Structure and Ubiquitination-Dependent Activation of TANK-Binding Kinase 1

Abstract

Summary Upon stimulation by pathogen-associated inflammatory signals, the atypical IκB kinase TBK1 induces type-I interferon expression and modulates NF-κB signaling. Here we describe the 2.4 Å-resolution crystal structure of nearly full-length TBK1 in complex with specific inhibitors. The structure reveals a novel dimeric assembly, created by an extensive network of interactions among the kinase, ubiquitin-like (ULD) and scaffold/dimerization (SDD) domains. An intact TBK1 dimer undergoes K63-linked polyubiquitination on Lysine 30 and Lysine 401, and these modifications are required for TBK1 activity. The ubiquitination sites and dimer contacts are conserved in the close homolog IKKε, but not in the canonical IκB kinase IKKβ, which assembles in an unrelated manner. The multidomain architecture of TBK1 provides a structural platform for integrating ubiquitination with kinase activation and IRF3 phosphorylation. The structure of TBK1 will facilitate studies of the atypical IκB kinases in normal and disease physiology and will further development of more specific inhibitors that may be useful as anti-cancer or anti-inflammatory agents.

Murine Gammaherpesvirus 68 Encoding Open Reading Frame 11 Targets TANK Binding Kinase 1 To Negatively Regulate the Host Type I Interferon Response

“…and lacks two coiled-coil domains at the C terminus, which serve as a scaffold/dimerization domain (SDD) (52)(53)(54). Consistent with the previous report (52), these two mutants failed to activate IFN-␤ production in reporter assays (Fig.…”

Hantavirus GnT Elements Mediate TRAF3 Binding and Inhibit RIG-I/TBK1-Directed Beta Interferon Transcription by Blocking IRF3 Phosphorylation

“…Replicating RNA viruses generate small amounts of cytoplasmic double-stranded RNA (dsRNA), or 5=-triphosphate-containing RNAs, that are detected by MDA5 or RIG-I sensors and result in the mitochondrial assembly of MAVS complexes (28)(29)(30)(31)(32)(33). MAVS binds tumor necrosis factor receptor-associated factor 3 (TRAF3), which recruits TBK1/IKKε kinases that activate NF-B and direct the phosphorylation of constitutively expressed IRF3 (27,(34)(35)(36)(37)(38)(39)(40). Activated IRFs and NF-B translocate to the nucleus and form transcriptional complexes with CBP/p300 and ATF2/cJUN on the IFN-␤ enhanceosome that transcriptionally induce IFN-␤ (28,29,32,41,42).…”

“…TRAF3 recruits TBK1 to signaling complexes that are essential for the induction of type I IFN (38). Interactions that mediate TRAF3 binding to TBK1 are regulated by ubiquitination and impacted by a growing list of factors (34,36,40,45). However, residues 440 to 442 within the TRAF-C domain of TRAF3 are Luciferase activity within lysates was determined 48 h posttransfection, normalized to Renilla luciferase activity, and reported as the fold increase compared to that of controls lacking inducer.…”

“…Overall, these findings indicate that degron binding to TRAF3 is not required for IFN regulation by hantavirus GnT proteins. However, we cannot exclude the possibility that TRAF3 binding facilitates GnT regulation of TBK1, IRF3, or ubiquitin ligases or alternatively TRAF3 heterotrimer formation with additional TRAF2/5 adapter proteins (36,40,58,62). GnT effects on TBK1-directed ISRE responses were evaluated as described for Fig.…”

Loss of TBK1 Induces Epithelial–Mesenchymal Transition in the Breast Cancer Cells by ERα Downregulation

“…Knockdown of IKKe sensitizes breast cancer cells to tamoxifeninduced growth arrest, whereas the ectopic expression of IKKe exerts the opposite effect through interaction with ERa in breast cancer cells (24). In particular, TBK1 has been validated as a potential therapeutic target for increasing synthetic lethality in K-Ras-driven cancers through its relationship with mutant K-Ras (26,27,40,41). Although IKKe has been extensively examined in breast cancer cells, the relationship between TBK1 and ERa expression status in different breast cancer subtypes remains to be elucidated.…”