Regulation of cyclooxygenase-2 expression by cAMP response element and mRNA stability in a human airway epithelial cell line exposed to zinc
Introduction
Cyclooxygenase (COX) is a heme-containing enzyme that catalyzes two sequential enzymatic reactions: the bis-oxygenation of arachidonic acid leading to the production of prostaglandin G2 (PGG2) and the reduction of 15-hydroperoxide of PGG2, leading to the formation of PGH2, a precursor of all PGs, thromboxanes, and prostacyclins, in concert with a series of cell-specific isomerases (Smith et al., 2000). Three COX isoforms, COX-1, COX-2, and COX-3, have been identified in mammals (Chandrasekharan et al., 2002). COX-1 is expressed constitutively in most tissues and appears to be responsible for the production of PGs that modulate physiological functions. COX-3 is an alternatively spliced form of COX-1, expressed primarily in brain and heart as a constitutive enzyme. In contrast, COX-2 is expressed at low or undetectable levels in most tissues and cells under basal conditions, but is rapidly inducible by a variety of stimuli such as lipopolysaccharide (LPS), inflammatory cytokines, growth factors, ultraviolet radiation, and chemicals (Fu et al., 1990, Zhang et al., 1998, Subbaramaiah et al., 2000, Chang et al., 2003, Huh et al., 2003).
The COX-2 gene is mapped to human chromosome 1q25.2-q25.3, approximately 8.3 kb in length with 10 exons, and is transcribed as a 4.4 kb mRNA (Tanabe and Tohnai, 2002). The human COX-2 5′-flanking region contains a canonical TATA box and several functionally important enhancer elements including a cyclic AMP response element (CRE), E box and activator protein 1 (AP-1) regulatory element complex situated very close to TATA, a CCAAT/enhancer binding protein (C/EBP) site and two κB sites (Tazawa et al., 1994). The proinflammatory stimuli can induce binding of different transcription factors to their specific DNA-binding sites in a cell type- and stimulus-specific fashion. The transcription factors that bind and activate COX-2 transcription involve C/EBPβ and C/EBPδ for the nuclear factor interleukin-6 (NF-IL-6) elements, AP-1, activating transcription factor (ATF) and CRE-binding protein (CREB) for the CRE element, and upstream stimulatory factor 1 (USF-1) for the E box (Murakami and Kudo, 2004). Posttranscriptional events also play an important role in modulating COX-2 mRNA levels (Dannenberg et al., 2005). The first 60 nucleotides of the 3′-untranslated region (UTR) of COX-2 mRNA are highly conserved and contain multiple copies of the regulatory sequence AUUUA. These well-known AU-rich elements (AREs), present within the 3′-UTRs of many proto-oncogene and cytokine mRNAs, confer posttranscriptional control of expression by acting as a mRNA instability determinant or as a translation inhibitory element that can affect both mRNA and protein translation (Caput et al., 1986, Xu et al., 1997). An ARE element within the 3′-UTR of COX-2 mRNA has been identified that can control both mRNA decay and protein translation (Dixon et al., 2000, Dixon et al., 2001).
Increased COX-2 protein expression has been implicated in the pathogenesis of lung diseases characterized by chronic airway inflammation, including asthma, chronic bronchitis, cystic fibrosis, and bronchiectasis (Ermert et al., 1998, Oguma et al., 2002). Expression of the COX-2 gene has been shown in human airway epithelial cells exposed to exogenous stimuli, such as air-borne residual oil fly ash (Samet et al., 2000), hydrochloric acid (Bonnans et al., 2006), peroxisome proliferator-activated receptor-gamma agonists (Patel et al., 2005), respiratory syncytial virus and Streptococcus pneumoniae infection (Liu et al., 2005, N'Guessan et al., 2006). Zinc (Zn) is an essential micronutrient involved in structural and regulatory cellular functions of a large number of proteins (Vallee and Falchuk, 1993). Zn is also a ubiquitous contaminant in ambient and occupational settings. It exists as a combustion-derived metal associated with ambient particulate matter (PM) and may contribute to the adverse health effects of ambient PM inhalation (Horner, 1996, Adamson et al., 2000). In this study, the regulation of COX-2 expression was studied in a human bronchial epithelial cell line BEAS-2B exposed to Zn2+. We report here that Zn2+ exposure increases COX-2 expression through the CRE site located in the COX-2 promoter region and stabilization of COX-2 mRNA.
