenicol acetyltransferase (CAT) and collagenase-1-CAT fusion gene expression in H4IIE cells through identical activator protein-1 (AP-1) motifs. In contrast, insulin and phorbol esters only stimulate collagenase-1-CAT and not ME-CAT fusion gene expression in HeLa cells. The experiments in this article were designed to explore the molecular basis for this differential cell type- and gene-specific regulation. The results highlight the influence of three variables, namely promoter context, AP-1 flanking sequence, and accessory elements that modulate insulin and phorbol ester signaling through the AP-1 motif. Thus, fusion gene transfection and proteolytic clipping gel retardation assays suggest that the AP-1 flanking sequence affects the conformation of AP-1 binding to the collagenase-1 and ME AP-1 motifs such that it selectively binds the latter in a fully activated state. However, this influence of ME AP-1 flanking sequence is dependent on promoter context. Thus, the ME AP-1 motif will mediate both an insulin and phorbol ester response in HeLa cells when introduced into either the collagenase-1 promoter or a specific heterologous promoter. But even in the context of the collagenase-1 promoter, the effects of both insulin and phorbol esters, mediated through the ME AP-1 motif are dependent on accessory factors.
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Affiliation(s)
- Julio E Ayala
- Department of Molecular Physiology and Biophysics, Vanderbilt University Medical School, Nashville, Tennessee 37232, USA
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36
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Nakopoulou L, Giannopoulou I, Stefanaki K, Panayotopoulou E, Tsirmpa I, Alexandrou P, Mavrommatis J, Katsarou S, Davaris P. Enhanced mRNA expression of tissue inhibitor of metalloproteinase-1 (TIMP-1) in breast carcinomas is correlated with adverse prognosis. J Pathol 2002; 197:307-13. [PMID: 12115876 DOI: 10.1002/path.1129] [Citation(s) in RCA: 68] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/10/2022]
Abstract
Tissue inhibitor of metalloproteinase-1 (TIMP-1) has emerged as a multifunctional protein with the contrasting activities of inhibiting tissue-degrading enzymes and promoting cellular growth. In an attempt to elucidate the clinical significance of TIMP-1 in breast cancer, the expression of TIMP-1 mRNA was evaluated in 117 invasive breast carcinomas by mRNA in situ hybridization, in correlation with clinicopathological parameters, immunohistochemical prognostic factors (Ki-67, c-erb-B-2, bcl-2) and clinical outcome. TIMP-1 was detected in stromal cells in areas within the tumours and at the tumour margin. High TIMP-1 mRNA expression in the marginal portion of the tumours was significantly correlated with lymph node metastasis (p<0.05) and c-erbB-2 expression (p<0.05). On the other hand, increased TIMP-1 mRNA expression within the tumours showed a statistically significant correlation with ER detection (p<0.01). Multivariate analysis revealed worse survival for patients with high TIMP-1 mRNA expression in the marginal portion of the tumours; the subgroup of these patients co-expressing high levels of TIMP-1 mRNA within the tumours as well had even worse survival (p=0.042). In conclusion, our data support the multifunctional role of TIMP-1, particularly its growth-promoting activity, on the basis of its significant correlation with lymph node metastasis and adverse prognosis. In addition to the latter property, a probable association of TIMP-1 with tumour cell differentiation is suggested by its topographical correlation with ER detection.
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MESH Headings
- Adult
- Aged
- Aged, 80 and over
- Biomarkers/analysis
- Breast Neoplasms/metabolism
- Breast Neoplasms/mortality
- Carcinoma, Ductal, Breast/metabolism
- Carcinoma, Ductal, Breast/mortality
- Carcinoma, Lobular/metabolism
- Carcinoma, Lobular/mortality
- Chi-Square Distribution
- Female
- Humans
- Immunohistochemistry/methods
- In Situ Hybridization/methods
- Ki-67 Antigen/analysis
- Lymphatic Metastasis
- Middle Aged
- Proto-Oncogene Proteins c-bcl-2/analysis
- RNA, Messenger/metabolism
- Receptor, ErbB-2/analysis
- Ribonuclease, Pancreatic
- Survival Rate
- Tissue Inhibitor of Metalloproteinase-1/genetics
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Affiliation(s)
- Lydia Nakopoulou
- Department of Pathology, Athens National University Medical School, Greece.
