title: Ca2+/calmodulin-dependent protein phosphatase calcineurin mediates the expression of iNOS through IKK and NF-kappaB activity in LPS-stimulated mouse peritoneal macrophages and RAW 264.7 cells.
abstract: Ca(2+) and Ca(2+)/calmodulin-dependent protein phosphatase calcineurin (CN) have been known to play crucial roles in immune response and inflammation. Using mouse peritoneal macrophages and RAW 264.7 macrophage cells, we demonstrated that LPS mobilized intracellular free Ca(2+) and induced CN phosphatase activity. iNOS expression and NO secretion in response to LPS were suppressed by Ca(2+) antagonists (TMB-8, BAPTA/AM, and nifedipine) and CN inhibitor (cyclosporin A). Transient expression of constitutively active CN in mouse peritoneal macrophages and RAW 264.7 macrophages strongly activated NF-kappaB, a key mediator of iNOS expression. We also found that CN mediates NF-kappaB activation via IkappaB-alpha hyperphosphorylation and degradation. Overexpression of dominant negative mutant of IKKalpha and -beta demonstrates that only IKKbeta is the target for CN. These results indicate that CN is required for full iNOS expression and the effective activation of NF-kappaB in RAW 264.7 and peritoneal macrophages.
authors.affiliation: Korea NanoBioTechnology Center, College of Natural Sciences, Pusan National University, 609-735, Busan, Republic of Korea.
authors.forename: YoungHee
authors.initials: Y
authors.lastname: Kim
authors.forename: Ji Sun
authors.initials: JS
authors.lastname: Moon
authors.forename: Kyoung Soon
authors.initials: KS
authors.lastname: Lee
authors.forename: Sun Young
authors.initials: SY
authors.lastname: Park
authors.forename: JaeHun
authors.initials: J
authors.lastname: Cheong
authors.forename: Ho Sung
authors.initials: HS
authors.lastname: Kang
authors.forename: Hak Young
authors.initials: HY
authors.lastname: Lee
authors.forename: Han Do
authors.initials: HD
authors.lastname: Kim
mesh_terms: D000818:Animals; D019703:Calcineurin / Q000378:metabolism*; D065095:Calcineurin Inhibitors; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D002614:Chelating Agents / Q000494:pharmacology; D016572:Cyclosporine / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D017930:Genes, Reporter / Q000187:drug effects / Q000235:genetics; D051550:I-kappa B Kinase; D008070:Lipopolysaccharides / Q000494:pharmacology*; D017737:Macrophages, Peritoneal / Q000187:drug effects / Q000201:enzymology*; D051379:Mice; D008810:Mice, Inbred C57BL; D016328:NF-kappa B / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D009569:Nitric Oxide / Q000096:biosynthesis; D019001:Nitric Oxide Synthase / Q000096:biosynthesis* / Q000235:genetics; D052247:Nitric Oxide Synthase Type II; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D014162:Transfection
chemical_list: D065095:Calcineurin Inhibitors; D002614:Chelating Agents; D008070:Lipopolysaccharides; D016328:NF-kappa B; D009569:Nitric Oxide; D016572:Cyclosporine; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496317:Nos2 protein, mouse; D017346:Protein Serine-Threonine Kinases; C496564:Chuk protein, mouse; D051550:I-kappa B Kinase; C496562:Ikbkb protein, mouse; C496566:Ikbke protein, mouse; D019703:Calcineurin; D002118:Calcium