Solving the IRAK-4 Enigma: Application of Kinase-Dead Knock-In Mice

  • M. Koziczak-Holbro
  • C. Joyce
  • A. Glück
  • B. Kinzel
  • M. Müller
  • H. Gram
Conference paper
Part of the Ernst Schering Foundation Symposium Proceedings book series (SCHERING FOUND, volume 2007/3)


Interleukin-1 receptor-associated kinase (IRAK-4) is an essential component of the signal transduction complex downstream of the interleukin (IL)-1- and Toll-like receptors. Though regarded as the first kinase in the signaling cascade, the role of IRAK-4 kinase activity versus its scaffold function has been controversial. In order to investigate the role of IRAK-4 kinase function in vivo, we generated “knock-in” mice where the wild-type IRAK-4 gene is replaced with a mutant gene encoding kinase-deficient IRAK-4 protein (IRAK-4 KD). IRAK-4 kinase is rendered inactive by mutating the conserved lysine residues in the ATP pocket essential for coordinating ATP. Analyses of embryonic fibroblasts and macrophages obtained from IRAK-4 KD mice demonstrated lack of cellular responsiveness to stimulation with IL-1β or Toll-like receptor 4 (TLR4) and TLR7 agonists. IRAK-4 KD cells were severely impaired in NF-κB, JNK, and p38 activation in response to IL-1β or TLR7 ligand. In addition, activation of JNK and p38 was affected in lipopolysaccharide (LPS)-stimulated IRAK-4 KD macrophages. As a consequence, IL-1 receptor/TLR4/TLR7-mediated production of cytokines and chemokines was largely absent in these cells. Additionally, microarray analysis identified IL-1β response genes and revealed that the induction of IL-1β-responsive mRNAs is largely ablated in IRAK-4 KD cells. In summary, our results suggest that kinase activity plays a critical role in IL-1R-, TLR4-, and TLR7-mediated induction of inflammatory responses.


Conserve Lysine Residue Proximal Kinase 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


