Brian T. Edelson

Batf3 Deficiency Reveals a Critical Role for CD8α+ Dendritic Cells in Cytotoxic T Cell Immunity

Although in vitro observations suggest that cross-presentation of antigens is mediated primarily by CD8α+ dendritic cells, in vivo analysis has been hampered by the lack of systems that selectively eliminate this cell lineage. We show that deletion of the transcription factor Batf3 ablated development of CD8α+ dendritic cells, allowing us to examine their role in immunity in vivo. Dendritic cells from Batf3–/– mice were defective in cross-presentation, and Batf3–/– mice lacked virus-specific CD8+ T cell responses to West Nile virus. Importantly, rejection of highly immunogenic syngeneic tumors was impaired in Batf3–/– mice. These results suggest an important role for CD8α+ dendritic cells and cross-presentation in responses to viruses and in tumor rejection.

Peripheral CD103+ dendritic cells form a unified subset developmentally related to CD8α+ conventional dendritic cells

Although CD103-expressing dendritic cells (DCs) are widely present in nonlymphoid tissues, the transcription factors controlling their development and their relationship to other DC subsets remain unclear. Mice lacking the transcription factor Batf3 have a defect in the development of CD8α+ conventional DCs (cDCs) within lymphoid tissues. We demonstrate that Batf3−/− mice also lack CD103+CD11b− DCs in the lung, intestine, mesenteric lymph nodes (MLNs), dermis, and skin-draining lymph nodes. Notably, Batf3−/− mice displayed reduced priming of CD8 T cells after pulmonary Sendai virus infection, with increased pulmonary inflammation. In the MLNs and intestine, Batf3 deficiency resulted in the specific lack of CD103+CD11b− DCs, with the population of CD103+CD11b+ DCs remaining intact. Batf3−/− mice showed no evidence of spontaneous gastrointestinal inflammation and had a normal contact hypersensitivity (CHS) response, despite previous suggestions that CD103+ DCs were required for immune homeostasis in the gut and CHS. The relationship between CD8α+ cDCs and nonlymphoid CD103+ DCs implied by their shared dependence on Batf3 was further supported by similar patterns of gene expression and their shared developmental dependence on the transcription factor Irf8. These data provide evidence for a developmental relationship between lymphoid organ–resident CD8α+ cDCs and nonlymphoid CD103+ DCs.

Zbtb46 expression distinguishes classical dendritic cells and their committed progenitors from other immune lineages

The zinc finger transcription factor Zbtb46 specifically marks cDCs and their committed precursors and, when overexpressed in BM progenitors, promotes cDC development at the expense of granulocytes.

MyD88-Dependent but Toll-Like Receptor 2-Independent Innate Immunity to Listeria: No Role for Either in Macrophage Listericidal Activity

We have assessed the requirements for Toll-like receptor (TLR) signaling in vivo during early infection with Listeria monocytogenes. Mice deficient for TLR2, a receptor required for the recognition of Gram-positive peptidoglycan, showed equivalent Listeria resistance to wild-type mice. However, mice deficient for MyD88, an adaptor molecule used by all TLRs, showed profound susceptibility with 3–4 logs greater Listeria burden and severe spleen and liver pathology at day 3 postinfection. Listeria-infected MyD88-deficient mice also showed markedly diminished IFN-γ, TNF-α, and NO responses, despite evidence of macrophage activation and up-regulation of MHC class II molecules. We demonstrate that although minor MyD88-independent responses to live Listeria do occur, these are insufficient for normal host defense. Lastly, we performed experiments in vitro in which macrophages deficient in TLR2 or MyD88 were directly infected with Listeria. Although TLR signaling was required for macrophage NO and cytokine production in response to Listeria, handling and direct killing of Listeria by activated macrophages occurred by TLR2- and MyD88-independent mechanisms.

Compensatory dendritic cell development mediated by BATF-IRF interactions

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.

Role for Spi-C in the development of red pulp macrophages and splenic iron homeostasis

Tissue macrophages comprise a heterogeneous group of cell types differing in location, surface markers and function. Red pulp macrophages are a distinct splenic subset involved in removing senescent red blood cells. Transcription factors such as PU.1 (also known as Sfpi1) and C/EBPα (Cebpa) have general roles in myelomonocytic development, but the transcriptional basis for producing tissue macrophage subsets remains unknown. Here we show that Spi-C (encoded by Spic), a PU.1-related transcription factor, selectively controls the development of red pulp macrophages. Spi-C is highly expressed in red pulp macrophages, but not monocytes, dendritic cells or other tissue macrophages. Spic-/- mice have a cell-autonomous defect in the development of red pulp macrophages that is corrected by retroviral Spi-C expression in bone marrow cells, but have normal monocyte and other macrophage subsets. Red pulp macrophages highly express genes involved in capturing circulating haemoglobin and in iron regulation. Spic-/- mice show normal trapping of red blood cells in the spleen, but fail to phagocytose these red blood cells efficiently, and develop an iron overload localized selectively to splenic red pulp. Thus, Spi-C controls development of red pulp macrophages required for red blood cell recycling and iron homeostasis.

