Silencing of the gene coding for DAP12 induces the appearance of an IDO-dependent, tolerogenic phenotype in CD8^- DCs in response to IFN-γ. (A) RT-PCR analysis of Tyrobp expression in CD8^- DCs treated with Tyrobp-specific siRNA. Control cells were treated with negative control (nc) siRNA. The expression of the Hcst gene, coding for DAP10, was also assayed as a specificity control. (B) Functional IDO activity was measured in terms of kynurenine levels by means of HPLC in supernatants of CD8⁺ or CD8^- DCs treated overnight with IFN-γ. The CD8^- DC fraction was used after Tyrobp gene silencing with specific siRNA or treatment with nc siRNA. Results (mean ± SD) are from 1 of 3 representative experiments. (C) Untreated CD8^- DCs alone or in combination with 5% treated CD8^- DCs (treatment consisting of nc or Tyrobp siRNA for 24 hours followed by IFN-γ for an additional 18 hours) were pulsed with NRP-A7 and transferred to recipient mice to be assayed for skin test reactivity to the eliciting peptide. Experimental groups included the use of cells treated with 2 μM 1-MT. ^*P < .001, experimental versus control footpads. Results are from 1 of 3 representative experiments.