title: Functional dissection of the TBK1 molecular network.
abstract: TANK-binding kinase 1 (TBK1) and inducible IκB-kinase (IKK-i) are central regulators of type-I interferon induction. They are associated with three adaptor proteins called TANK, Sintbad (or TBKBP1) and NAP1 (or TBKBP2, AZI2) whose functional relationship to TBK1 and IKK-i is poorly understood. We performed a systematic affinity purification-mass spectrometry approach to derive a comprehensive TBK1/IKK-i molecular network. The most salient feature of the network is the mutual exclusive interaction of the adaptors with the kinases, suggesting distinct alternative complexes. Immunofluorescence data indicated that the individual adaptors reside in different subcellular locations. TANK, Sintbad and NAP1 competed for binding of TBK1. The binding site for all three adaptors was mapped to the C-terminal coiled-coil 2 region of TBK1. Point mutants that affect binding of individual adaptors were used to reconstitute TBK1/IKK-i-deficient cells and dissect the functional relevance of the individual kinase-adaptor edges within the network. Using a microarray-derived gene expression signature of TBK1 in response virus infection or poly(I∶C) stimulation, we found that TBK1 activation was strictly dependent on the integrity of the TBK1/TANK interaction.
authors.affiliation: CeMM-Research Center for Molecular Medicine, Austrian Academy of Sciences, Vienna, Austria.
authors.forename: Adriana
authors.initials: A
authors.lastname: Goncalves
authors.forename: Tilmann
authors.initials: T
authors.lastname: Bürckstümmer
authors.forename: Evelyn
authors.initials: E
authors.lastname: Dixit
authors.forename: Ruth
authors.initials: R
authors.lastname: Scheicher
authors.forename: Maria W
authors.initials: MW
authors.lastname: Górna
authors.forename: Evren
authors.initials: E
authors.lastname: Karayel
authors.forename: Cristina
authors.initials: C
authors.lastname: Sugar
authors.forename: Alexey
authors.initials: A
authors.lastname: Stukalov
authors.forename: Tiina
authors.initials: T
authors.lastname: Berg
authors.forename: Robert
authors.initials: R
authors.lastname: Kralovics
authors.forename: Melanie
authors.initials: M
authors.lastname: Planyavsky
authors.forename: Keiryn L
authors.initials: KL
authors.lastname: Bennett
authors.forename: Jacques
authors.initials: J
authors.lastname: Colinge
authors.forename: Giulio
authors.initials: G
authors.lastname: Superti-Furga
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D006367:HeLa Cells; D006801:Humans; D051550:I-kappa B Kinase / Q000378:metabolism; D042541:Intracellular Space / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D025941:Protein Interaction Mapping; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000378:metabolism*; D017433:Protein Structure, Secondary; D021381:Protein Transport; D011506:Proteins / Q000378:metabolism; D012359:tRNA Methyltransferases
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D011506:Proteins; C484406:TANK protein, human; C524349:TBKBP1 protein, human; C498548:TRMO protein, human; D012359:tRNA Methyltransferases; D017346:Protein Serine-Threonine Kinases; C403191:TBK1 protein, human; D051550:I-kappa B Kinase; C496565:IKBKE protein, human