Tetraspanin 6 (TSPAN6) Negatively Regulates Retinoic Acid-inducible Gene I-like Receptor-mediated Immune Signaling in a Ubiquitination-dependent Manner

Yetao Wang,Xiaomei Tong,Ehimwenma Sheena Omoregie,Wenjun Liu,Songdong Meng,Xin Ye

doi:10.1074/jbc.m112.390401

Abstract

The recognition between retinoic acid-inducible gene I-like receptors (RLRs) and viral RNA triggers an intracellular cascade of signaling to induce the expression of type I IFNs. Both positive and negative regulation of the RLR signaling pathway are important for the host antiviral immune response. Here, we demonstrate that the tetraspanin protein TSPAN6 inhibits RLR signaling by affecting the formation of the adaptor MAVS (mitochondrial antiviral signaling)-centered signalosome. We found that overexpression of TSPAN6 impaired RLR-mediated activation of IFN-stimulated response element, NF-κB, and IFN-β promoters, whereas knockdown of TSPAN6 enhanced the RLR-mediated signaling pathway. Interestingly, as the RLR pathway was activated, TSPAN6 underwent Lys-63-linked ubiquitination, which promoted its association with MAVS. The interaction of TSPAN6 and MAVS interfered with the recruitment of RLR downstream molecules TRAF3, MITA, and IRF3 to MAVS. Further study revealed that the first transmembrane domain of TSPAN6 is critical for its ubiquitination and association with MAVS as well as its inhibitory effect on RLR signaling. We concluded that TSPAN6 functions as a negative regulator of the RLR pathway by interacting with MAVS in a ubiquitination-dependent manner.

Highlights

The RIG-I-mediated signaling pathway is important for the antiviral immune response
TSPAN6 Inhibits retinoic acid-inducible gene I-like receptors (RLRs) Signaling at the MAVS Level—The genomes of influenza A virus and Sendai virus (SeV) can be recognized by RIG-I and trigger downstream signaling [17, 18]
To further examine whether TSPAN6 is involved in negative regulation of the RLR pathway, we analyzed the effect of TSPAN6 on influenza A virus- or SeV-induced activation of the IFN-␤ promoter

Summary

Background

The RIG-I-mediated signaling pathway is important for the antiviral immune response. Results: TSPAN6 inhibits the formation of the MAVS-centered signalosome. Upon detecting viral RNAs, RLRs are recruited to the mitochondrial adaptor MAVS (mitochondrial antiviral signaling; called VISA, IPS-1, and Cardif) [2,3,4,5], other RLR signaling components such as MITA (mediator of IRF3 activation), TRAF2/3/6 (TNF receptor-associated factor 2/3/ 6), TRADD, and RIP1 are subsequently recruited to MAVS, forming a MAVS-centered signalosome, which further triggers TBK1- and I␬B kinase-mediated activation of IRF3/7 (IFN regulatory factor 3/7) and NF-␬B These signaling events lead to the induction of type I IFNs and inflammatory cytokines [6]. We found that TSPAN6 inhibits the RLR-mediated immune response It impairs RIG-I-, MDA5-, and MAVS-mediated (but not TBK1- and Toll-like receptor adaptor TRIF-mediated) activation of IFN-␤. As the RLR signaling pathway is activated, TSPAN6 undergoes Lys-63-linked ubiquitination, which is critical for its interaction with MAVS and the inhibitory effect on the recruitment of TRAF3, MITA, and IRF3 to MAVS. We propose that TSPAN6 is a negative regulator of the RLR-mediated signaling pathway

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION