title: Rapid Up-regulation of IkappaBbeta and abrogation of NF-kappaB activity in peritoneal macrophages stimulated with lipopolysaccharide.
abstract: Lipopolysaccharide (LPS) administration to mice elicited the activation of nuclear factor kappaB (NF-kappaB) in several tissues including liver and macrophages. Maximal activation was observed 1 h after treatment but declined at 3 and 6 h. The levels of IkappaBalpha and IkappaBbeta were analyzed during this period in an attempt to correlate NF-kappaB activity with IkappaB resynthesis. Degradation of IkappaBalpha was very rapid and was followed by recovery 1 h after LPS administration. IkappaBbeta degradation, which has been associated with persistent NF-kappaB activation, was complete at 1 h. However, a rapid recovery of IkappaBbeta in these tissues was observed at 3 h in parallel with the abrogation of NF-kappaB activity. Immunolocalization of newly synthesized IkappaBbeta by confocal microscopy revealed its preferential accumulation in the cytosol. Analysis of IkappaBbeta by Western blot using high resolution polyacrylamide gel electrophoresis showed the presence of two bands in cytosolic extracts of LPS-treated macrophages at 3 h, but only one band with the same mobility as the control was detected at 6 h. Moreover, treatment of extracts of resynthesized IkappaBbeta with alkaline phosphatase resulted in the accumulation of the protein of slightly higher electrophoretic mobility, indicating the prevalence of a rapid phosphorylation of the newly synthesized IkappaBbeta. At the mRNA level, up-regulation of IkappaBbeta was observed in macrophages stimulated for 1 h with LPS. When the effect of pro-inflammatory cytokines was investigated, tumor necrosis factor alpha, but not interleukin-1 or interferon-gamma, promoted an important degradation of IkappaBbeta followed by an increase in the mRNA at 1 h. These results suggest the existence of LPS- and tumor necrosis factor alpha- specific pathways involved in a rapid IkappaBbeta degradation and resynthesis and might explain the transient period of activation of NF-kappaB in these tissues upon stimulation with these factors. This rapid control of NF-kappaB function may contribute to the attenuation of the inflammatory response of these cells.
authors.affiliation: Instituto de Bioquímica (Consejo Superior de Investigaciones Cientificas), Facultad de Farmacia, Universidad Complutense, 28040 Madrid, Spain.
authors.forename: M
authors.initials: M
authors.lastname: Velasco
authors.forename: M J
authors.initials: MJ
authors.lastname: Díaz-Guerra
authors.forename: P
authors.initials: P
authors.lastname: Martín-Sanz
authors.forename: A
authors.initials: A
authors.lastname: Alvarez
authors.forename: L
authors.initials: L
authors.lastname: Boscá
mesh_terms: D000210:Acute-Phase Reaction / Q000378:metabolism; D000818:Animals; D002451:Cell Compartmentation; D002467:Cell Nucleus / Q000378:metabolism; D002478:Cells, Cultured; D003600:Cytosol / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000096:biosynthesis* / Q000235:genetics; D020672:I-kappa B Proteins; D008070:Lipopolysaccharides / Q000494:pharmacology; D017737:Macrophages, Peritoneal / Q000187:drug effects / Q000378:metabolism*; D051379:Mice; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000378:metabolism*; D012333:RNA, Messenger / Q000096:biosynthesis; D012772:Shock, Septic / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D015854:Up-Regulation
chemical_list: D004268:DNA-Binding Proteins; C083487:I kappa B beta protein; D020672:I-kappa B Proteins; D008070:Lipopolysaccharides; D016328:NF-kappa B; C000606205:Nfkbia protein, mouse; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D000072000:NF-KappaB Inhibitor alpha