Kenneth M. Murphy

Dendritic cell regulation of TH1-TH2 development.

Understanding the control exerted by cytokines on T helper cell subsets 1 and 2 (TH1-TH2) development has progressed to a fairly satisfying knowledge of intracellular signals and transcription factors. Less is understood about the molecular basis of TH1-TH2 development exerted by other parameters, such as how the antigen presenting cell can influence this process. Recent work suggests that dendritic cell subsets contribute significant polarizing influences on T helper differentiation, but how this comes about is less clear. In some cases known pathways may be used, as in the dendritic cell subset 1 exerting TH1 polarization by interleukin 12 (IL-12) production and STAT4 activation. In others, the effects are still in need of explanation.

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.

BTLA is a lymphocyte inhibitory receptor with similarities to CTLA-4 and PD-1

During activation, T cells express receptors for receiving positive and negative costimulatory signals. Here we identify the B and T lymphocyte attenuator (BTLA), an immunoglobulin domain–containing glycoprotein with two immunoreceptor tyrosine-based inhibitory motifs. BTLA is not expressed by naive T cells, but it is induced during activation and remains expressed on T helper type 1 (TH1) but not TH2 cells. Crosslinking BTLA with antigen receptors induces its tyrosine phosphorylation and association with the Src homology domain 2 (SH2)-containing protein tyrosine phosphatases SHP-1 and SHP-2, and attenuates production of interleukin 2 (IL-2). BTLA-deficient T cells show increased proliferation, and BTLA-deficient mice have increased specific antibody responses and enhanced sensitivity to experimental autoimmune encephalomyelitis. B7x, a peripheral homolog of B7, is a ligand of BTLA. Thus, BTLA is a third inhibitory receptor on T lymphocytes with similarities to cytotoxic T lymphocyte–associated antigen 4 (CTLA-4) and programmed death 1 (PD-1).

A validated regulatory network for Th17 cell specification.

Th17 cells have critical roles in mucosal defense and are major contributors to inflammatory disease. Their differentiation requires the nuclear hormone receptor RORγt working with multiple other essential transcription factors (TFs). We have used an iterative systems approach, combining genome-wide TF occupancy, expression profiling of TF mutants, and expression time series to delineate the Th17 global transcriptional regulatory network. We find that cooperatively bound BATF and IRF4 contribute to initial chromatin accessibility and, with STAT3, initiate a transcriptional program that is then globally tuned by the lineage-specifying TF RORγt, which plays a focal deterministic role at key loci. Integration of multiple data sets allowed inference of an accurate predictive model that we computationally and experimentally validated, identifying multiple new Th17 regulators, including Fosl2, a key determinant of cellular plasticity. This interconnected network can be used to investigate new therapeutic approaches to manipulate Th17 functions in the setting of inflammatory disease.

Activation of human CD4+ cells with CD3 and CD46 induces a T-regulatory cell 1 phenotype

The immune system must distinguish not only between self and non-self, but also between innocuous and pathological foreign antigens to prevent unnecessary or self-destructive immune responses. Unresponsiveness to harmless antigens is established through central and peripheral processes. Whereas clonal deletion and anergy are mechanisms of peripheral tolerance, active suppression by T-regulatory 1 (Tr1) cells has emerged as an essential factor in the control of autoreactive cells. Tr1 cells are CD4+ T lymphocytes that are defined by their production of interleukin 10 (IL-10) and suppression of T-helper cells; however, the physiological conditions underlying Tr1 differentiation are unknown. Here we show that co-engagement of CD3 and the complement regulator CD46 in the presence of IL-2 induces a Tr1-specific cytokine phenotype in human CD4+ T cells. These CD3/CD46-stimulated IL-10-producing CD4+ cells proliferate strongly, suppress activation of bystander T cells and acquire a memory phenotype. Our findings identify an endogenous receptor-mediated event that drives Tr1 differentiation and suggest that the complement system has a previously unappreciated role in T-cell-mediated immunity and tolerance.

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.

Regulation of interleukin 12 p40 expression through an NF-kappa B half-site.

Interleukin 12 (IL-12) is an inducible cytokine composed of 35- and 40-kDa subunits that is critical for promoting T helper type 1 development and cell-mediated immunity against pathogens. The 40-kDa subunit, expressed by activated macrophages and B cells, is induced by several pathogens in vivo and in vitro and is augmented or inhibited by gamma interferon (IFN-gamma) or IL-10, respectively. Control of IL-12 p40 expression is therefore important for understanding resistance and susceptibility to a variety of pathogens, including Leishmania major and perhaps human immunodeficiency virus. In this report, we provide the first characterization of IL-12 p40 gene regulation in macrophages. We localize inducible activity of the promoter to the sequence -122GGGGAATTTTA-132 not previously recognized to bind Rel family transcription factors. We demonstrate binding of this sequence to NF-kappa B (p50/p65 and p50/c-Rel) complexes in macrophages activated by several p40-inducing pathogens and provide functional data to support a role for NF-kappa B family members in IL-12 p40 activation. Finally, we find that IFN-gamma treatment of cells enhances this binding interaction, thus potentially providing a mechanism for IFN-gamma augmentation of IL-12 production by macrophages.

Host type I IFN signals are required for antitumor CD8+ T cell responses through CD8α+ dendritic cells

The generation of antitumor CD8+ T cell responses requires type I interferon responsiveness in host antigen-presenting cells

B and T lymphocyte attenuator regulates T cell activation through interaction with herpesvirus entry mediator

B and T lymphocyte attenuator (BTLA) provides an inhibitory signal to B and T cells. Previously, indirect observations suggested that B7x was a ligand for BTLA. Here we show that BTLA does not bind B7x; instead, we identify herpesvirus entry mediator (HVEM) as the unique BTLA ligand. BTLA bound the most membrane-distal cysteine-rich domain of HVEM, distinct from regions where the ligands LIGHT and lymphotoxin-α bound HVEM. HVEM induced BTLA tyrosine phosphorylation and association of the tyrosine phosphatase SHP-2 and repressed antigen-driven T cell proliferation, providing an example of reverse signaling to a non–tumor necrosis factor family ligand. The conservation of the BTLA-HVEM interaction between mouse and human suggests that this system is an important pathway regulating lymphocyte activation and/or homeostasis in the immune response.

Type I interferon is selectively required by dendritic cells for immune rejection of tumors

Dendritic cell responsiveness to type I interferon is required for the generation of antitumor T cell responses and tumor rejection.