Molecular characterization and expression analysis of rockfish (Sebastes schlegelii) viperin, and its ability to enervate RNA virus transcription and replication in vitro.

Abstract

Viperin, also known as RSAD2 (Radical S-adenosyl methionine domain containing 2), is an interferon-induced endoplasmic reticulum-associated antiviral protein. Previous studies have shown that viperin levels are elevated in the presence of viral RNA, but it has rarely been characterized in marine organisms. This study was designed to functionally characterize rockfish viperin (SsVip), to examine the effects of different immune stimulants on its expression, and to determine its subcellular localization. SsVip is a 349 amino acid protein with a predicted molecular mass of 40.24\u202fkDa. It contains an S-adenosyl l-methionine binding conserved domain with a CNYKCGFC sequence. Unchallenged tissue expression analysis using quantitative real time PCR (qPCR) revealed SsVip expression to be the highest in the blood, followed by the spleen. When challenged with poly I:C, SsVip was upregulated by approximately 60-fold in the blood after 24\u202fh, and approximately 50-fold in the spleen after 12\u202fh. Notable upregulation was detected throughout the poly I:C challenge experiment in both tissues. Significant expression of SsVip was detected in the blood following Streptococcus iniae and lipopolysaccharide challenge, and viral hemorrhagic septicemia virus (VHSV) gene transcription was significantly downregulated during SsVip overexpression. Furthermore, cell viability assay and virus titer quantification with the presence of SsVip revealed a significant reduction in virus replication. As with previously identified viperin counterparts, SsVip was localized in the endoplasmic reticulum. Our findings show that SsVip is an antiviral protein crucial to innate immune defense.