In vitro and in vivo bioactivity of single-chain interleukin-12

Interleukin-12 is a heterodimeric cytokine with potent immunoregulatory pro perties, making it a potential vaccine adjuvant and an immune response modu lator. The study of its function is confounded by its heterodimeric structu re. 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 porc ine interleukin-12 was bioactive in vitro on both human and porcine cells a s measured by its ability to induce proliferation of lymphoblasts and inter feron-gamma secretion by lymph node cells. In contrast, the p40 subunit of porcine interleukin-12 alone did not induce proliferation or inhibit the ac tivity of the single-chain porcine interkeukin-12. The in vivo bioactivity of single-chain porcine interleukin-12 was demonstrated in an oral immuniza tion 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 fa cilitate its in vivo delivery and study of its structure and function.