Feeding of mice with Arabidopsis thaliana expressing the HIV-1 subtype C p24 antigen gives rise to systemic immune responses

Development of transgenic edible plants, to be used as production, storage and delivery systems for recombinant vaccine antigens, is a promising strategy to obtain cost effective vaccines against infectious diseases, not least for use in developing countries. Therefore, we used Agrobacterium tumefaciens-mediated gene transfer to introduce the p24 gag gene encoding the nucleocapsid protein from HIV-1 subtype C into the Arabidopsis thaliana plant genome. Eighteen plant lines were confirmed positive for the p24 gene by PCR; four of these lines showed an apparent homozygous phenotype when grown on selective medium and these lines also showed transcription of the p24 gene into its corresponding mRNA. The mRNA in all four cases generated the p24 protein in plants, as verified by Western blot analysis. The plants were shown to contain between 0.2 mu g and 0.5 mu g p24 protein per g of fresh tissue. Analysis of the localisation of the p24 protein showed that stem tissue contained the largest amount of protein, more than twice as much as leaf tissue, whereas no p24 protein was detected in roots. By using Southern blotting, we found that 4, 2-3, 2 and 1 T-DNA insertion events took place in the four lines 1, 2, 7, and 10, respectively. The genetic insertions of line 1 were stable from the T-2 to the T-5 generation and gave rise to the p24 protein in all cases, as verified by Western blotting. In mice fed with fresh transgenic A. thaliana (line 10), anti-gag IgG was obtained in serum after a booster injection with recombinant p37Gag. No immune response was observed after equal booster injection of untreated mice or mice fed with A. thaliana WT plants.