A Novel Transcript Isoform of TBK1 Negatively Regulates Type I IFN Production by Promoting Proteasomal Degradation of TBK1 and Lysosomal Degradation of IRF3.

Jie Zhang,Ming Xian Chang,Yi Wei Hu,Xiao Man Wu

doi:10.3389/fimmu.2020.580864

Abstract

TANK-binding kinase 1 (TBK1), an IKK-related serine/threonine kinase, is pivotal for the induction of antiviral type I interferon (IFN) by TLR and RLR signaling pathways. In a previous study, we demonstrated that TBK1 spliced isoforms (TBK1_tv1 and TBK1_tv2) from zebrafish were dominant negative regulators in the RLR antiviral pathway by targeting the functional TBK1–IRF3 complex formation. In this study, we show that the third TBK1 isoform (namely TBK1_tv3) inhibits zebrafish type I IFN production by promoting TBK1 and IRF3 degradation. First, ectopic expression of TBK1_tv3 suppresses poly(I:C)- and Spring viremia of carp virus-induced type I IFN response, and also inhibits the up-regulation of IFN promoter activities stimulated by RIG-I, MDA5, MAVS, TBK1, and IRF3. Second, TBK1_tv3 targets TBK1 and IRF3 to impair the formation of TBK1 dimer, TBK1–IRF3 complex, and IRF3 dimer. Notably, TBK1_tv3 promotes the degradation of TBK1 through the ubiquitin–proteasome pathway and the degradation of IRF3 through the lysosomal pathway. Further analysis demonstrates that TBK1_tv3 promotes the degradation of TBK1 for K48-linked ubiquitination by targeting the K251, K256, and K271 sites of TBK1. Collectively, our results suggest a novel TBK1 isoform-mediated negative regulation mechanism, which serves to balance the production of type I IFN and ISGs.

Highlights

Type I interferon (IFN) plays a vital role in inducing cell-intrinsic antimicrobial states in infected and neighboring cells to limit the spread of viral pathogens, modulating innate immune response in a balanced manner and activating the adaptive immune system to promote the development of T- and B-cell responses [1]
By sequence aligning with TANK-binding kinase 1 (TBK1) normal form, TBK1_tv3 lacks exons 14–17, 9 bp at the C terminal of exon 13, 34 bp at the N terminal of exon 18, and 17 bp in the middle of exon 19 (Figure 1A), which leads to an in-frame deletion of the C-terminal CCD2 domain of TBK1
Our previous study showed that zebrafish TBK1_tv1 and TBK1_tv2 isoforms inhibited RIG-I, MAVS, TBK1, and IRF3-mediated activation of IFN promoters in response to Spring viremia of carp virus (SVCV) infection, disrupted the formation of a functional TBK1–IRF3 complex, and impeded the phosphorylation of IRF3 mediated by TBK1 [18]

Summary

Introduction

Type I interferon (IFN) plays a vital role in inducing cell-intrinsic antimicrobial states in infected and neighboring cells to limit the spread of viral pathogens, modulating innate immune response in a balanced manner and activating the adaptive immune system to promote the development of T- and B-cell responses [1]. Activated TBK1 phosphorylates IRF3 and induces the production of type I IFN [6, 7]. The E3 ubiquitin ligase RNF128 activates TBK1 by promoting the K63-linked polyubiquitination of TBK1 at Lys and Lys401 [11]. Many molecules such as TRIM26 and IFIT3 associate with TBK1 to facilitate the formation of TBK1-containing complexes; other molecules such as NLRC3 and NLRX1 prevent the interaction of TBK1 with other adaptors [12,13,14,15]

Methods

Results

Conclusion