Section snippets
Materials and reagents
American Chemical Society-grade zinc sulfate, Triton X-100, and polyacrylamide were purchased from Sigma Chemical Co. (St. Louis, MO). SDS-PAGE supplies such as molecular mass standards and buffers were from Bio-Rad (Richmond, CA). Anti-human COX-2 polyclonal antibody was obtained from Cayman Chemical (Ann Arbor, MI). β-actin antibody was purchased from USBiological (Swampscott, MA). Horseradish peroxidase (HRP)-conjugated goat anti-rabbit or goat anti-mouse IgG was obtained from Santa Cruz
Zn2+ exposure increases COX-2 mRNA and protein expression, and PGE2 production in BEAS-2B cells
As demonstrated in our previous study (Wu et al., 2003), exposure of BEAS-2B cells to 50 μM Zn2+ for 8 h did not result in significant alterations in cell viability, as assessed by assay of lactate dehydrogenase activity released into the culture medium. Exposure of BEAS-2B cells to 25 or 50 μM Zn2+ for 8 h caused a marked increase in COX-2 mRNA expression (Fig. 1A). The highest dose utilized (50 μM) induced elevations in COX-2 mRNA which reached about 11 fold at 4 h of exposure and remained
Discussion
COX-2 is an important pharmacological target for the treatment of diseases ranging from inflammation to cancer. The molecular mechanisms controlling expression of COX-2 are not fully defined. This study demonstrates that Zn2+ stimulation markedly increases COX-2 mRNA expression through a combination of transcriptional activation and mRNA stabilization in a human airway epithelial cell line, BEAS-2B. This results in a marked increase in expression of functional COX-2 protein. In the context of
Acknowledgments
We greatly appreciate the technical assistance from Lisa Dailey, Drs Ilona Jaspers and William Reed. This work was supported by the United States Environmental Protection Agency Cooperative Agreement CR83346301 awarded to the Center for Environmental Medicine, Asthma and Lung Biology, University of North Carolina. The research described in this article has been reviewed by the National Health and Environmental Effects Research Laboratory and National Risk Management Laboratory, U.S. EPA, and
References (53)
- et al.
Zinc is the toxic factor in the lung response to an atmospheric particulate sample
Toxicol. Appl. Pharmacol.
(2000) - et al.
Lipoxin A(4) regulates bronchial epithelial cell responses to acid injury
Am. J. Pathol.
(2006) - et al.
Proinflammatory cytokines induce cyclooxygenase-2 mRNA and protein expression in human pulp cell cultures
J. Endod.
(2003) - et al.
The 3′-untranslated region of murine cyclooxygenase-2 contains multiple regulatory elements that alter message stability and translational efficiency
J. Biol. Chem.
(2001) - et al.
The proximal region of the 3′-untranslated region of cyclooxygenase-2 is recognized by a multimeric protein complex containing HuR, TIA-1, TIAR, and the heterogeneous nuclear ribonucleoprotein U
J. Biol. Chem.
(2003) - et al.
Post-transcriptional control of cyclooxygenase-2 gene expression. The role of the 3′-untranslated region
J. Biol. Chem.
(2000) - et al.
The induction and suppression of prostaglandin H2 synthase (cyclooxygenase) in human monocytes
J. Biol. Chem.
(1990) - et al.
Identification of TIAR as a protein binding to the translational regulatory AU–rich element of tumor necrosis factor alpha mRNA
J. Biol. Chem.
(1999) - et al.
Differentiation status-dependent regulation of cyclooxygenase-2 expression and prostaglandin E2 production by epidermal growth factor via mitogen-activated protein kinase in articular chondrocytes
J. Biol. Chem.
(2003) - et al.
AP-1 function and regulation
Curr. Opin. Cell Biol.
(1997)
Zn2+-induced NF-kappaB-dependent transcriptional activity involves site-specific p65/RelA phosphorylation
Cell. Signal.
RSV-induced prostaglandin E2 production occurs via cPLA2 activation: role in viral replication
Virology
An E-box region within the prostaglandin endoperoxide synthase-2 (PGS-2) promoter is required for transcription in rat ovarian granulosa cells
J. Biol. Chem.
Recent advances in molecular biology and physiology of the prostaglandin E2-biosynthetic pathway
Prog. Lipid Res.
Differential modulation of COX-2 expression in A549 airway epithelial cells by structurally distinct PPAR(gamma) agonists: evidence for disparate functional effects which are independent of NF-(kappa)B and PPAR(gamma)
Cell. Signal.
Selective suppression of CCAAT/enhancer-binding protein beta binding and cyclooxygenase-2 promoter activity by sodium salicylate in quiescent human fibroblasts
J. Biol. Chem.
Hypoxia induces cyclooxygenase-2 via the NF-kappaB p65 transcription factor in human vascular endothelial cells
J. Biol. Chem.
Regulation of cyclooxygenase-2 expression in monocytes by ligation of the receptor for advanced glycation end products
J. Biol. Chem.
Microtubule-interfering agents stimulate the transcription of cyclooxygenase-2. Evidence for involvement of ERK1/2 AND p38 mitogen-activated protein kinase pathways
J. Biol. Chem.
Regulation of cyclooxgenase-2 mRNA stability by taxanes: evidence for involvement of p38, MAPKAPK-2, and HuR
J. Biol. Chem.
Cyclooxygenase isozymes and their gene structures and expression
Prostaglandins Other Lipid Mediat.
Characterization of the genomic structure, chromosomal location and promoter of human prostaglandin H synthase-2 gene
Biochem. Biophys. Res. Commun.
Activation of thromboxane receptor alpha induces expression of cyclooxygenase-2 through multiple signaling pathways in A549 human lung adenocarcinoma cells
Biochem. Pharmacol.
Zinc-induced PTEN protein degradation through the proteasome pathway in human airway epithelial cells
J. Biol. Chem.
Dihydroxy bile acids activate the transcription of cyclooxygenase-2
J. Biol. Chem.
Structure of the human cyclooxygenase-2 gene
Biochem. J.
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