Repression of SUMOylation pathway by grass carp reovirus contributes to the upregulation of PKR in an IFN-independent manner

Fei Yu,Liqun Lu,Jialu Sheng,Longlong Wang,Hao Wang

doi:10.18632/oncotarget.20309

Abstract

SUMOylation, a post-translational modification, is involved in interaction between hosts and viruses, and participates in diverse cellular processes including inflammatory responses and innate immunity. Here, we investigated the interaction between reovirus infection and the cellular SUMOylation machinery using grass carp reovirus (GCRV) as a model. Full-length cDNAs of grass carp SUMO-1 and SUMO-2 were obtained and phylogenetic analysis indicated that they shared high homology with those of higher vertebrates. The two modifiers and SUMO conjugating enzyme 9 (Ubc9) were ubiquitously expressed in all tested tissues of grass carp. During GCRV infection in CIK cells, transcriptional expressions of SUMO1/2 and Ubc9 were significantly inhibited; while UV-inactivated GCRV failed to inhibit the expression of the three molecules, which suggested that SUMOylation system was suppressed during viral replication. In CIK cells treated with inhibitor 2-D08 for SUMOylation, GCRV replication was not interfered; however, transcriptional analysis of immune genes involved in anti-viral interferon (IFN) response indicated that IRF2 and PKR were significantly up-regulated in CIK cells treated with inhibitor in contrast to IRF1, IRF7 and IFNI. Furthermore, 2-D08 treatment coupled with GCRV challenge resulted in higher IRF2 and PKR level during infection in comparison to those of CIK cells infected with GCRV only. These results indicated that inhibition of SUMOylation should result in the induction of PKR via IFN-independent manner, and both IFN-signaling and IFN-independent signaling seemed to involve in the upregulation of PKR during the process of GCRV infection. Repression of SUMOylation by GCRV might represent a cellular antiviral mechanism.

Highlights

SUMOylation plays an important role on the maintenance of life activity and diversity of protein function, and is involved in cancer, neurological disease, heart disease and intrinsic innate immunity [1, 2]
This study aims to monitor the SUMOylation machinery during grass carp reovirus (GCRV) infection by quantitatively detecting the transcription levels of SUMO1, SUMO2 and SUMO Conjugating Enzyme 9 (Ubc9), to investigate whether SUMOylation was necessary for GCRV replication in Ctenopharyngodon idellus kidney (CIK) cells through a specific SUMOylation-inhibitor assay, and to probe the effect of regulated SUMOylation level on host antiviral response with a focus on immune genes involved in IFN response
Four Small ubiquitin-like modifier (SUMO) paralogs in human have been identified, among which SUMO2, 3 and 4 share high homology with each other, and SUMO1, 2, and 3 can act as protein modifiers, whereas SUMO4 seems to be expressed only in limited tissues and may have no ability to be conjugated to substrate proteins [20, 21]

Summary

Introduction

SUMOylation plays an important role on the maintenance of life activity and diversity of protein function, and is involved in cancer, neurological disease, heart disease and intrinsic innate immunity [1, 2]. Similar to the ubiquitin pathway, SUMO isoforms (SUMO-1, SUMO-2/3, SUMO4) in eukaryotes are conserved small proteins, which are covalently conjugated to substrate proteins by a different set of enzymes: E1 activating enzyme, E2 conjugating enzyme and E3 ligase [3]. Grass carp Ubc bound to the N-terminal coiledcoil domain of GCRV-104 fiber protein and promoted viral infection efficiency [8, 9]. It remains to be clarified whether direct SUMOylation of viral target or SUMOylation-mediated innate immune response is responsible for the pro-viral effect of Ubc. Whether SUMOylation influences innate immunity pathway of bony fish is still not clear

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