Cellular Thioredoxin 2 Interacts With The E2 Protein And Regulates The Replication Of Classical Swine Fever Virus

Classical swine fever(CSF) is an OIE-listed animal disease notifiable to the World Organization for Animal Health(OIE). CSF is characterized by fever, high mortality, immunosuppression and reproduction failure. Classical swine fever virus(CSFV), the causative agent of CSF, belongs to the Pestivirus genus of the Flaviviridae family. The structural components of CSFV include C, Erns, E1 and E2 proteins. The E2 glycoprotein is one of the virulence-associated factors and the most important protective antigen of CSFV in pigs. It is also involved in virus attachment to and entry into the cell. To date, however, the E2-interacting cellular proteins and their involvement in viral replication have been poorly documented.In this study, the CSFV E2 protein was used as bait in a yeast two-hybrid to screen from a porcine macrophage cDNA library, which resulted in the identification of thioredoxin 2(Trx2) as a novel interacting partner of E2. Trx2 is a mitochondria-associated thioredoxin that participates in diverse cellular events. Trx2 is expressed in different tissues of pigs. The Trx2-E2 interaction was further confirmed by GST pull-down, in situ proximity ligation and laser confocal assays. The thioredoxin domain of Trx2 was shown to be critical to the interaction. Silencing of the Trx2 expression in PK-15 cells by small interfering RNAs significantly promoted CSFV replication, and conversely, overexpression of Trx2 markedly inhibited the replication of CSFV. CSFV, as well as E2, was shown to reduce the Trx2 protein expression in PK-15 cells.In summary, we demonstrate that the Trx2 interacts with the E2 protein and the thioredoxin domain of Trx2 is essential to the interaction, Trx2 inhibits CSFV replication, and the CSFV infection downregulates Trx2 expression. This study will help to better understand the mechanisms involved in the replication of CSFV.