(316e) Inferring Relevant Control Mechanisms for Interleukin-12 Signaling in Naive CD4+ T Cells

Interleukin-12 (IL-12) is a key cytokine involved in shaping cell-mediated immunity to intracellular pathogens. IL-12 initiates the cellular response via its receptor, IL-12R. The IL-12 signaling pathway is a member of the JAK/STAT family of signaling networks. The JAK/STAT pathway includes several regulatory elements; however, the dynamics of these mechanisms are not fully understood. Therefore, the objective of this study was to probe the relative importance of regulatory mechanisms that modulate the activation of STAT4 in primary naive CD4+ T cells. Naive CD4+ CD62L+ T cells were isolated from murine Balb/c splenocytes and activated for 44 hours to provide nonspecific activation and co-stimulation of the T cell receptor. Activated T cells were stimulated with anti-IL-4 and IL-12 and stained with markers for IL-12Rbeta1, IL-12Rbeta2, and phosphorylated STAT4. Dynamic changes in protein expression and activity were measured using flow cytometry and analyzed using R/Bioconductor. These data were used to calibrate a mathematical model of IL-12 signaling, which included the cell's natural programming of IL-12R expression and control feedback loops. An empirical Bayesian approach was used to infer the relative strength of different regulatory mechanism in the system. The model predicted that IL-12 receptor expression is regulated via modulation of the receptor concentration, which occurred via a dynamic, autonomous program for IL-12R expression in control cells as well as those stimulated with IL-12. Additionally, time scale analysis identified the signaling events that limited the cellular response, including ligand binding and receptor degradation.