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37
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Gebhardt C, Breitenbach U, Tuckermann JP, Dittrich BT, Richter KH, Angel P. Calgranulins S100A8 and S100A9 are negatively regulated by glucocorticoids in a c-Fos-dependent manner and overexpressed throughout skin carcinogenesis. Oncogene 2002; 21:4266-76. [PMID: 12082614 DOI: 10.1038/sj.onc.1205521] [Citation(s) in RCA: 102] [Impact Index Per Article: 4.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/10/2001] [Revised: 03/06/2002] [Accepted: 03/26/2002] [Indexed: 01/01/2023]
Abstract
The two calgranulins S100A8 and S100A9 were found to be differentially expressed at sites of acute and chronic inflammation. Here we have employed the phorbol ester-induced multistage skin carcinogenesis protocol in mice to determine the expression of both genes in inflamed skin and in skin tumors. We show that expression is coordinately induced by the phorbol ester TPA in epithelial cells as well as infiltrating leukocytes. By comparing S100A8 and S100A9 mRNA levels in wild type and c-Fos deficient mice (c-fos(-/-)) we found that expression is negatively regulated by c-Fos/AP-1. Glucocorticoids, which exhibit potent anti-inflammatory and anti-tumor promoting activities repressed TPA-mediated S100A8 and S100A9 induction in wild type, but not in c-fos(-/-) mice, thus identifying both genes as the first examples of AP-1 target genes whose repression of TPA-induced transcription by glucocorticoids depends on c-Fos. Finally, we show that enhanced expression is not restricted to the initial TPA-induced inflammatory response but is observed at all stages of skin carcinogenesis. These data identify S100A8 and S100A9 as novel, tumor-associated genes and may point to an as yet unrecognized function of both genes in the development of epithelial skin tumors.
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MESH Headings
- Animals
- Anti-Inflammatory Agents/pharmacology
- Antigens, Differentiation/biosynthesis
- Antigens, Differentiation/genetics
- Antineoplastic Agents, Hormonal/pharmacology
- Calcium/physiology
- Calcium-Binding Proteins/biosynthesis
- Calcium-Binding Proteins/genetics
- Calgranulin A
- Calgranulin B
- Carcinogens/pharmacology
- Carcinogens/toxicity
- Carcinoma, Squamous Cell/chemically induced
- Carcinoma, Squamous Cell/genetics
- Carcinoma, Squamous Cell/metabolism
- Dexamethasone/pharmacology
- Disease Progression
- Drug Eruptions/etiology
- Drug Eruptions/genetics
- Drug Eruptions/metabolism
- Female
- Gene Expression Regulation/drug effects
- Gene Expression Regulation, Neoplastic/drug effects
- Genes, fos
- Keratinocytes/drug effects
- Keratinocytes/metabolism
- Leukocytes/drug effects
- Leukocytes/metabolism
- Mice
- Mice, Inbred C57BL
- Mice, Inbred Strains
- Mice, Knockout
- Papilloma/chemically induced
- Papilloma/genetics
- Papilloma/metabolism
- Protein Kinase C/antagonists & inhibitors
- Proto-Oncogene Proteins c-fos/deficiency
- Proto-Oncogene Proteins c-fos/physiology
- S100 Proteins/biosynthesis
- S100 Proteins/genetics
- Skin Neoplasms/chemically induced
- Skin Neoplasms/genetics
- Skin Neoplasms/metabolism
- Specific Pathogen-Free Organisms
- Tetradecanoylphorbol Acetate/pharmacology
- Tetradecanoylphorbol Acetate/toxicity
- Transcription Factor AP-1/physiology
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Affiliation(s)
- Christoffer Gebhardt
- Deutsches Krebsforschungszentrum, Division of Signal Transduction and Growth Control, 69120 Heidelberg, Germany
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38
