Ubiquitin-specific Protease 15 Negatively Regulates Virus-induced Type I Interferon Signaling via Catalytically-dependent and -independent Mechanisms.

Huan Zhang,Rui Luo,Liurong Fang,Huijuan Zhong,Dezhi Liu,Shaobo Xiao,Dang Wang,Min Shang,Huanchun Chen

doi:10.1038/srep11220

Abstract

Viral infection triggers a series of signaling cascades, which converge to activate the transcription factors nuclear factor-κB (NF-κB) and interferon regulatory factor 3 (IRF3), thereby inducing the transcription of type I interferons (IFNs). Although not fully characterized, these innate antiviral responses are fine-tuned by dynamic ubiquitination and deubiquitination processes. In this study, we report ubiquitin-specific protease (USP) 15 is involved in regulation of the retinoic acid-inducible gene I (RIG-I)-dependent type I IFN induction pathway. Knockdown of endogenous USP15 augmented cellular antiviral responses. Overexpression of USP15 inhibited the transcription of IFN-β. Further analyses identified histidine 862 as a critical residue for USP15’s catalytic activity. Interestingly, USP15 specifically removed lysine 63-linked polyubiquitin chains from RIG-I among the essential components in RIG-I-like receptor-dependent pathway. In addition, we demonstrated that in contrast to USP15 de-ubiquitinating (DUB) activity, USP15-mediated inhibition of IFN signaling was not abolished by mutations eliminating the catalytic activity, indicating that a fraction of USP15-mediated IFN antagonism was independent of the DUB activity. Catalytically inactive USP15 mutants, as did the wild-type protein, disrupted virus-induced interaction of RIG-I and IFN-β promoter stimulator 1. Taken together, our data demonstrate that USP15 acts as a negative regulator of RIG-I signaling via DUB-dependent and independent mechanisms.

Highlights

The infection of viral induces a strong antiviral immune response characterized by robust production of type I interferons (IFNs) and proinflammatory cytokines
By using reporter assays in HEK293T cells to screen a pool of 162 siRNAs targeting 54 human ubiquitin-specific protease (USP) genes for the capabilities to modulate Sendai virus (SEV)-induced activation of the ISRE promoter, we found USP15 is a potential candidate in previous study[24]
Further supporting the notion that USP15 impacts the downstream cascade that amplifies type I IFN expression are the data shown in Supplementary Fig. S1, which suggest that knockdown of USP15 potentiated the SEV-induced expression of IFN-stimulated genes (ISGs) (ISG15, ISG20, ISG54, ISG56, IP-10)

Summary

Introduction

The infection of viral induces a strong antiviral immune response characterized by robust production of type I interferons (IFNs) and proinflammatory cytokines. Emerging evidence indicates that ubiquitin plays a pivotal role in a myriad of biological processes, including protein degradation, cell-cycle regulation, kinase activation, and cell signal transduction[14]. Ubiquitination and deubiquitination are critically involved in regulating the virus-induced type I IFN pathway. To verify the specific mechanism of USP15-mediated IFN inhibition, we identified the DUB activity site His[862] of USP15 in vivo and in vitro, and we presented evidence that USP15 functions as a RIG-I deubiquitinase and removes Lys63-linked polyubiquitin chains. We identified USP15 as a multifunctional regulator of the virus-induced type I IFN signaling pathway via DUB-dependent and independent mechanisms

Methods

Results

Conclusion