Poster Presentation: IL-12p35 and IL-12p40 Functionally Synergize to Activate NK Cells

Interleukin-12 (IL-12) is a member of a family of heterodimeric cytokines that play critical roles in regulating the immune response toward different effector pathways. The formation of functional IL-12 requires association of the IL-12p35 subunit with the IL-12p40 subunit, and the secretion of IL-12 relies on p40 and p35 being expressed and assembled within dendritic cells and macrophages. IL-12p40 can also be released from these hematopoietic cells as a monomer and is usually found in great excess over IL-12 in serum and cell culture supernatants. The precise biological significance of the secreted IL-12p40 monomer is not clear, although recent studies indicate that it has immunomodulatory properties. Previous studies have reported that p40 and p35 can assemble to form functionally active IL-12 in vitro. IL-12p40 associates with p35 extracellularly to generate IL-12–like activities on T cells, but the biological significance on NK cells remains unclear.

In this study, we show that only high concentrations of IL-12p35 could activate NK cells to secrete IFN-gamma and induce cell proliferation and IL-12p40 had no effect on NK cell activation. Interestingly, IL-12p35 and p40 functionally synergized to activate NK cells by inducing IFN-gamma secretion and cell proliferation in vitro. Surprisingly, pretreatment of NK cells with IL-12p35 or p40 didn’t affect IL-12 induced IFN-gamma secretion and cell proliferation in NK cells. Furthermore, a functional ELISA binding assay revealed that IL-12p35 binds to both IL-12R beta1 and IL-12R beta2 with a low affinity, while IL-12p40 only binds to IL-12R beta1 with a high affinity. Fluorescent conjugated human IL-12, IL-12p35 and IL-12p40 proteins all significantly bound to human NK cells and IL-12p35 and p40 didn’t significantly interfere with IL-12 binding to these cells. Collectively, the data we show herein indicates that IL-12 activity can come from the collaboration between the IL-12p35 and p40 subunits and impact NK cell activation.