title: Role of the Rho GTPase in bradykinin-stimulated nuclear factor-kappaB activation and IL-1beta gene expression in cultured human epithelial cells.
abstract: Recent evidence suggests a novel role of bradykinin (BK) in stimulating gene transcription. This study examined the effect of BK on nuclear factor kappaB (NF-kappaB) activation and IL-1beta synthesis in human epithelial cells. Stimulation of A549 cells and primary bronchial epithelial cells with BK rapidly activated NF-kappaB. BK also increased the level of secreted immunoreactive IL-1beta in A549 culture supernatants, an effect that was blocked by actinomycin D and the B2 BK receptor antagonist HOE-140. The role of NF-kappaB activation in BK-induced IL-1beta synthesis was demonstrated by the ability of BK to stimulate increased chloramphenicol acetyltransferase (CAT) activity in A549 cells transfected with a reporter plasmid containing three kappaB enhancers from the IL-1beta gene, while deletion of the kappaB enhancer sequences eliminated BK-stimulated CAT activity. C3 transferase exoenzyme, an inhibitor of Rho, abolished BK-induced NF-kappaB activation at 10 microg/ml and significantly inhibited BK-stimulated IL-1beta synthesis at 5 microg/ml. A dominant-negative form of RhoA (T19N) inhibited BK-stimulated reporter gene expression in a dose-dependent and kappaB-dependent manner. Cotransfection of A549 cells with an expression vector encoding a constitutively active form of RhoA (Q63L) along with the IL-1beta promoter-CAT reporter plasmid resulted in a marked increase in NF-kappaB activity compared with transfection with the IL-1beta promoter-CAT reporter plasmid alone. These results demonstrate that BK stimulates NF-kappaB activation and IL-1beta synthesis in A549 cells, and that RhoA is both necessary and sufficient to mediate this effect.
authors.affiliation: Department of Molecular and Experimental Medicine, The Scripps Research Institute, La Jolla, CA 92037, USA.
authors.forename: Z K
authors.initials: ZK
authors.lastname: Pan
authors.forename: R D
authors.initials: RD
authors.lastname: Ye
authors.forename: S C
authors.initials: SC
authors.lastname: Christiansen
authors.forename: M A
authors.initials: MA
authors.lastname: Jagels
authors.forename: G M
authors.initials: GM
authors.lastname: Bokoch
authors.forename: B L
authors.initials: BL
authors.lastname: Zuraw
mesh_terms: D001920:Bradykinin / Q000494:pharmacology*; D001980:Bronchi / Q000378:metabolism; D002478:Cells, Cultured; D015500:Chloramphenicol O-Acetyltransferase / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D004847:Epithelial Cells / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000502:physiology*; D006801:Humans; D007375:Interleukin-1 / Q000096:biosynthesis / Q000235:genetics*; D016328:NF-kappa B / Q000378:metabolism*; D020742:rhoA GTP-Binding Protein
chemical_list: D007375:Interleukin-1; D016328:NF-kappa B; D004247:DNA; D015500:Chloramphenicol O-Acetyltransferase; D019204:GTP-Binding Proteins; D020742:rhoA GTP-Binding Protein; D001920:Bradykinin