IFN-regulatory factor 3-dependent gene expression is defective in Tbk1-deficient mouse embryonic fibroblasts

Proc Natl Acad Sci U S A. 2004 Jan 6;101(1):233-8. doi: 10.1073/pnas.2237236100. Epub 2003 Dec 16.

Abstract

Virus infection, double-stranded RNA, and lipopolysaccharide each induce the expression of genes encoding IFN-alpha and -beta and chemokines, such as RANTES (regulated on activation, normal T cell expressed and secreted) and IP-10 (IFN-gamma inducible protein 10). This induction requires the coordinate activation of several transcription factors, including IFN-regulatory factor 3 (IRF3). The signaling pathways leading to IRF3 activation are triggered by the binding of pathogen-specific products to Toll-like receptors and culminate in the phosphorylation of specific serine residues in the C terminus of IRF3. Recent studies of human cell lines in culture have implicated two noncanonical IkappaB kinase (IKK)-related kinases, IKK-epsilon and Traf family member-associated NF-kappaB activator (TANK)-binding kinase 1 (TBK1), in the phosphorylation of IRF3. Here, we show that purified recombinant IKK-epsilon and TBK1 directly phosphorylate the critical serine residues in IRF3. We have also examined the expression of IRF3-dependent genes in mouse embryonic fibroblasts (MEFs) derived from Tbk1(-/-) mice, and we show that TBK1 is required for the activation and nuclear translocation of IRF3 in these cells. Moreover, Tbk1(-/-) MEFs show marked defects in IFN-alpha and -beta, IP-10, and RANTES gene expression after infection with either Sendai or Newcastle disease viruses or after engagement of the Toll-like receptors 3 and 4 by double-stranded RNA and lipopolysaccharide, respectively. Finally, TRIF (TIR domain-containing adapter-inducing IFN-beta), fails to activate IRF3-dependent genes in Tbk1(-/-) MEFs. We conclude that TBK1 is essential for IRF3-dependent antiviral gene expression.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • DNA-Binding Proteins / chemistry
  • DNA-Binding Proteins / genetics*
  • DNA-Binding Proteins / metabolism
  • Fibroblasts / immunology
  • Fibroblasts / metabolism
  • Gene Expression
  • I-kappa B Kinase
  • In Vitro Techniques
  • Interferon Regulatory Factor-3
  • Interferon-beta / genetics
  • Mice
  • Mice, Knockout
  • Phosphorylation
  • Protein-Serine-Threonine Kinases / deficiency*
  • Protein-Serine-Threonine Kinases / genetics
  • Protein-Serine-Threonine Kinases / metabolism
  • Recombinant Proteins / metabolism
  • Transcription Factors / chemistry
  • Transcription Factors / genetics*
  • Transcription Factors / metabolism
  • Virus Diseases / genetics
  • Virus Diseases / immunology
  • Virus Diseases / metabolism

Substances

  • DNA-Binding Proteins
  • Interferon Regulatory Factor-3
  • Irf3 protein, mouse
  • Recombinant Proteins
  • Transcription Factors
  • Interferon-beta
  • Tbk1 protein, mouse
  • Protein-Serine-Threonine Kinases
  • Chuk protein, mouse
  • I-kappa B Kinase
  • Ikbkb protein, mouse
  • Ikbke protein, mouse