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Shah OJ, Iniguez-Lluhi JA, Romanelli A, Kimball SR, Jefferson LS. The activated glucocorticoid receptor modulates presumptive autoregulation of ribosomal protein S6 protein kinase, p70 S6K. J Biol Chem 2002; 277:2525-33. [PMID: 11705993 DOI: 10.1074/jbc.m105935200] [Citation(s) in RCA: 21] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/02/2023] Open
Abstract
Protein metabolism in eukaryotic organisms is defined by a synthesis-degradation equilibrium that is subject to regulation by hormonal and nutritional signals. In mammalian tissues such as skeletal muscle, glucocorticoid hormones specify a catabolic response that influences both protein synthetic and protein degradative pathways. With regard to the former, glucocorticoids attenuate mRNA translation at two levels: translational efficiency, i.e. translation initiation, and translational capacity, i.e. ribosome biogenesis. Glucocorticoids may impair translational capacity through the ribosomal S6 protein kinase (p70 S6K), a recognized glucocorticoid target and an effector of ribosomal protein synthesis. We demonstrate here that the reduction in growth factor-activated p70 S6K activity by glucocorticoids depends upon a functional glucocorticoid receptor (GR) and that the GR is both necessary and sufficient to render p70 S6K subject to glucocorticoid regulation. Furthermore, the DNA binding and transcriptional activation but not repression properties of the GR are indispensable for p70 S6K regulation. Finally, a mutational analysis of the p70 S6K carboxyl terminus indicates that this region confers glucocorticoid sensitivity, and thus glucocorticoids may facilitate autoinhibition of the enzyme ultimately reducing the efficiency with which T389 is phosphorylated.
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Affiliation(s)
- O Jameel Shah
- Department of Cellular and Molecular Physiology, Pennsylvania State University College of Medicine, Hershey, Pennsylvania 17033-0850, USA
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39
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Distelhorst CW. Recent insights into the mechanism of glucocorticosteroid-induced apoptosis. Cell Death Differ 2002; 9:6-19. [PMID: 11803370 DOI: 10.1038/sj.cdd.4400969] [Citation(s) in RCA: 222] [Impact Index Per Article: 9.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/20/2001] [Revised: 09/07/2001] [Accepted: 10/03/2001] [Indexed: 01/08/2023] Open
Abstract
Glucocorticosteroid hormones induce apoptosis in lymphocytes. Therefore, glucocorticoids are commonly used as immunosuppressive and chemotherapeutic agents. This review examines many facets of the process by which glucocorticoids induce apoptosis. This process is divided into three stages, an initiation stage that involves glucocorticoid receptor-mediated gene regulation, a decision stage that involves the counterbalancing influence of prosurvival and proapoptotic factors, and the execution stage which involves caspase and endonuclease activation. Many aspects of glucocorticoid-induced apoptosis, such as mitochondrial dysfunction and caspase activation, are important steps in virtually all forms of apoptosis. But the process glucocorticoid-induced apoptosis differs from other forms of apoptosis in terms of initiation at the transcriptional level and involvement of the multicatalytic proteasome and calcium. Moreover, the abundant opportunity for crosstalk between the glucocorticoid receptor and other signaling pathways increases the complexity of glucocorticoid-induced apoptosis and its regulation.
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Affiliation(s)
- C W Distelhorst
- Division of Hematology/Oncology and Comprehensive Cancer Center, Departments of Medicine and Pharmacology, Case Western Reserve University School of Medicine and University Hospitals of Cleveland, Cleveland, OH 44106-4937, USA.
