IDO-orchestrated crosstalk between pDCs and Tregs inhibits autoimmunity

Lippens, Carla; Duraes, Fernanda V.; Dubrot, Juan; Brighouse, Dale; Lacroix, Mathilde; Irla, Magali; Aubry-Lachainaye, Jean-Pierre; Reith, Walter; Mandl, Judith N.; Hugues, Stéphanie

To clearly demonstrate that MHCII-restricted Ag specific interactions between pDCs and T cells are necessary to induce IDO in pDCs, we used genetically deficient mice that selectively lack MHCII expression by pDCs. These mice have been described before and are deficient for the promoters III and IV (pIII + IV) of CIITA, the master regulator for MHCII expression [38]. In mice, CIITA is under the control of cell specific promoters, pI, pIII and pIV [49], [50] (Supplementary Fig. 2). Absence of pIV leads to MHCII abrogation on cortical thymic epithelial cells (cTECs), resulting in the lack of CD4+ T cell positive selection. To restore CD4+ T cell thymic positive selection by MHCII competent cTECs, bone marrow (BM) cell precursors from pIII + IV−/− mice need to be injected into irradiated WT recipients (pIII + IV−/− → WT) and compared to WT → WT controls. pIII + IV−/− → WT mice exhibiting genetic deficiencies compared to WT → WT, it is possible that distinct immunological environments will affect IDO expression by pDCs. Therefore, to immerse MHCII competent and deficient pDCs in an identical milieu, we performed mixed BM chimeric mice using BM cells from Ubi-eGFP WT and pIII + IV−/− mice (ratio 1:1) that were co-injected into irradiated recipient mice expressing the congenic marker CD45.1 (Fig. 2A). LN cells were sorted 2 months later as pDCs and cDCs from donor BM cells (gated on CD45.2+) and further separated as WT (eGFP+) or pIII + IV−/− (eGFP−) cells (Fig. 2A and B). We confirmed that cDCs expressed little IDO mRNA (Fig. 2C). IDO expression by MHCII deficient pDCs was impaired compared to MHCII competent pDCs in mixed BM chimeric mice (Fig. 2C). Since cells were isolated from the same LNs, decreased IDO expression in absence of MHCII expression by pDCs was not related to different cytokine expression profiles, but linked to a defective MHCII expression by pDCs. An alternative explanation is that CIITA directly acts as a transcription factor regulating IDO gene expression. However, we observed a similar reduction of IDO mRNA in pDCs isolated from H2-Db−/− mice (not shown), ruling out this hypothesis. Altogether, our data demonstrated that MHCII-restricted Ag specific interactions with Tregs are required for the induction of IDO expression by pDCs in steady-state LNs.