Transcriptome profiling analysis of lung tissue of Chinese soft-shell turtle infected by Trionyx sinensis Hemorrhagic Syndrome Virus.

Trionyx sinensis Hemorrhagic Syndrome Virus (TSHSV) is the firstly discovered aquatic arterivirus inducing high mortality of Trionyx sinensis. So far, the lack of genomic resources has hindered further research on revealing the immunological characteristics of T. sinensis in response to TSHSV. In the present study, we performed a transcriptome analysis from the lungs of T. sinensis challenged by TSHSV using Illumina-based RNA-Seq. The validity of transcriptomic data was confirmed with the gradual increase of TSHSV RNA copies detected in lung. A total of 103079339 clean reads were generated, and 58374764 unique mapped reads were analyzed. Assembly of the sequence data allowed identifying 16383 unigenes consisting of 36 significant differentially expressed genes (DEGs). These DEGs were categorized into 30 GO-enriched bioprocesses and 9 KEGG pathways. The combinational analysis of GO-enriched bioprocesses and KEGG pathways demonstrated that TSHSV modulated several immune genes of T. sinensis related to various biological processes, including virus recognition (RIG-I/MDA-5), immune initiation (IFIT-1 and IFIT-5), endocytosis (CUBN, ENPP2 and LRP2) and steroid metabolism (FCNIL and STAR). In summary, the finding of this study revealed several immune pathways and candidated genes involved in the immune response of T. sinensis against TSHSV-infection. These results will provide helpful information to investigate molecular mechanism of T. sinensis in response to TSHSV.

Site-directed mutagenesis of the Nidovirus replicative endoribonuclease NendoU exerts pleiotropic effects on the arterivirus life cycle.

The highly conserved NendoU replicative domain of nidoviruses (arteriviruses, coronaviruses, and roniviruses) belongs to a small protein family whose cellular branch is prototyped by XendoU, a Xenopus laevis endoribonuclease involved in nucleolar RNA processing. Recently, sequence-specific in vitro endoribonuclease activity was demonstrated for the NendoU-containing nonstructural protein (nsp) 15 of several coronaviruses. To investigate the biological role of this novel enzymatic activity, we have characterized a comprehensive set of arterivirus NendoU mutants. Deleting parts of the NendoU domain from nsp11 of equine arteritis virus was lethal. Site-directed mutagenesis of conserved residues exerted pleiotropic effects. In a first-cycle analysis, replacement of two conserved Asp residues in the C-terminal part of NendoU rendered viral RNA synthesis and virus production undetectable. In contrast, mutagenesis of other conserved residues, including two putative catalytic His residues that are absolutely conserved in NendoU and cellular homologs, produced viable mutants displaying reduced plaque sizes (20 to 80% reduction) and reduced yields of infectious progeny of up to 5 log units. A more detailed analysis of these mutants revealed a moderate reduction in RNA synthesis, with subgenomic RNA synthesis consistently being more strongly affected than genome replication. Our data suggest that the arterivirus nsp11 is a multifunctional protein with a key role in viral RNA synthesis and additional functions in the viral life cycle that are as yet poorly defined.