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40
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Rahmani M, Péron P, Weitzman J, Bakiri L, Lardeux B, Bernuau D. Functional cooperation between JunD and NF-kappaB in rat hepatocytes. Oncogene 2001; 20:5132-42. [PMID: 11526502 DOI: 10.1038/sj.onc.1204678] [Citation(s) in RCA: 33] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/22/2001] [Revised: 05/29/2001] [Accepted: 05/30/2001] [Indexed: 11/10/2022]
Abstract
AP-1 and NF-kappaB are rapidly activated during liver regeneration. Whether these parallel inductions have potential functional implications is not known. Isolated rat hepatocytes were stimulated with two mitogens, epidermal growth factor or hepatocyte growth factor and with tumor necrosis factor alpha, a cytokine involved in the liver regenerative response in vivo and a strong inducer of NF-kappaB. All three cytokines increased AP-1 and NF-kappaB binding to their cognate cis-element and induced a 2.5-fold activation of NF-kappaB-dependent transcription. Inactivation of AP-1 by TAM67, a dominant negative mutant of AP-1 drastically inhibited basal and cytokine-induced NF-kappaB transactivation. Overexpression of Jun D, but not of the other Jun or Fos proteins increased by threefold NF-kappaB transactivation. Functional cooperation between JunD and p65 was demonstrated in a simple Gal-hybrid system. Finally, a twofold decrease in NF-kappaB transactivation was found in hepatocytes isolated from JunD(-/-) mice compared with hepatocytes from JunD(+/+) mice. Altogether these data demonstrate a functional cooperation of p65 with JunD, a major constituent of AP-1 in normal hepatocytes.
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Affiliation(s)
- M Rahmani
- Laboratoire de Biologie cellulaire, INSERM U 327, Faculté de Médecine Xavier Bichat et Université Paris 7 Denis Diderot, Paris, France
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41
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Shi MJ, Park SR, Kim PH, Stavnezer J. Roles of Ets proteins, NF-kappa B and nocodazole in regulating induction of transcription of mouse germline Ig alpha RNA by transforming growth factor-beta 1. Int Immunol 2001; 13:733-46. [PMID: 11369700 DOI: 10.1093/intimm/13.6.733] [Citation(s) in RCA: 29] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/18/2023] Open
Abstract
Antibody class switch recombination (CSR) occurs after antigen activation of B cells. CSR is directed to specific heavy chain isotypes by cytokines and B cell activators that induce transcription from the unrearranged, or germline (GL), C(H) region genes. Transforming growth factor (TGF)-beta1 is essential for switch recombination to IgA due to its ability to induce transcription from GL Ig alpha genes. It has been shown that the promoters which regulate transcription of mouse and human GL alpha RNAs contain a TGF-beta1-responsive element that binds Smad and core binding factor (CBFalpha)/AML/PEBPalpha/RUNX: They also contain other elements which bind the transcription factors CREB, BSAP and Ets family proteins. In this manuscript we demonstrate that two tandem Ets sites in the mouse GL alpha promoter bind the transcription factors Elf-1 and PU.1, and that the 3' site is essential for expression of a luciferase reporter gene driven by the GL alpha promoter. Binding of Elf-1 to the GL alpha promoter is inducible by lipopolysaccharide in nuclear extracts from splenic B cells. An NF-kappaB site is identified, although it does not contribute to expression of the promoter in reporter gene assays. Since CSR to IgA is greatly reduced in NF-kappaB/p50-deficient mice, these data support the hypothesis that NF-kappaB has roles in switching in addition to regulation of GL transcription. Finally, we demonstrate that nocodazole, which disrupts microtubules that sequester Smad proteins in the cytoplasm, stimulates transcription from the GL alpha promoter.
