Inhibition of NF-kappa B by S-nitrosylation

Biochemistry. 2001 Feb 13;40(6):1688-93. doi: 10.1021/bi002239y.


It is not clear if redox regulation of transcription is the consequence of direct redox-related modifications of transcription factors, or if it occurs at some other redox-sensitive step. One obstacle has been the inability to demonstrate redox-related modifications of transcription factors in vivo. The redox-sensitive transcriptional activator NF-kappaB (p50-p65) is a case in point. Its activity in vitro can be inhibited by S-nitrosylation of a critical thiol in the DNA-interacting p50 subunit, but modulation of NF-kappaB activity by nitric oxide synthase (NOS) has been attributed to other mechanisms. Herein we show that cellular NF-kappaB activity is in fact regulated by S-nitrosylation. We observed that both S-nitrosocysteine and cytokine-activated NOS2 inhibited NF-kappaB in human respiratory cells or murine macrophages. This inhibition was reversed by addition of the denitrosylating agent dithiothreitol to cellular extracts, whereas NO bioactivity did not affect the TNFalpha-induced degradation of IkappaBalpha or the nuclear translocation of p65. Recapitulation of these conditions in vitro resulted in S-nitrosylation of recombinant p50, thereby inhibiting its binding to DNA, and this effect was reversed by dithiothreitol. Further, an increase in S-nitrosylated p50 was detected in cells, and the level was modulated by TNFalpha. Taken together, these data suggest that S-nitrosylation of p50 is a physiological mechanism of NF-kappaB regulation.

MeSH terms

  • Active Transport, Cell Nucleus / drug effects
  • Animals
  • Cell Line
  • Cysteine / analogs & derivatives*
  • Cysteine / metabolism*
  • Cysteine / pharmacology
  • Cytokines / pharmacology
  • DNA / antagonists & inhibitors
  • DNA / metabolism
  • DNA-Binding Proteins / antagonists & inhibitors
  • DNA-Binding Proteins / metabolism
  • Down-Regulation / drug effects
  • Enzyme Inhibitors / pharmacology
  • Humans
  • I-kappa B Proteins*
  • Mercaptoethanol*
  • Mice
  • NF-KappaB Inhibitor alpha
  • NF-kappa B / antagonists & inhibitors*
  • NF-kappa B / genetics
  • NF-kappa B / isolation & purification
  • NF-kappa B / metabolism*
  • NF-kappa B / physiology
  • NF-kappa B p50 Subunit
  • Nitric Oxide Synthase / antagonists & inhibitors
  • Nitric Oxide Synthase / metabolism
  • Nitrosation / drug effects
  • Nitroso Compounds / metabolism*
  • Nitroso Compounds / pharmacology
  • Protein Binding / drug effects
  • S-Nitrosothiols*
  • Transcription, Genetic / drug effects
  • Transfection


  • Cytokines
  • DNA-Binding Proteins
  • Enzyme Inhibitors
  • I-kappa B Proteins
  • NF-kappa B
  • NF-kappa B p50 Subunit
  • NFKBIA protein, human
  • Nfkbia protein, mouse
  • Nitroso Compounds
  • S-Nitrosothiols
  • NF-KappaB Inhibitor alpha
  • Mercaptoethanol
  • S-nitrosomercaptoethanol
  • DNA
  • S-nitrosocysteine
  • Nitric Oxide Synthase
  • Cysteine