Antiviral activity of ISG15 against classical swine fever virus replication in porcine alveolar macrophages via inhibition of autophagy by ISGylating BECN1

Cheng Li,Wang Dong,Kangkang Guo,Jihui Lin,Huifang Lv,Yanming Zhang,Hongqing Zheng,Yifan Wang

doi:10.1186/s13567-020-00753-5

Abstract

Interferons (IFNs) induce the expression of interferon-stimulated genes (ISGs) for defense against numerous viral infections, including classical swine fever virus (CSFV). However, the mechanisms underlying the effect of ISGs on CSFV infection are rarely reported. In this study, we demonstrate that IFN-α treatment induces upregulation of ISG15 and thus attenuates CSFV replication. To determine whether ISG15 is critical for controlling CSFV replication, we established porcine alveolar macrophages (PAMs) with stable overexpression or knockdown of ISG15. Overexpression of Flag-ISG15 significantly prevented CSFV replication, whereas loss of ISG15 led to abnormal proliferation of CSFV. Furthermore, upregulated ISG15 promoted beclin-1 (BECN1) ISGylation and dysfunction and subsequently inhibited autophagy, which is indispensable for CSFV replication. In addition, HECT and RLD domain containing E3 ubiquitin protein ligase 5 (HERC5), which functions to catalyze conjugation of ISG15 protein, was confirmed to interact with BECN1. Collectively, these results indicate that IFN-α restricts CSFV replication through ISG15-mediated BECN1 ISGylation and autophagy inhibition, providing insight into the mechanism of CSFV replication control by type I IFN. This mechanism may not be the only antiviral mechanism of ISG15; nonetheless, this study may contribute to the development of CSFV treatment and prevention strategies.

Highlights

Classical swine fever (CSF), which is caused by the CSF virus (CSFV) of family Flaviviridae, is a highly contagious viral disease of pig and has serious socioeconomic implications [1]
ISG15 has been proposed as an efficient host effector that defends against viral infections including that of human cytomegalovirus [17], human immune deficiency virus (HIV) [23], West Nile virus [24], and porcine reproductive and respiratory syncytial virus [25]
We provided the first strong evidence that ISG15 is an antiviral factor against CSFV in immune cells

Summary

Introduction

Classical swine fever (CSF), which is caused by the CSF virus (CSFV) of family Flaviviridae, is a highly contagious viral disease of pig and has serious socioeconomic implications [1]. The production of type I IFN relies on the recognition of pathogen-associated molecular patterns by pattern recognition receptors, which induce an antiviral response by regulating synthesis of hundreds of proteins, inducing interferon-stimulated genes (ISGs) [3, 4]. 2′,5′-oligoadenylate synthetase-like protein (OASL), myxovirus resistance protein 1 (Mx1), viperin, guanylate-binding protein 1 (GBP1), and IFN-inducible transmembrane proteins (IFITMs) have been identified to inhibit CSFV infection or replication [6,7,8,9,10,11]. It has been shown that CSFV infection triggers a complete autophagic response along with upregulation of autophagic markers, and induces formation of the membranes of autophagosome-like vesicles to promote viral replication in porcine kidney cells (PK-15) and porcine alveolar macrophages (PAMs) [14]. BECN1 regulates autophagic phosphatidylinositol 3-phosphate generation and recruits additional ATG proteins to orchestrate autophagosome formation [15]

Methods

Results

Conclusion