Recombinant adenovirus expression of FMDV P1-2A and 3C protein and its immune response in mice

The purpose of this research was to develop a new type of recombinant adenovirus vaccine against foot-and-mouth disease (FMD) virus and confirm whether both 2A and 3C proteases can fully process FMDV P1 polyprotein into VP1 protein. We constructed and characterized recombinant adenoviruses, designated as rAd-P1, rAd-P1-2A, rAd-L-P1, rAd-L-2A, and rAd-3C. The construct was further confirmed by the enzyme digestion, polymerase chain reaction (PCR) testing, sequencing, and transfection. The infective replication-defective adenovirus rAd-3C was used to co-infect Vero cells with rAd-P1, rAd-P1-2A, rAd-L-P1, and rAd-L-2A, respectively; the supernatant of cracked cells was then collected to immunize mice. Notably, the supernatant of the Vero cells co-infected with rAd-L-2A/rAd-3C or rAd-P1-2A/rAd-3C was shown to induce specific humoral and cellular immune responses in the study animals. Most importantly, we confirmed that 3C protease successfully processed P1 polyprotein into VP1 protein without L protein only under the precondition of 2A protease; however, P1-2A and L-2A polyprotein were not fully processed. Our study highlighted that P1 polyprotein cannot be fully processed into structural protein VP1 only under the condition of 2A and 3C proteases. The results also suggest that the recombinant adenovirus may be an attractive candidate as a vaccine for preventing the disease associated with FMD virus infection.