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Affiliation(s)
- M J Shi
- Department of Molecular Genetics and Microbiology, Program in Immunology and Virology, University of Massachusetts Medical School, Worcester, MA 01655-0122, USA
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42
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Mao CS, Stavnezer J. Differential regulation of mouse germline Ig gamma 1 and epsilon promoters by IL-4 and CD40. JOURNAL OF IMMUNOLOGY (BALTIMORE, MD. : 1950) 2001; 167:1522-34. [PMID: 11466373 DOI: 10.4049/jimmunol.167.3.1522] [Citation(s) in RCA: 53] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/19/2022]
Abstract
Before Ig class switching, RNA transcription through the specific S regions undergoing recombination is induced by cytokines and other activators that induce and direct switching. The resulting germline (GL) transcripts are essential for switch recombination. To understand the differential regulation of mouse IgG1 and IgE, we compared the promoters for GL gamma1 and epsilon transcripts. We addressed the question of why the promoter that regulates GL epsilon transcription is more responsive to IL-4 than the gamma1 promoter and also why GL epsilon transcription is more dependent on IL-4 than is gamma1 transcription. We found that the IL-4-responsive region of the GL epsilon promoter is more inducible than that of the gamma1 promoter, although each promoter contains a binding site for the IL-4-inducible transcription factor Stat6, located immediately adjacent to a binding site for a basic region leucine zipper (bZip) family protein. However, the arrangement and sequences of the sites differ between the epsilon and gamma1 promoters. The GL epsilon promoter binds Stat6 with a 10-fold higher affinity than does the gamma1 promoter. Furthermore, the bZip elements of the two promoters bind different transcription factors, as the GL epsilon promoter binds and is activated by AP-1, whereas the gamma1 promoter binds and is activated by activating transcription factor 2. C/EBPbeta and C/EBPgamma also bind the gamma1 bZip element, although they inhibit rather than activate transcription. However, inhibition of promoter activity by C/EBPbeta does not require the bZip element and may instead occur via inhibiting the activity of NF-kappaB.
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Affiliation(s)
- C S Mao
- Department of Molecular Genetics, Program in Immunology and Virology, University of Massachusetts Medical School, 55 Lake Avenue North, Worcester MA 01655, USA
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43
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Ramirez-Carrozzi VR, Kerppola TK. Control of the orientation of Fos-Jun binding and the transcriptional cooperativity of Fos-Jun-NFAT1 complexes. J Biol Chem 2001; 276:21797-808. [PMID: 11259418 DOI: 10.1074/jbc.m101494200] [Citation(s) in RCA: 16] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
Heterodimeric transcription regulatory proteins can bind to palindromic recognition elements in two opposite orientations. We have developed a gel-based fluorescence resonance energy transfer assay for quantifying heterodimer orientation preferences. Fos-Jun heterodimers bind in opposite orientations to AP-1 sites with different flanking sequences. The effects of individual amino acid and base pair substitutions on heterodimer binding orientation were quantified. Base pairs at positions +/-6 and +/-10 relative to the center of the AP-1 site were the principal determinants of Fos-Jun binding orientation. Amino acid residues of opposite charge adjacent to the basic regions of Fos and Jun had independent effects on heterodimer orientation. Exchange of these amino acid residues between the basic region-leucine zipper domains of Fos and Jun reversed the binding orientation. Heterodimers formed by full-length Fos and Jun exhibited the same changes in binding orientation in response to amino acid and base pair substitutions. The preferred orientation of heterodimer binding affected the stability of Fos-Jun-NFAT1 complexes at composite regulatory elements. Changes in heterodimer orientation preference altered the transcriptional activity and the promoter selectivity of Fos-Jun-NFAT1 complexes. Consequently, the orientation of Fos-Jun binding can influence transcriptional activity by altering cooperative interactions with other transcription regulatory proteins.
