Expression of Bovine Interleukin-1β in a Bovine Herpesvirus-1 Vector:In VitroAnalysis

Abstract

In order to evaluate whether bovine herpesvirus-1 (BHV-1) could be used as a live viral vector for the expression of cytokines, we constructed a recombinant BHV-1 expressing bovine interleukin-1β (boIL-1β). The boIL-1β coding sequence, corresponding to the cleaved mature product, was fused with the BHV-1 glycoprotein C (gC) signal peptide sequence; the resultant gC-boIL-1β fusion gene was recombined into the gC locus of the BHV-1 genome. Southern blot analysis confirmed the proper genomic configuration of the recombinant virus. Results from transcript analysis showed that boIL-1β was expressed in infected cells with kinetics similar to that of gC. Indirect immunofluorescence and immunoprecipitation assays showed that the recombinant protein was produced in both cell-associated and secreted forms. Western blot analysis detected a 19.3-kDa protein. Further analysis, using an IL-1β bioassay demonstrated that both the cellular and secreted forms of recombinant boIL-1β possessed biological activity. The expression of the boIL-1β protein did not affect thein vitrogrowth efficiency of the virus, which exhibited similar growth kinetics to that of a simple gC deletion mutant. The results from this study demonstrate that BHV-1 can be used to express a functional cytokine, thereby establishing the basis to further study recombinant BHV-1 expressing cytokines as an alternative means to attenuate the virus and also as a potentialin situcytokine delivery system to modulate immune responses against BHV-1 and other cattle pathogens.