In Vitro and In Vivo Bioactivity of Single‐Chain Interleukin‐12

Interleukin‐12 is a heterodimeric cytokine with potent immunoregulatory properties, making it a potential vaccine adjuvant and an immune response modulator. The study of its function is confounded by its heterodimeric structure. In order to facilitate the study of interleukin‐12 in both in vitro and in vivo models, we constructed a single‐chain porcine interleukin‐12 gene and expressed the recombinant protein in Pichia pastoris. Single‐chain porcine interleukin‐12 was bioactive in vitro on both human and porcine cells as measured by its ability to induce proliferation of lymphoblasts and interferon‐γ secretion by lymph node cells. In contrast, the p40 subunit of porcine interleukin‐12 alone did not induce proliferation or inhibit the activity of the single‐chain porcine interkeukin‐12. The in vivo bioactivity of single‐chain porcine interleukin‐12 was demonstrated in an oral immunization model where it increased antigen‐specific IgA and IgG in jejunal mucus. These results indicate that binding of interleukin‐12 to its receptor and transduction of intracellular signals requires both p40 and p35 subunits. The bioactivity of interleukin‐12 expressed as a single polypeptide will facilitate its in vivo delivery and study of its structure and function.