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Affiliation(s)
- V R Ramirez-Carrozzi
- Howard Hughes Medical InstituteM Department of Biological Chemistry, University of Michigan Medical School, Ann Arbor, Michigan 48109-0650, USA
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44
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Lazennec G, Thomas JA, Katzenellenbogen BS. Involvement of cyclic AMP response element binding protein (CREB) and estrogen receptor phosphorylation in the synergistic activation of the estrogen receptor by estradiol and protein kinase activators. J Steroid Biochem Mol Biol 2001; 77:193-203. [PMID: 11457657 DOI: 10.1016/s0960-0760(01)00060-7] [Citation(s) in RCA: 55] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/19/2022]
Abstract
Estrogen receptor (ER) and cAMP signaling pathways interact in a number of estrogen target tissues including mammary and uterine tissues. One aspect of this interaction is that estradiol and protein kinase A (PKA) activators can cooperate synergistically to activate ER-mediated transcription of both endogenous genes and reporter genes containing only estrogen response elements. The purpose of this study was to investigate the molecular mechanism of this interaction between signaling pathways. Site-directed mutagenesis of the potential PKA phosphorylation sites in the ER indicated that phosphorylation of these sites was not necessary for the observed transcriptional synergy. In transient transfection assays in two different cell lines using reporter constructs containing either cAMP response elements, estrogen response elements or both types of elements, with the addition or absence of cAMP response element binding protein (CREB) expression plasmid, we observed that only one of these cell lines exhibited estrogen/PKA transcriptional synergy. Experiments demonstrated that CREB itself was involved in the transcriptional synergy, and that transfection of CREB restored transcriptional synergy in the cell line in which it was lacking. A functional interaction between ER and CREB was also demonstrated using a mammalian cell protein interaction assay; a dominant negative mutant of CREB did not exhibit this interaction. Therefore, these data indicate that CREB protein is required for the transcriptional synergy between cAMP and estrogen signaling pathways. Furthermore, CREB cooperated with the ER on genes that did not contain cAMP response elements, but contained only estrogen response elements. We propose that activated CREB is recruited to estrogen responsive genes by an ER--coactivator complex containing proteins such as CREB binding protein (CBP) and that the interaction of CREB with ER may assist in stabilizing its interaction with CBP and in promoting estrogen-ER and PKA transcriptional synergy.
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Affiliation(s)
- G Lazennec
- Department of Molecular and Integrative Physiology, University of Illinois at Urbana-Champaign, 524 Burrill Hall, 407 South Goodwin Ave, Urbana, IL 61801, USA
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45
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Wallberg AE, Wright A, Gustafsson JA. Chromatin-remodeling complexes involved in gene activation by the glucocorticoid receptor. VITAMINS AND HORMONES 2001; 60:75-122. [PMID: 11037622 DOI: 10.1016/s0083-6729(00)60017-1] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 02/18/2023]
Affiliation(s)
- A E Wallberg
- Karolinska Institute, Department of Biosciences, NOVUM, Huddinge, Sweden
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46
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Rinehart-Kim J, Johnston M, Birrer M, Bos T. Alterations in the gene expression profile of MCF-7 breast tumor cells in response to c-Jun. Int J Cancer 2000; 88:180-90. [PMID: 11004666 DOI: 10.1002/1097-0215(20001015)88:2<180::aid-ijc6>3.0.co;2-h] [Citation(s) in RCA: 33] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/07/2022]
Abstract
MCF7 breast tumor cells overexpressing human c-Jun exhibit a transformed phenotype characterized not only by increased tumorigenicity but also by enhanced motility and invasion. The cellular phenotypic response to c-Jun overexpression is likely due, at least in part, to altered patterns of gene expression. In order to begin to understand the complexities by which elevated production of c-Jun alters the state of the cell, we have profiled the expression of 588 different genes by comparative hybridization. By using this approach, we have identified a total of 21 upregulated or downregulated gene targets responsive to c-Jun overexpression. Interestingly, 8 of these genes have been previously found associated with c-Jun or AP-1 activity and therefore provide internal validation for this approach to target gene discovery. The remaining 13 genes represent potential new c-Jun regulated target genes. Genomic sequence information was available for 15 of the 21 genes identified in this screen. Analysis of these genomic sequences revealed the presence of AP-1 or AP-1-like sequences in 12 of the 15 genes examined. Consistent with a direct mechanism of target regulation by c-Jun, gel shift analysis of selected AP-1-containing promoter regions revealed elevated and specific binding by proteins present in nuclear extracts of c-Jun expressing MCF7 cells.
