Antigen presentation by mature dendritic cells (DCs) is the first step for initiating adaptive immune responses. DCs are composed of heterogeneous functional subsets; however, the molecular mechanisms that regulate differentiation of specific DC subsets are not understood. Here, we report that the basic leucine zipper transcription factor NFIL3/E4BP4 is essential for the development of CD8α+ conventional DCs (cDCs). Nfil3−/− mice specifically lack CD8α+ cDCs but not CD8α− cDCs or plasmacytoid DCs in lymphoid tissues. Flt3 ligand–dependent generation of CD8α+ cDCs in lymphoid tissues and CD8α+-equivalent cDCs from Nfil3−/− bone marrow cells was also impaired. NFIL3 regulates CD8α+ cDC development in part through Batf3 expression. Importantly, Nfil3−/− mice exhibited impaired cross-priming of CD8+ T cells against cell-associated antigen, a process normally performed by CD8α+ cDCs, and failed to produce IL-12 after TLR3 stimulation. Thus, NFIL3 plays an essential role in the development of CD8α+ cDCs
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