Inhibitors of the proteasome pathway interfere with induction of nitric oxide synthase in macrophages by blocking activation of transcription factor NF-κB

Jeanette M. Griscavage, Sherwin Wilk, Louis J. Ignarro

Research output: Contribution to journalArticlepeer-review

248 Scopus citations

Abstract

The objective of this study was to elucidate the role of the proteasome pathway or multicatalytic proteinase complex in the induction of immunologic nitric oxide (NO) synthase (iNOS) in rat alveolar macrophages activated by lipopolysaccharide. Macrophages were incubated in the presence of lipopolysaccharide plus test agent for up to 24 hr. Culture media were analyzed for accumulation of stable oxidation products of NO (NO2- + NO3- , designated as NO(X)-), cellular RNA was extracted for determination of iNOS mRNA levels by Northern blot analysis, and nuclear extracts were prepared for determination of NF-κB by electrophoretic mobility-shift assay. Inhibitors of calpain (α-N-acetyl-Leu-Leu-norleucinal; N-benzyloxycarbonyl- Leu-leucinal) and the proteasome (N-benzyloxycarbonyl-Ile-Glu-(O-t-Bu)-Ala- leucinal) markedly inhibited or abolished the induction of iNOS in macrophages. The proteinase inhibitors interfered with lipopolysaccharide- induced NO(X)- production by macrophages, and this effect was accompanied by comparable interference with the appearance of both iNOS mRNA and NF-κB. Calpain inhibitors elicited effects at concentrations of 1-100 μM, whereas the proteasome inhibitor was 1000-fold more potent, producing significant inhibitory effects at 1 nM. The present findings indicate that the proteasome pathway is essential for lipopolysaccharide-induced expression of the iNOS gene in rat alveolar macrophages. Furthermore, the data support the view that the proteasome pathway is directly involved in promoting the activation of NF-κB and that the induction of iNOS by lipopolysaccharide involves the transcriptional action of NF-κB.

Original languageEnglish
Pages (from-to)3308-3312
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume93
Issue number8
DOIs
StatePublished - 16 Apr 1996

Keywords

  • NO synthase induction
  • calpain
  • multicatalytic proteinase complex
  • proteinase inhibitors
  • transcription factors

Fingerprint

Dive into the research topics of 'Inhibitors of the proteasome pathway interfere with induction of nitric oxide synthase in macrophages by blocking activation of transcription factor NF-κB'. Together they form a unique fingerprint.

Cite this