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Affiliation(s)
- J Rinehart-Kim
- Department of Microbiology and Molecular Cell Biology, Eastern Virginia Medical School, Norfolk, Virginia, USA
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Basso J, Briggs J, Findlay C, Bos T. Directed mutation of the basic domain of v-Jun alters DNA binding specificity and abolishes its oncogenic activity in chicken embryo fibroblasts. Oncogene 2000; 19:4876-85. [PMID: 11039905 DOI: 10.1038/sj.onc.1203863] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/08/2022]
Abstract
Overexpression of v-Jun in chicken embryo fibroblasts (CEF) leads to oncogenic transformation phenotypically characterized by anchorage independent growth and release from contact inhibition (focus formation). The mechanisms involved in this oncogenic conversion however, are not yet clear. Because Jun is a transcription factor, it has been assumed that oncogenic transformation results directly from deregulated AP-1 target gene expression. However, a number of experimental observations in avian cell culture models fail to correlate oncogenesis with AP-1 activity suggesting that transformation induced by v-Jun may occur through an indirect mechanism. To test this possibility, we introduced point mutations into the basic DNA binding domain of v-Jun and created mutants that exhibit altered binding specificity. When expressed in CEF, these mutants fail to deregulate three known v-Jun target genes (JTAP-1, apolipoprotein A1, c-Jun) thus demonstrating in vivo specificity changes. Each of the binding specificity mutants was also tested for its ability to induce oncogenic transformation. Interestingly, expression of these mutants in CEF results in a phenotype indistinguishable from the vector control with respect to growth rate, focus formation and the ability to form colonies in soft agar. These results are consistent with a model requiring direct AP-1 target deregulation as a prerequisite of v-Jun induced cell transformation. With this in mind, we generated a series of additional mutants that retain the ability to bind AP-1 sequence elements, but vary in their oncogenic potential. We demonstrate the use of these mutants to screen v-Jun induced gene targets for a functional role in cell transformation.
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Affiliation(s)
- J Basso
- Department of Microbiology and Molecular Cell Biology, Eastern Virginia Medical School, Norfolk 23501, USA
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48
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Radoja N, Komine M, Jho SH, Blumenberg M, Tomic-Canic M. Novel mechanism of steroid action in skin through glucocorticoid receptor monomers. Mol Cell Biol 2000; 20:4328-39. [PMID: 10825196 PMCID: PMC85800 DOI: 10.1128/mcb.20.12.4328-4339.2000] [Citation(s) in RCA: 83] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/16/1999] [Accepted: 03/20/2000] [Indexed: 11/20/2022] Open
Abstract
Glucocorticoids (GCs), important regulators of epidermal growth, differentiation, and homeostasis, are used extensively in the treatment of skin diseases. Using keratin gene expression as a paradigm of epidermal physiology and pathology, we have developed a model system to study the molecular mechanism of GCs action in skin. Here we describe a novel mechanism of suppression of transcription by the glucocorticoid receptor (GR) that represents an example of customizing a device for transcriptional regulation to target a specific group of genes within the target tissue, in our case, epidermis. We have shown that GCs repress the expression of the basal-cell-specific keratins K5 and K14 and disease-associated keratins K6, K16, and K17 but not the differentiation-specific keratins K3 and K10 or the simple epithelium-specific keratins K8, K18, and K19. We have identified the negative recognition elements (nGREs) in all five regulated keratin gene promoters. Detailed footprinting revealed that the function of nGREs is to instruct the GR to bind as four monomers. Furthermore, using cotransfection and antisense technology we have found that, unlike SRC-1 and GRIP-1, which are not involved in the GR complex that suppresses keratin genes, histone acetyltransferase and CBP are. In addition, we have found that GR, independently from GREs, blocks the induction of keratin gene expression by AP1. We conclude that GR suppresses keratin gene expression through two independent mechanisms: directly, through interactions of keratin nGREs with four GR monomers, as well as indirectly, by blocking the AP1 induction of keratin gene expression.
