RNA interference targeting glycoprotein D inhibits infectious bovine rhinotracheitis virus replication in MDBK cells

Abstract

Infectious bovine rhinotracheitis (IBR) is an acute, febrile, contagious disease caused by infectious bovine rhinotracheitis virus (IBRV). IBRV vaccine is considered to be partially effective but not yet completely successful. Therefore, establishing a new antiviral approach is critical. RNA interference (RNAi) has been rapidly developed in recent years as an antiviral therapy for several viruses. This study showed that RNAi could suppress IBRV replication via knockdown of a virion glycoprotein. Two recombinant lentiviral vectors containing short hairpin RNAs (shRNAs) (H1-RNA-121, H1-RNA-304) against the glycoprotein gD of IBRV and a pcDNA3-gD vector containing a FLAG tag were constructed. pcDNA3-gD and the individual shRNA recombinant lentiviral vectors were co-transfected into 293T cells, and the efficiency of RNAi was verified using Western blotting. H1-RNA-304 strongly suppressed the transient expression of the FLAG-tagged gD fusion protein. The recombinant H1-RNA lentivirus was packaged by transfecting the 293T cells with the recombinant H1-RNA vector and two helper plasmids using Lipofectamine, and the lentivirus was then used to infect MDBK cells. When the MDBK cells were infected with IBRV after infection with the H1-shRNAs, H1-shRNA-304 was more effective at markedly silencing viral gD gene expression and it inhibited IBRV replication. These results indicate that shRNAs targeting the gD gene have substantial antiviral properties and inhibit IBRV replication in a sequence-specific manner, demonstrating their potential for clinical application.