Endotoxin tolerance impairs IL-1 receptor-associated kinase (IRAK) 4 and TGF-beta-activated kinase 1 activation, K63-linked polyubiquitination and assembly of IRAK1, TNF receptor-associated factor 6, and IkappaB kinase gamma and increases A20 expression

J Biol Chem. 2011 Mar 11;286(10):7905-7916. doi: 10.1074/jbc.M110.182873. Epub 2011 Jan 10.


Endotoxin tolerance reprograms Toll-like receptor 4 responses by impairing LPS-elicited production of pro-inflammatory cytokines without inhibiting expression of anti-inflammatory or anti-microbial mediators. In septic patients, Toll-like receptor tolerance is thought to underlie decreased pro-inflammatory cytokine expression in response to LPS and increased incidence of microbial infections. The impact of endotoxin tolerance on recruitment, post-translational modifications and signalosome assembly of IL-1 receptor-associated kinase (IRAK) 4, IRAK1, TNF receptor-associated factor (TRAF) 6, TGF-β-activated kinase (TAK) 1, and IκB kinase (IKK) γ is largely unknown. We report that endotoxin tolerization of THP1 cells and human monocytes impairs LPS-mediated receptor recruitment and activation of IRAK4, ablates K63-linked polyubiquitination of IRAK1 and TRAF6, compromises assembly of IRAK1-TRAF6 and IRAK1-IKKγ platforms, and inhibits TAK1 activation. Deficiencies in these signaling events in LPS-tolerant cells coincided with increased expression of A20, an essential deubiquitination enzyme, and sustained A20-IRAK1 associations. Overexpression of A20 inhibited LPS-induced activation of NF-κB and ablated NF-κB reporter activation driven by ectopic expression of MyD88, IRAK1, IRAK2, TRAF6, and TAK1/TAB1, while not affecting the responses induced by IKKβ and p65. A20 shRNA knockdown abolished LPS tolerization of THP1 cells, mechanistically linking A20 and endotoxin tolerance. Thus, deficient LPS-induced activation of IRAK4 and TAK1, K63-linked polyubiquitination of IRAK1 and TRAF6, and disrupted IRAK1-TRAF6 and IRAK1-IKKγ assembly associated with increased A20 expression and A20-IRAK1 interactions are new determinants of endotoxin tolerance.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Cell Line
  • DNA-Binding Proteins
  • Drug Resistance / drug effects*
  • Gene Expression Regulation / drug effects*
  • Humans
  • I-kappa B Kinase / genetics
  • I-kappa B Kinase / metabolism*
  • Interleukin-1 Receptor-Associated Kinases / genetics
  • Interleukin-1 Receptor-Associated Kinases / metabolism*
  • Intracellular Signaling Peptides and Proteins / genetics
  • Lipopolysaccharides / pharmacology*
  • MAP Kinase Kinase Kinases / genetics
  • MAP Kinase Kinase Kinases / metabolism*
  • Monocytes / metabolism*
  • Myeloid Differentiation Factor 88 / genetics
  • Myeloid Differentiation Factor 88 / metabolism
  • Nuclear Proteins / biosynthesis*
  • Nuclear Proteins / genetics
  • TNF Receptor-Associated Factor 6 / genetics
  • TNF Receptor-Associated Factor 6 / metabolism*
  • Tumor Necrosis Factor alpha-Induced Protein 3
  • Ubiquitination / drug effects*


  • DNA-Binding Proteins
  • Intracellular Signaling Peptides and Proteins
  • Lipopolysaccharides
  • MYD88 protein, human
  • Myeloid Differentiation Factor 88
  • Nuclear Proteins
  • TNF Receptor-Associated Factor 6
  • IRAK1 protein, human
  • IRAK4 protein, human
  • Interleukin-1 Receptor-Associated Kinases
  • I-kappa B Kinase
  • MAP Kinase Kinase Kinases
  • MAP kinase kinase kinase 7
  • TNFAIP3 protein, human
  • Tumor Necrosis Factor alpha-Induced Protein 3