title: IFN-regulatory factor 3-dependent gene expression is defective in Tbk1-deficient mouse embryonic fibroblasts.
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.
authors.affiliation: Department of Molecular and Cellular Biology, Harvard University, 7 Divinity Avenue, Cambridge, MA 02138, USA.
authors.forename: Sarah M
authors.initials: SM
authors.lastname: McWhirter
authors.forename: Katherine A
authors.initials: KA
authors.lastname: Fitzgerald
authors.forename: Jacqueline
authors.initials: J
authors.lastname: Rosains
authors.forename: Daniel C
authors.initials: DC
authors.lastname: Rowe
authors.forename: Douglas T
authors.initials: DT
authors.lastname: Golenbock
authors.forename: Tom
authors.initials: T
authors.lastname: Maniatis
mesh_terms: D000818:Animals; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D005347:Fibroblasts / Q000276:immunology / Q000378:metabolism; D015870:Gene Expression; D051550:I-kappa B Kinase; D066298:In Vitro Techniques; D050838:Interferon Regulatory Factor-3; D016899:Interferon-beta / Q000235:genetics; D051379:Mice; D018345:Mice, Knockout; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000172:deficiency* / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D014777:Virus Diseases / Q000235:genetics / Q000276:immunology / Q000378:metabolism
chemical_list: D004268:DNA-Binding Proteins; D050838:Interferon Regulatory Factor-3; C494233:Irf3 protein, mouse; D011994:Recombinant Proteins; D014157:Transcription Factors; D016899:Interferon-beta; C498588:Tbk1 protein, mouse; D017346:Protein Serine-Threonine Kinases; C496564:Chuk protein, mouse; D051550:I-kappa B Kinase; C496562:Ikbkb protein, mouse; C496566:Ikbke protein, mouse