title: Involvement of Ras/Raf-1/p44/42 MAPK in YC-1-induced cyclooxygenase-2 expression in human pulmonary epithelial cells.
abstract: Our previous study demonstrated that 3-(5'-hydroxymethyl-2'-furyl)-1-benzylindazole (YC-1) might activate the soluble guanylate cyclase (sGC)/cGMP/protein kinase G (PKG) pathway to induce cyclooxygenase-2 (COX-2) expression in human pulmonary epithelial cells (A549). In this study, we further investigated the role of Raf-1 in YC-1-induced nuclear factor-kappaB (NF-kappaB) activation and COX-2 expression in A549 cells. YC-1-induced COX-2 expression was attenuated by a Raf-1 inhibitor (GW 5074) in a concentration-dependent manner. Treatment of A549 cells with YC-1 or 8-bromo-cGMP, a cell-permeable cGMP analogue, induced Raf-1 Ser338 phosphorylation in a time-dependent manner. YC-1-mediated Raf-1 activation was inhibited by an sGC inhibitor (ODQ), a PKG inhibitor (KT-5823), a Ras inhibitor (manumycin A), a dominant negative Ras mutant (RasN17), a protein kinase C-alpha (PKC-alpha) inhibitor (Ro 32-0432), and a phosphoinositide-3-OH-kinase (PI3K) inhibitor (LY 294002). Pretreatment of A549 cells with either manumycin A or GW 5074 attenuated YC-1-induced p44/42 MAPK activation. The YC-1-mediated increase in IKKalpha/beta activation and kappaB-luciferase activity were attenuated by GW 5074, a MAPK/ERK kinase (MEK) inhibitor (PD 98059), and an ERK2 inhibitor (AG 126). Furthermore, YC-1-induced COX-2 promoter activity was also inhibited by GW 5074, PD 98059, and AG 126. These results indicate that YC-1 might activate the sGC/cGMP/PKG pathway to elicit Ras/Raf-1/p44/42 MAPK activation, which in turn induces IKKalpha/beta and NF-kappaB activation, and ultimately causes COX-2 expression in A549 cells. Moreover, PKC-alpha and PI3K signal might be involved in YC-1-induced Raf-1 activation.
authors.affiliation: Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei 110, Taiwan.
authors.forename: Ming-Shyan
authors.initials: MS
authors.lastname: Chang
authors.forename: Bing-Chang
authors.initials: BC
authors.lastname: Chen
authors.forename: Chih-Ming
authors.initials: CM
authors.lastname: Weng
authors.forename: Wen-Sen
authors.initials: WS
authors.lastname: Lee
authors.forename: Chien-Huang
authors.initials: CH
authors.lastname: Lin
mesh_terms: D045744:Cell Line, Tumor; D051546:Cyclooxygenase 2 / Q000235:genetics* / Q000378:metabolism; D020536:Enzyme Activators / Q000494:pharmacology*; D004847:Epithelial Cells / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D051550:I-kappa B Kinase / Q000378:metabolism; D007191:Indazoles / Q000494:pharmacology*; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D016328:NF-kappa B / Q000235:genetics* / Q000378:metabolism; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism*; D011650:Pulmonary Alveoli / Q000166:cytology; D015398:Signal Transduction; D018631:ras Proteins / Q000378:metabolism
chemical_list: D020536:Enzyme Activators; D007191:Indazoles; D016328:NF-kappa B; C090937:3-(5'-hydroxymethyl-2'-furyl)-1-benzylindazole; D051546:Cyclooxygenase 2; D019908:Proto-Oncogene Proteins c-raf; C496559:CHUK protein, human; D051550:I-kappa B Kinase; C496560:IKBKB protein, human; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D018631:ras Proteins