MicroRNA-like viral small RNA from porcine reproductive and respiratory syndrome virus negatively regulates viral replication by targeting the viral nonstructural protein 2.

Na Li,Jiming Gao,Gaiping Zhang,Angke Zhang,Chong Zhang,Jinbu Jia,Yunhuan Yan,En-Min Zhou,Gaopeng Hou,Shuqi Xiao,Xue Wang

doi:10.18632/oncotarget.12703

Abstract

Many viruses encode microRNAs (miRNAs) that are small non-coding single-stranded RNAs which play critical roles in virus-host interactions. Porcine reproductive and respiratory syndrome virus (PRRSV) is one of the most economically impactful viruses in the swine industry. The present study sought to determine whether PRRSV encodes miRNAs that could regulate PRRSV replication. Four viral small RNAs (vsRNAs) were mapped to the stem-loop structures in the ORF1a, ORF1b and GP2a regions of the PRRSV genome by bioinformatics prediction and experimental verification. Of these, the structures with the lowest minimum free energy (MFE) values predicted for PRRSV-vsRNA1 corresponded to typical stem-loop, hairpin structures. Inhibition of PRRSV-vsRNA1 function led to significant increases in viral replication. Transfection with PRRSV-vsRNA1 mimics significantly inhibited PRRSV replication in primary porcine alveolar macrophages (PAMs). The time-dependent increase in the abundance of PRRSV-vsRNA1 mirrored the gradual upregulation of PRRSV RNA expression. Knockdown of proteins associated with cellular miRNA biogenesis demonstrated that Drosha and Argonaute (Ago2) are involved in PRRSV-vsRNA1 biogenesis. Moreover, PRRSV-vsRNA1 bound specifically to the nonstructural protein 2 (NSP2)-coding sequence of PRRSV genome RNA. Collectively, the results reveal that PRRSV encodes a functional PRRSV-vsRNA1 which auto-regulates PRRSV replication by directly targeting and suppressing viral NSP2 gene expression. These findings not only provide new insights into the mechanism of the pathogenesis of PRRSV, but also explore a potential avenue for controlling PRRSV infection using viral small RNAs.

Highlights

Porcine reproductive and respiratory syndrome (PRRS) is one of the most prevalent viral diseases in swine
Among these viral small RNAs (vsRNAs), Porcine reproductive and respiratory syndrome virus (PRRSV)-vsRNA1 exhibited stem-loop structures with the lowest minimum free energy (MFE) value, -35.4 kcal/mol, which mirrored the stability of a typical hairpin structure (Figure 1B and Table 1)
We identified and characterized miRNA-like small RNAs encoded by PRRSV using bioinformatic prediction and experimental verification tools

Summary

Introduction

Porcine reproductive and respiratory syndrome (PRRS) is one of the most prevalent viral diseases in swine. Infection progressively leads to reproductive failure in pregnant sows and respiratory distress in young pigs, causing significant economic losses to the swine industry worldwide each year [1,2,3]. The causative pathogen of PRRS is the PRRS virus (PRRSV), a small enveloped linear, positive-sense single-stranded RNA virus, which is a member of the order Nidovirales in the family Arteriviridae [4]. In China, HP-PRRS coexists with the long-established, low pathogenic North American type PRRSV strains. Because these coexistent strains cannot be controlled with the same vaccine, current vaccination strategies cannot effectively control PRRSV infection. It is imperative to better understand the mechanisms of PRRSV pathogenesis in order to facilitate the development of more effective control measures

Methods

Results

Conclusion