title: Leukotriene B(4) BLT receptor signaling regulates the level and stability of cyclooxygenase-2 (COX-2) mRNA through restricted activation of Ras/Raf/ERK/p42 AUF1 pathway.
abstract: Recent studies suggest that active resolution of the inflammatory response in animal models of arthritis may involve leukotriene B(4) (LTB(4))-dependent stimulation of "intermediate" prostaglandin production, which in turn favors the synthesis of "downstream" anti-inflammatory and pro-resolving lipoxins, resolvins, and protectins. We explored a putative mechanism involving LTB(4)-dependent control of cyclooxygenase-2 (COX-2) expression, the rate-limiting step in inflammatory prostaglandin biosynthesis. Indeed, LTB(4) potently up-regulated/stabilized interleukin-1beta-induced COX-2 mRNA and protein expression under conditions of COX-2 inhibitor-dependent blockade of PGE(2) release in human synovial fibroblasts (EC(50) = 16.5 + or - 1.7 nm for mRNA; 19 + or - 2.4 nm for protein, n = 4). The latter response was pertussis toxin-sensitive, and semi-quantitative reverse transcription-PCR confirmed the quantitative predominance of the BLT2 receptor. Transfection experiments, using human COX-2 promoter plasmids and chimeric luciferase-COX-2 mRNA 3'-untranslated region (3'-UTR) reporter constructs, revealed that LTB(4) exerted its stabilizing effect at the post-transcriptional level through a 116-bp adenylate/uridylate-rich sequence in the proximal region of the COX-2 3'-UTR. Using luciferase-COX-2 mRNA 3'-UTR reporter constructs and Ras/c-Raf expression and mutant constructs, we showed that the Ras/c-Raf/MEK1/2/ERK1/2 signaling pathway mediated LTB(4)-dependent COX-2 mRNA stabilization. Knockdown experiments with specific short hairpin RNAs confirmed that LTB(4) stabilization of COX-2 mRNA was apparently mediated through the RNA-binding protein, p42 AUF1. The nuclear export of p42 AUF1 was driven by c-Raf/MEK1/2/ERK1/2 signaling and sensitive to leptomycin B treatment, suggesting a CRM1-dependent mechanism. We conclude that LTB(4) may support the resolution phase of the inflammatory response by stabilizing COX-2, ensuring a reservoir of ambient pro-resolution lipid mediators.
authors.affiliation: Department of Medicine, McGill University, Montreal, Quebec H3A 1A1.
authors.forename: Beibei
authors.initials: B
authors.lastname: Zhai
authors.forename: Huiqing
authors.initials: H
authors.lastname: Yang
authors.forename: Arturo
authors.initials: A
authors.lastname: Mancini
authors.forename: QingWen
authors.initials: Q
authors.lastname: He
authors.forename: John
authors.initials: J
authors.lastname: Antoniou
authors.forename: John A
authors.initials: JA
authors.lastname: Di Battista
mesh_terms: D020413:3' Untranslated Regions; D018844:Cyclin-Dependent Kinases / Q000378:metabolism; D051546:Cyclooxygenase 2 / Q000378:metabolism*; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism; D006367:HeLa Cells; D000081035:Heterogeneous Nuclear Ribonucleoprotein D0; D034502:Heterogeneous-Nuclear Ribonucleoprotein D / Q000378:metabolism*; D006801:Humans; D007975:Leukotriene B4 / Q000737:chemistry*; D008055:Lipids / Q000737:chemistry; D018102:Receptors, Leukotriene B4 / Q000378:metabolism; D015398:Signal Transduction; D013583:Synovial Membrane / Q000166:cytology*; D048490:raf Kinases / Q000378:metabolism*; D018631:ras Proteins / Q000378:metabolism*; D000095043:Cyclin-Dependent Kinase-Activating Kinase
chemical_list: D020413:3' Untranslated Regions; C000630986:HNRNPD protein, human; D000081035:Heterogeneous Nuclear Ribonucleoprotein D0; D034502:Heterogeneous-Nuclear Ribonucleoprotein D; C513240:LTB4R protein, human; D008055:Lipids; D018102:Receptors, Leukotriene B4; D007975:Leukotriene B4; D051546:Cyclooxygenase 2; D048490:raf Kinases; D018844:Cyclin-Dependent Kinases; D048049:Extracellular Signal-Regulated MAP Kinases; D018631:ras Proteins; D000095043:Cyclin-Dependent Kinase-Activating Kinase