Intracellular Antibody Neutralizes Listeria Growth

We previously reported that treatment of mice with a neutralizing mAb against listeriolysin O (LLO), the pore-forming toxin of Listeria monocytogenes, provided resistance to this intracellular bacterium. We evaluated whether anti-LLO mAb would affect Listeria handling by macrophages, essential cells in Listeria resistance. Macrophages infected in the presence of anti-LLO mAb showed a marked reduction in intracellular Listeria growth, with a concomitant block in LLO-dependent Listeria passage from phagosome to cytosol. Anti-LLO mAb did not opsonize Listeria but, rather, acted within macrophages to neutralize LLO. Importantly, anti-LLO mAb effects on Listeria growth were independent of Fcgamma receptor expression, IFNgamma signaling, and production of nitric oxide and superoxide. These results identify a novel mechanism for antibody control of bacteria within macrophages.

Immunity to Listeria infection

Infection with Listeria monocytogenes is a well studied model for understanding host resistance to intracellular bacteria. Recent advances in the study of Listeria have carefully quantitated the response of CD8(+) T cells to infection and analyzed the effector functions of these cells in vivo. A surprising role for antibody in mediating resistance to Listeria has also recently emerged, providing new insight into the mechanisms of host defense.

CX3CR1+ CD8α+ dendritic cells are a steady-state population related to plasmacytoid dendritic cells

Lymphoid organs are characterized by a complex network of phenotypically distinct dendritic cells (DC) with potentially unique roles in pathogen recognition and immunostimulation. Classical DC (cDC) include two major subsets distinguished in the mouse by the expression of CD8α. Here we describe a subset of CD8α+ DC in lymphoid organs of naïve mice characterized by expression of the CX3CR1 chemokine receptor. CX3CR1+ CD8α+ DC lack hallmarks of classical CD8α+ DC, including IL-12 secretion, the capacity to cross-present antigen, and their developmental dependence on the transcriptional factor BatF3. Gene-expression profiling showed that CX3CR1+ CD8α+ DC resemble CD8α− cDC. The microarray analysis further revealed a unique plasmacytoid DC (PDC) gene signature of CX3CR1+ CD8α+ DC. A PDC relationship of the cells is supported further by the fact that they harbor characteristic D–J Ig gene rearrangements and that development of CX3CR1+ CD8α+ DC requires E2-2, the critical transcriptional regulator of PDC. Thus, CX3CR1+ CD8α+ DC represent a unique DC subset, related to but distinct from PDC. Collectively, the expression-profiling data of this study refine the resolution of previous DC definitions, sharpen the border of classical CD8α+ and CD8α− DC, and should assist the identification of human counterparts of murine DC subsets.

Batf3-Dependent CD11blow/− Peripheral Dendritic Cells Are GM-CSF-Independent and Are Not Required for Th Cell Priming after Subcutaneous Immunization

Dendritic cells (DCs) subsets differ in precursor cell of origin, functional properties, requirements for growth factors, and dependence on transcription factors. Lymphoid-tissue resident CD8α+ conventional DCs (cDCs) and CD11blow/−CD103+ non-lymphoid DCs are developmentally related, each being dependent on FMS-like tyrosine kinase 3 ligand (Flt3L), and requiring the transcription factors Batf3, Irf8, and Id2 for development. It was recently suggested that granulocyte/macrophage colony stimulating factor (GM-CSF) was required for the development of dermal CD11blow/−Langerin+CD103+ DCs, and that this dermal DC subset was required for priming autoreactive T cells in experimental autoimmune encephalitis (EAE). Here, we compared development of peripheral tissue DCs and susceptibility to EAE in GM-CSF receptor deficient (Csf2rb −/−) and Batf3 −/− mice. We find that Batf3-dependent dermal CD11blow/−Langerin+ DCs do develop in Csf2rb −/− mice, but that they express reduced, but not absent, levels of CD103. Further, Batf3 −/− mice lacking all peripheral CD11blow/− DCs show robust Th cell priming after subcutaneous immunization and are susceptible to EAE. Our results suggest that defective T effector priming and resistance to EAE exhibited by Csf2rb −/− mice does not result from the absence of dermal CD11blow/−Langerin+CD103+ DCs.