Compensatory dendritic cell development mediated by BATF-IRF interactions

Abstract

The AP1 transcription factor Batf3 is required for homeostatic development of CD8α(+) classical dendritic cells that prime CD8 T-cell responses against intracellular pathogens. Here we identify an alternative, Batf3-independent pathway in mice for CD8α(+) dendritic cell development operating during infection with intracellular pathogens and mediated by the cytokines interleukin (IL)-12 and interferon-γ. This alternative pathway results from molecular compensation for Batf3 provided by the related AP1 factors Batf, which also functions in T and B cells, and Batf2 induced by cytokines in response to infection. Reciprocally, physiological compensation between Batf and Batf3 also occurs in T cells for expression of IL-10 and CTLA4. Compensation among BATF factors is based on the shared capacity of their leucine zipper domains to interact with non-AP1 factors such as IRF4 and IRF8 to mediate cooperative gene activation. Conceivably, manipulating this alternative pathway of dendritic cell development could be of value in augmenting immune responses to vaccines.

Figure 1. Intracellular pathogens or IL-12 restore lymphoid CD8α⁺ cDCs and tissue-resident CD103⁺ cDCs in Batf3^−/− mice

a, Wild type (WT) and Batf3^−/− (BATF3 KO) 129SvEv mice were uninfected (CTL) or infected with Mtb, and spleens harvested and analyzed by FACS at the indicated time. Histograms for indicated markers are gated as autofluorescent⁻MHCII^highCD11c⁺ cells. Numbers are percent of cells in the gate. b, Serum IL-12 was measured from individual mice (a) at the indicated time. c, Wild type (WT) and Batf3^−/− (BATF3 KO) 129SvEv mice were treated with vehicle (PBS) or IL-12 (IL12) and analyzed by FACS after 3 days as in (a).

Figure 2. IL-12-induced CD8α⁺ cDCs in Batf3^−/− mice can cross-present and mediate tumor rejection

a, From mice in Fig. 1c, DCs were purified by sorting as CD3⁻DX5⁻MHCII⁺CD11c⁺Sirp-α⁻CD24⁺DEC205⁺ DCs (CD8DC) and CD3⁻DX5⁻MHCII⁺CD11c⁺Sirp-α⁺CD24⁻DEC205⁻ DCs (CD4DC) and assayed for cross-presentation. OT-I proliferation in response to cDCs mixed with the indicated number of MHC class I-deficient ovalbumin (Ova)-loaded splenocytes is shown. b, Wild type (WT) or Batf3^−/− (BATF3 KO) mice treated with vehicle (PBS) or with IL-12 (IL12) were inoculated with 1×10⁶ H31m1 fibrosarcomas. Tumor size in individual mice is shown. c, Mice in (b) were analyzed by FACS 11 days after H31m1 inoculation for CD8 T cell infiltration into tumors.

Figure 3. Batf compensates for CD8α⁺ cDC development in Batf3^−/− mice

a, Wild type (WT), or Batf3^−/− (BATF3KO) BM cells were infected with GFP-RV (Empty) or retrovirus expressing the indicated cDNA and cultured with Flt3L. Histograms for the indicated markers are for B220⁻CD11c⁺ cells on day 10. Numbers are the percent of cells in the gate. b, Inguinal lymph nodes from WT, Batf3^−/− (BATF3 KO) or Batf^−/−Batf3^−/− mice (BATF1/3 DKO) on 129SvEv or C57BL/6 backgrounds were analyzed by FACS. Shown are histograms for DEC205 and CD8α. c, CD4 T cells of the indicated genotype were differentiated twice under T_H2 conditions and analyzed by FACS for intracellular IL-10.

Figure 4. Batf2 compensates for Batf3 in CD8α⁺ and CD103⁺ cDC development during T. gondii infection

a, Wild-type (WT) and Batf2^−/− (BATF2KO) mice were infected with T. gondii and monitored for survival. n=29 for WT (dashed line) and Batf2^−/− (solid line) mice. b, Shown are percentages of lung CD103⁺ DCs of total CD45.2⁺ cells for uninfected and infected (T. gondii) mice on day 10. n=5 from one of three experiments. c, WT or Batf3^−/− (BATF3KO) BM cells were infected with the indicated retrovirus, cultured with Flt3L and analyzed by FACS on day 10. d, Groups of 5 mice each of the indicated genotypes were treated with vehicle (PBS) or IL-12 (IL12) and analyzed by FACS after 3 days. Shown are percentages of CD8α⁺ cDCs as a total of splenic cDCs.

Figure 5. BATF leucine zipper interactions with non-AP-1 factors mediate lineage-specific actions

a, Structures of chimeric proteins are shown below a diagram of c-Fos. DNA binding domain (DB), hinge (H), leucine zipper (LZ), amino- (5′) and carboxy-terminus (3′). Flt3L-treated WT or Batf3^−/− (BATF3 KO) BM infected with the indicated retrovirus were analyzed after 10 days. b, 293FT cells expressing both Batf and Irf4 (upper panel) or Batf and Irf4 as indicated (lower panel) were analyzed by EMSA with the indicated probes and antibodies c, 293FT cells expressing Irf4 (+) and the indicated Batf chimera were analyzed by EMSA with the AICE1 probe. d, B cells were analyzed by EMSA with the indicated probe and competitor oligonucleotides (comp). e, Batf3^−/− (BATF3 KO) BM infected with the indicated Batf retroviruses s encoding Batf were analyzed as in (a). f, 293FT cells expressing the indicated Batf mutants were analyzed by EMSA with the AICE1 probe.