Viperin protein inhibits the replication of caprine parainfluenza virus type 3 (CPIV 3) by interaction with viral N protein

摘要

Caprine parainfluenza virus type3 (CPIV3) is a newly identified member of Paramyxoviridae family. CPIV3 is highly prevalence in China and showed pathogenicity to goats; in addition, CPIV3 infection causes severe clinical disease under stress and/or co-infection conditions. Viperin is one of the hundreds of interferon-stimulated genes (ISGs), and possesses a wide range of antiviral activities. The aim of this study was to systemically explore the anti-CPIV3 activity of ruminants' Viperin. CPIV3 infection up-regulated Viperin transcription but not protein expression in MDBK cells. Bovine and caprine Viperin genes (bVi and gVi) were amplified and analyzed by BLAST and multiple alignment. The obtained bVi/gVi amino acid sequences showed 99.5%-100% identity with previously submitted sequences and has variants at N-terminal domain (1-70aa) between each other. The pcDNA3.1 plasmids containing bVi and gVi genes were constructed to over-express the target proteins. CPIV3 was inoculated in MDBK cells over-expressing bVi/gVi and viral load was detected by qRT-PCR, virus titration and Western blot. Both of the bVi and gVi significantly inhibited CPIV3 genome copy numbers and viral titers at 24 and 48 hpi (P < 0.01); and viral N protein expression was also decreased, comparing with those of mock transfected group. The last 50aa C-terminal region was crucial for its anti-CPIV3 activity. In addition, the overexpression of bVi/gVi did not influence CPIV3 binding, entry and release in the cells. These results indicated the anti-CPIV3 activity occurred in viral RNA/protein synthesis progress of the viral replication cycle. The Viperin also showed similar inhibitory effect on different CPIV3 strains. The potential interaction of Viperin with viral proteins (N, P, C and V) was determined by confocal laser scanning microscopy and Co-IP assay. Co-localization of Viperin with N, P or C, but not V, was observed; while only N protein direct interacted with Viperin in Co-IP test, no matter using viral protein expressing plasmids transfected or CPIV3 infected cell samples. In conclusion, the bVi and gVi Viperin effectively inhibited CPIV3 replication potentially via the interaction of Viperin with viral N protein. The present results gave more information about antiviral activity of ruminants Viperin and provided foundation for further studies of the interaction of Viperin with CPIV3 and other related viruses.