The inhibitory effect of RAPA on DC maturation is IL-4 dependent and mediated via FKBP-12 binding. BM-derived DCs were generated with GM-CSF with or without IL-4 and analyzed on day 7. (A-B) In the presence of IL-4, RAPA inhibited the cell surface expression of CD40, CD80, CD86, and MHC class II molecules and the allostimulatory activity of purified CD11c⁺ DCs, whereas FK506 exhibited no effect. Competition for RAPA's intracellular receptor FKBP12 by a molar excess of FK506 (panel A, second column from right) antagonized the inhibitory effects of RAPA on DC maturation. (C-E) In the absence of IL-4 (C-D), or in IL-4Rα—deficient mice (E), RAPA exerted no inhibitory effect on DC surface expression of CD40, CD80, CD86, MHC class II molecules, or T-cell allostimulatory acitivity. (A,C,E) Cells were gated on CD11c. The incidence of CD11c⁺ cells expressing the antigen of interest is indicated. Results show representative data from 10 (A), 3 (B,E), 5 (C), and 2 (D) similar experiments.