TAX1BP1 and A20 Inhibit Antiviral Signaling by Targeting TBK1-IKKi Kinases

Abstract

Induction of type I interferons by the transcription factor IRF3 is essential in the initiation of antiviral innate immunity. Activation of IRF3 requires C-terminal phosphorylation by the upstream kinases TBK1-IKKi, where IRF3 phosphorylation promotes dimerization, and subsequent nuclear translocation to the IFN␤ promoter. Recent studies have described the ubiquitin-editing enzyme A20 as a negative regulator of IRF3 signaling by associating with TBK1-IKKi; however, the regulatory mechanism of A20 inhibition remains unclear. Here we describe the adaptor protein, TAX1BP1, as a key regulator of A20 function in terminating signaling to IRF3. Murine embryonic fibroblasts (MEFs) deficient in TAX1BP1 displayed increased amounts of IFN␤ production upon viral challenge compared with WT MEFs. TAX1BP1 inhibited virus-mediated activation of IRF3 at the level of TBK1-IKKi. TAX1BP1 and A20 blocked antiviral signaling by disrupting Lys 63 -linked polyubiquitination of TBK1-IKKi independently of the A20 deubiquitination domain. Furthermore, TAX1BP1 was required for A20 effector function because A20 was defective for the targeting and inactivation of TBK1 and IKKi in Tax1bp1 ؊/؊ MEFs. Additionally, we found the E3 ubiquitin ligase TRAF3 to play a critical role in promoting TBK1-IKKi ubiquitination. Collectively, our results demonstrate TBK1-IKKi to be novel substrates for A20 and further identify a novel mechanism whereby A20 and TAX1BP1 restrict antiviral signaling by disrupting a TRAF3-TBK1-IKKi signaling complex.

Aryl Hydrocarbon Receptor Interacting Protein Targets IRF7 to Suppress Antiviral Signaling and the Induction of Type I Interferon

“…IFN-␣4 luciferase reporter was provided by Dr. Shunbin Ning (East Tennessee State University) (43). IFN-␤ and NF-B reporters were described previously (22,44). Flag-IRF3, GFP-⌬RIG-I, GFP-TBK1, and Flag-A20 were previously described (22,23,45).…”

Ubiquitin in the activation and attenuation of innate antiviral immunity

“…Furthermore, there is increasing evidence of ubiquitin-mediated cross-talk between TRAFs. TRAF3 promotes IFN-I expression by activating TBK1/IRF3 (Parvatiyar et al, 2010), whereas TRAF6 activates mitogen-activated protein kinase kinase kinase 1 (MEKK1) to activate NF-κB, which also enhances IFN-I expression (Yoshida et al, 2008). Simultaneously, TRAF3 suppresses NF-κB by inhibiting IKK activation upon binding TRAF2 (Zarnegar et al, 2008), likely as a mechanism to skew innate immune effector molecule expression as required.…”

“…TBK1 polyubiquitination by Nrdp1 activates TBK1 in TRIF-mediated IFN-I expression, which simultaneously K48-polyubiquitinates and down-regulates MyD88-mediated NF-κB activation . Conversely, the ubiquitin-editing enzyme A20 inhibits ubiquitin-mediated activation of TRAF6, inhibiting NF-κB activation via the TLR and NLR cascades, as well as TBK1, inhibiting IFN-I expression Turer et al, 2008;Parvatiyar et al, 2010). Additional avenues of signaling cross-talk include Smurf1 and Smurf2, which degrade MAVS and inhibit IFN-I activation.…”

“…Conversely, NOD2-driven NF-κB activation is enhanced by LUB AC, a negative regulator of RLR signaling, as well as X-linked IAP (XIAP), which respectively ligate M1-and K63-linked polyubiquitin chains to NOD2 and RIPK2 (Damgaard et al, 2012). These activating ubiquitin chains may be antagonized by the ubiquitin-editing enzyme A20 , which also disrupts ubiquitin-mediated TBK1 activation in the RLR signaling cascade as well as ubiquitin-mediated TRAF6 activation in the TLR cascade Parvatiyar et al, 2010).…”

“…This facilitates interactions between TBK1, inhibitor of NF-κB kinase subunit ε (IKKε) and TRAFs, particularly TRAF3. Upon RLR activation, these proteins are colocalized at the cytosolic surface of the mitochondrial outer membrane (Parvatiyar et al, 2010;van Zuylen et al, 2012), coordinated by the mitochondrial antiviral-signaling protein (MAVS; also termed VISA/Cardif/IPS-1).…”