  1. Burns K, Janssens S, Brissoni B, Olivos N, Beyaert R, Tschopp J (2003) Inhibition of interleukin 1 receptor/Toll-like receptor signaling through the alternatively spliced, short form of MyD88 is due to its failure to recruit IRAK-4. J Exp Med 197:263–268PubMedCrossRefGoogle Scholar
  2. Cheng H, Addona T, Keshishian H, Dahlstrand E, Lu C, Dorsch M, Li Z, Wang A, Ocain TD, Li P, Parsons TF, Jaffee B, Xu Y (2007) Regulation of IRAK-4 kinase activity via autophosphorylation within its activation loop. Biochem Biophys Res Commun 352:609–616PubMedCrossRefGoogle Scholar
  3. Cohen SB (2004) The use of anakinra an interleukin-1 receptor antagonist in the treatment of rheumatoid arthritis. Rheum Dis Clin North Am 30:365–80 viiPubMedCrossRefGoogle Scholar
  4. Davis CN, Tabarean I, Gaidarova S, Behrens MM, Bartfai T (2006) IL-1beta induces a MyD88-dependent and ceramide-mediated activation of Src in anterior hypothalamic neurons. J Neurochem 98:1379–1389PubMedCrossRefGoogle Scholar
  5. Huang Q, Yang J, Lin Y, Walker C, Cheng J, Liu ZG, Su B (2004) Differential regulation of interleukin 1 receptor and Toll-like receptor signaling by MEKK3. Nat Immunol 5:98–103PubMedCrossRefGoogle Scholar
  6. Janssens S, Beyaert R (2003) Functional diversity and regulation of different interleukin-1 receptor-associated kinase (IRAK) family members. Mol Cell 11:293–302PubMedCrossRefGoogle Scholar
  7. Jiang Z, Ninomiya-Tsuji J, Qian Y, Matsumoto K, Li X (2002) Interleukin-1 (IL-1) receptor-associated kinase-dependent IL-1-induced signaling complexes phosphorylate TAK1 and TAB2 at the plasma membrane and activate TAK1 in the cytosol. Mol Cell Biol 22:7158–7167PubMedCrossRefGoogle Scholar
  8. Kawagoe T, Sato S, Jung A, Yamamoto M, Matsui K, Kato H, Uematsu S, Takeuchi O, Akira S (2007) Essential role of IRAK-4 protein and its kinase activity in Toll-like receptor-mediated immune responses but not in TCR signaling. J Exp Med 204:1013–1024PubMedCrossRefGoogle Scholar
  9. Kim TW, Staschke K, Bulek K, Yao J, Peters K, Oh KH, Vandenburg Y, Xiao H, Qian W, Hamilton T, Min B, Sen G, Gilmour R, Li X (2007) A critical role for IRAK4 kinase activity in Toll-like receptor-mediated innate immunity. J Exp Med 204:1025–1036PubMedCrossRefGoogle Scholar
  10. Kollewe C, Mackensen AC, Neumann D, Knop J, Cao P, Li S, Wesche H, Martin MU (2004) Sequential autophosphorylation steps in the interleukin-1 receptor-associated kinase-1 regulate its availability as an adapter in interleukin-1 signaling. J Biol Chem 279:5227–5236PubMedCrossRefGoogle Scholar
  11. Koziczak-Holbro M, Joyce C, Glück A, Kinzel B, Müller M, Tschopp C, Mathison JC, Davis CN, Gram H (2007) IRAK-4 kinase activity is required for interleukin-1 (IL-1) receptor- and toll-like receptor 7-mediated signaling and gene expression. J Biol Chem 282:13552–13560PubMedCrossRefGoogle Scholar
  12. Li S, Strelow A, Fontana EJ, Wesche H (2002) IRAK-4: a novel member of the IRAK family with the properties of an IRAK-kinase. Proc Natl Acad Sci USA 99:5567–5572PubMedCrossRefGoogle Scholar
  13. Lye E, Mirtsos C, Suzuki N, Suzuki S, Yeh WC (2004) The role of interleukin 1 receptor-associated kinase-4 (IRAK-4) kinase activity in IRAK-4-mediated signaling. J Biol Chem 279:40653–40658PubMedCrossRefGoogle Scholar
  14. Martin MU, Wesche H (2002) Summary and comparison of the signaling mechanisms of the Toll/interleukin-1 receptor family. Biochim Biophys Acta 1592:265–280PubMedCrossRefGoogle Scholar
  15. Means TK, Golenbock DT, Fenton MJ (2000) The biology of Toll-like receptors. Cytokine Growth Factor Rev 11:219–232PubMedCrossRefGoogle Scholar
  16. Medvedev AE, Thomas K, Awomoyi A, Kuhns DB, Gallin JI, Li X, Vogel SN (2005) Cutting edge: expression of IL-1 receptor-associated kinase-4 (IRAK-4) proteins with mutations identified in a patient with recurrent bacterial infections alters normal IRAK-4 interaction with components of the IL-1 receptor complex. J Immunol 174:6587–6591PubMedGoogle Scholar
  17. Picard C, Puel A, Bonnet M, Ku CL, Bustamante J, Yang K, Soudais C, Dupuis S, Feinberg J, Fieschi C, Elbim C, Hitchcock R, Lammas D, Davies G, Al-Ghonaium A, Al-Rayes H, Al-Jumaah S, Al-Hajjar S, Al-Mohsen IZ, Frayha HH, Rucker R, Hawn TR, Aderem A, Tufenkeji H, Haraguchi S, Day NK, Good RA, Gougerot-Pocidalo MA, Ozinsky A, Casanova JL (2003) Pyogenic bacterial infections in humans with IRAK-4 deficiency. Science 299:2076–2079PubMedCrossRefGoogle Scholar
  18. Qin J, Jiang Z, Qian Y, Casanova JL, Li X (2004) IRAK4 kinase activity is redundant for interleukin-1 (IL-1) receptor-associated kinase phosphorylation and IL-1 responsiveness. J Biol Chem 279:26748–26753PubMedCrossRefGoogle Scholar
  19. Suzuki N, Suzuki S, Duncan GS, Millar DG, Wada T, Mirtsos C, Takada H, Wakeham A, Itie A, Li S, Penninger JM, Wesche H, Ohashi PS, Mak TW, Yeh WC (2002a) Severe impairment of interleukin-1 and Toll-like receptor signalling in mice lacking IRAK-4. Nature 416:750–756PubMedCrossRefGoogle Scholar
  20. Suzuki N, Suzuki S, Yeh WC (2002b) IRAK-4 as the central TIR signaling mediator in innate immunity. Trends Immunol 23:503–506PubMedCrossRefGoogle Scholar
  21. Tak PP, Firestein GS (2001) NF-kappaB: a key role in inflammatory diseases. J Clin Invest 107:7–11PubMedCrossRefGoogle Scholar
  22. Takeda K, Akira S (2004) TLR signaling pathways. Semin Immunol 16:3–9PubMedCrossRefGoogle Scholar
  23. Takeda K, Akira S (2005) Toll-like receptors in innate immunity. Int Immunol 17:1–14PubMedCrossRefGoogle Scholar
  24. van den Berg WB (2001) Uncoupling of inflammatory and destructive mechanisms in arthritis. Semin Arthritis Rheum 30:7–16PubMedCrossRefGoogle Scholar
  25. Wang C, Deng L, Hong M, Akkaraju GR, Inoue J, Chen ZJ (2001) TAK1 is a ubiquitin-dependent kinase of MKK and IKK. Nature 412:346–351PubMedCrossRefGoogle Scholar
  26. Wang Z, Liu J, Sudom A, Ayres M, Li S, Wesche H, Powers JP, Walker NP (2006) Crystal structures of IRAK-4 kinase in complex with inhibitors: a serine/threonine kinase with tyrosine as a gatekeeper. Structure 14:1835–1844PubMedCrossRefGoogle Scholar
  27. Wisdom R (1999) AP-1: one switch for many signals. Exp Cell Res 253:180–185PubMedCrossRefGoogle Scholar
  28. Yamin TT, Miller DK (1997) The interleukin-1 receptor-associated kinase is degraded by proteasomes following its phosphorylation. J Biol Chem 272:21540–21547PubMedCrossRefGoogle Scholar
  29. Zheng J, Knighton DR, ten Eyck LF, Karlsson R, Xuong N, Taylor SS, Sowadski JM (1993) Crystal structure of the catalytic subunit of cAMP-dependent protein kinase complexed with MgATP and peptide inhibitor. Biochemistry 32:2154–2161PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2008

Authors and Affiliations

  • M. Koziczak-Holbro
    • 1
  • C. Joyce
    • 1
  • A. Glück
    • 1
  • B. Kinzel
    • 1
  • M. Müller
    • 1
  • H. Gram
    • 1
  1. 1.Novartis Institutes for BioMedical ResearchBaselSwitzerland

Personalised recommendations