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Affiliation(s)
- N Radoja
- The Ronald O. Perelman Department of Dermatology, New York University School of Medicine, New York, New York 10016, USA
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Cheng YH, Nicholson RC, King B, Chan EC, Fitter JT, Smith R. Glucocorticoid stimulation of corticotropin-releasing hormone gene expression requires a cyclic adenosine 3',5'-monophosphate regulatory element in human primary placental cytotrophoblast cells. J Clin Endocrinol Metab 2000; 85:1937-45. [PMID: 10843178 DOI: 10.1210/jcem.85.5.6552] [Citation(s) in RCA: 14] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/19/2022]
Abstract
Production of placental CRH, which is identical to the peptide synthesized and secreted in the hypothalamus, has been linked to human parturition. Glucocorticoids stimulate placental CRH secretion and messenger ribonucleic acid expression, in contrast to their inhibition of CRH synthesis in the hypothalamus. A positive feedforward loop involving glucocorticoid-CRH-ACTH-glucocorticoid is thought to drive the exponential increase in placental CRH leading to delivery. Tissue-specific effects of glucocorticoids on CRH expression are therefore of interest. Using human primary placental cells, we investigated the mechanism by which glucocorticoids stimulate placental CRH gene expression. Nuclear run-on transcription shows that in human placental cells glucocorticoids up-regulate transcription of human CRH (hCRH). Using transient transfection assays we demonstrate that dexamethasone up-regulates both basal and cAMP-stimulated hCRH promoter activity, correlating well with the increase in endogenous CRH peptide levels. Through mutagenesis and deletion analyses we show that dexamethasone stimulation of hCRH gene transcription requires a functional cAMP regulatory element (CRE); this CRE is adequate to confer dexamethasone stimulation upon a heterologous promoter, and electrophoretic mobility shift assay studies show that a placental nuclear protein specifically binds to the hCRH CRE.
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Affiliation(s)
- Y H Cheng
- Mothers and Babies Research Center, Endocrine Unit, John Hunter Hospital, Newcastle, New South Wales, Australia
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50
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Shanmugam A, Shi MJ, Yauch L, Stavnezer J, Kenter AL. Evidence for class-specific factors in immunoglobulin isotype switching. J Exp Med 2000; 191:1365-80. [PMID: 10770803 PMCID: PMC2193137 DOI: 10.1084/jem.191.8.1365] [Citation(s) in RCA: 40] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/19/1999] [Accepted: 01/21/2000] [Indexed: 11/04/2022] Open
Abstract
Immunoglobulin class switch recombination (SR) occurs by a B cell-specific, intrachromosomal deletional process between switch regions. We have developed a plasmid-based transient transfection assay for SR to test for the presence of transacting switch activities. The plasmids are novel in that they lack a eukaryotic origin of DNA replication. The recombination activity of these switch substrates is restricted to a subset of B cell lines that support isotype switching on their endogenous loci and to mitogen-activated normal splenic B cells. The factors required for extrachromosomal plasmid recombination are constitutively expressed in proliferating splenic B cells and in B cell lines capable of inducibly undergoing immunoglobulin SR on their chromosomal genes. These studies suggest that mitogens that induce switching on the chromosome induce accessibility rather than switch recombinase activity. Finally, we provide evidence for two distinct switching activities which independently mediate mu-->alpha and mu-->gamma3 SR.
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Affiliation(s)
- Ananth Shanmugam
- Department of Microbiology and Immunology, University of Illinois College of Medicine, Chicago, Illinois 60612
| | - Meng-Jiao Shi
- Department of Molecular Genetics and Microbiology and the Program in Immunology and Virology, University of Massachusetts Medical School, Worcester, Massachusetts 01655
| | - Lauren Yauch
- Department of Microbiology and Immunology, University of Illinois College of Medicine, Chicago, Illinois 60612
| | - Janet Stavnezer
- Department of Molecular Genetics and Microbiology and the Program in Immunology and Virology, University of Massachusetts Medical School, Worcester, Massachusetts 01655
| | - Amy L. Kenter
- Department of Microbiology and Immunology, University of Illinois College of Medicine, Chicago, Illinois 60612
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