Truncation of C-terminal 20 amino acids in PA-X contributes to adaptation of swine influenza virus in pigs.

Guanlong Xu,Qiming He,Jinhua Liu,Yipeng Sun,Xin Guo,Hanchun Yang,Ming Jiang,Qinfang Liu,Juan Pu,Honglei Sun,Yu Wang,Xuxiao Zhang,Xin Xiong

doi:10.1038/srep21845

Abstract

The PA-X protein is a fusion protein incorporating the N-terminal 191 amino acids of the PA protein with a short C-terminal sequence encoded by an overlapping ORF (X-ORF) in segment 3 that is accessed by + 1 ribosomal frameshifting, and this X-ORF exists in either full length or a truncated form (either 61-or 41-condons). Genetic evolution analysis indicates that all swine influenza viruses (SIVs) possessed full-length PA-X prior to 1985, but since then SIVs with truncated PA-X have gradually increased and become dominant, implying that truncation of this protein may contribute to the adaptation of influenza virus in pigs. To verify this hypothesis, we constructed PA-X extended viruses in the background of a “triple-reassortment” H1N2 SIV with truncated PA-X, and evaluated their biological characteristics in vitro and in vivo. Compared with full-length PA-X, SIV with truncated PA-X had increased viral replication in porcine cells and swine respiratory tissues, along with enhanced pathogenicity, replication and transmissibility in pigs. Furthermore, we found that truncation of PA-X improved the inhibition of IFN-I mRNA expression. Hereby, our results imply that truncation of PA-X may contribute to the adaptation of SIV in pigs.

Highlights

The natural reservoir of influenza A viruses is aquatic birds; some influenza strains have been spread and adapted stably in terrestrial birds or mammals[1,2]
To explore the relationship between truncation of PA-X and viral adaptation in swine, we analyzed the frequency of the PA-X truncations in swine H1N1 influenza viruses (SIVs) by year based on all SIV sequences available in the National Center for Biotechnology Information (NCBI) Influenza Viruses Resource, regardless of subtype and lineage
For the viral replication in the tissues of inoculated pigs, the replication titers of Sw-41X(WT) in nasal turbinates, tracheas and bronchoalveolar lavage fluids (BALFs) were significantly higher than Sw-61X at 3 dpi and/or 5 dpi (P < 0.05) (Fig. 6A–C). These results suggested that PA-X with 41 amino acids in X-ORF lead to increased viral replication in vivo

Summary

Introduction

The natural reservoir of influenza A viruses is aquatic birds; some influenza strains have been spread and adapted stably in terrestrial birds or mammals[1,2]. The avian-origin gene segment 3 of “triple-reassortment” (TR) SIV expresses a truncated PA-X; in contrast, PA-X proteins from avian influenza viruses are full-length[5]. These facts suggest that SIV with truncated PA-X may potentially present a selective advantage in pigs. To determine the effects of the truncation of PA-X for adaptation of SIV in pigs, we used reverse genetics to construct two TR swine influenza viruses, only differing in PA-X These included truncated PA-X and full-length PA-X, and we compared their biological characteristics in vitro and in vivo

Methods

Results

Conclusion