Joonsoo Kang

CD4+ regulatory T cells require CTLA-4 for the maintenance of systemic tolerance

Cytotoxic T lymphocyte antigen-4 (CTLA-4) plays a critical role in negatively regulating T cell responses and has also been implicated in the development and function of natural FOXP3+ regulatory T cells. CTLA-4–deficient mice develop fatal, early onset lymphoproliferative disease. However, chimeric mice containing both CTLA-4–deficient and –sufficient bone marrow (BM)–derived cells do not develop disease, indicating that CTLA-4 can act in trans to maintain T cell self-tolerance. Using genetically mixed blastocyst and BM chimaeras as well as in vivo T cell transfer systems, we demonstrate that in vivo regulation of Ctla4−/− T cells in trans by CTLA-4–sufficient T cells is a reversible process that requires the persistent presence of FOXP3+ regulatory T cells with a diverse TCR repertoire. Based on gene expression studies, the regulatory T cells do not appear to act directly on T cells, suggesting they may instead modulate the stimulatory activities of antigen-presenting cells. These results demonstrate that CTLA-4 is absolutely required for FOXP3+ regulatory T cell function in vivo.

Essential role of the Wnt pathway effector Tcf-1 for the establishment of functional CD8 T cell memory

Immune protection from intracellular pathogens depends on the generation of terminally differentiated effector and of multipotent memory precursor CD8 T cells, which rapidly regenerate effector and memory cells during recurrent infection. The identification of factors and pathways involved in CD8 T cell differentiation is of obvious importance to improve vaccination strategies. Here, we show that mice lacking T cell factor 1 (Tcf-1), a nuclear effector of the canonical Wingless/Integration 1 (Wnt) signaling pathway, mount normal effector and effector memory CD8 T cell responses to infection with lymphocytic choriomeningitis virus (LCMV). However, Tcf-1–deficient CD8 T cells are selectively impaired in their ability to expand upon secondary challenge and to protect from recurrent virus infection. Tcf-1–deficient mice essentially lack CD8 memory precursor T cells, which is evident already at the peak of the primary response, suggesting that Tcf-1 programs CD8 memory cell fate. The function of Tcf-1 to establish CD8 T cell memory is dependent on the catenin-binding domain in Tcf-1 and requires the Tcf-1 coactivators and Wnt signaling intermediates β-catenin and γ-catenin. These findings demonstrate that the canonical Wnt signaling pathway plays an essential role for CD8 central memory T cell differentiation under physiological conditions in vivo. They raise the possibility that modulation of Wnt signaling may be exploited to improve the generation of CD8 memory T cells during vaccination or for therapies designed to promote sustained cytotoxic CD8 T cell responses against tumors.

Identification of transcriptional regulators in the mouse immune system

The differentiation of hematopoietic stem cells into cells of the immune system has been studied extensively in mammals, but the transcriptional circuitry that controls it is still only partially understood. Here, the Immunological Genome Project gene-expression profiles across mouse immune lineages allowed us to systematically analyze these circuits. To analyze this data set we developed Ontogenet, an algorithm for reconstructing lineage-specific regulation from gene-expression profiles across lineages. Using Ontogenet, we found differentiation stage–specific regulators of mouse hematopoiesis and identified many known hematopoietic regulators and 175 previously unknown candidate regulators, as well as their target genes and the cell types in which they act. Among the previously unknown regulators, we emphasize the role of ETV5 in the differentiation of γδ T cells. As the transcriptional programs of human and mouse cells are highly conserved, it is likely that many lessons learned from the mouse model apply to humans.

Dual function of CTLA-4 in regulatory T cells and conventional T cells to prevent multiorgan autoimmunity

Cytotoxic T lymphocyte antigen-4 (CTLA-4) is an inhibitory receptor on T cells essential for maintaining T cell homeostasis and tolerance to self. Mice lacking CTLA-4 develop an early onset, fatal breakdown in T cell tolerance. Whether this autoimmune disease occurs because of the loss of CTLA-4 function in regulatory T cells, conventional T cells, or both is unclear. We show here that lack of CTLA-4 in regulatory T cells leads to aberrant activation and expansion of conventional T cells. However, CTLA-4 expression in conventional T cells prevents aberrantly activated T cells from infiltrating and fatally damaging nonlymphoid tissues. These results demonstrate that CTLA-4 has a dual function in maintaining T cell tolerance: CTLA-4 in regulatory T cells inhibits inappropriate naïve T cell activation and CTLA-4 in conventional T cells prevents the harmful accumulation of self-reactive pathogenic T cells in vital organs.

Tec kinase Itk in γδT cells is pivotal for controlling IgE production in vivo

In conventional αβ T cells, the Tec family tyrosine kinase Itk is required for signaling downstream of the T cell receptor (TCR). Itk also regulates αβ T cell development, lineage commitment, and effector function. A well established feature of Itk−/− mice is their inability to generate T helper type 2 (Th2) responses that produce IL-4, IL-5, and IL-13; yet these mice have spontaneously elevated levels of serum IgE and increased numbers of germinal center B cells. Here we show that the source of this phenotype is γδ T cells, as normal IgE levels are observed in Itk−/−Tcrd−/− mice. When stimulated through the γδ TCR, Itk−/− γδ T cells produce high levels of Th2 cytokines, but diminished IFNγ. In addition, activated Itk−/− γδ T cells up-regulate costimulatory molecules important for B cell help, suggesting that they may directly promote B cell activation and Ig class switching. Furthermore, we find that γδ T cells numbers are increased in Itk−/− mice, most notably the Vγ1.1+Vδ6.3+ subset that represents the dominant population of γδ NKT cells. Itk−/− γδ NKT cells also have increased expression of PLZF, a transcription factor required for αβ NKT cells, indicating a common molecular program between αβ and γδ NKT cell lineages. Together, these data indicate that Itk signaling regulates γδ T cell lineage development and effector function and is required to control IgE production in vivo.

The Necroptosis Adaptor RIPK3 Promotes Injury-Induced Cytokine Expression and Tissue Repair

Programmed necrosis or necroptosis is an inflammatory form of cell death that critically requires the receptor-interacting protein kinase 3 (RIPK3). Here we showed that RIPK3 controls a separate, necrosis-independent pathway of inflammation by regulating cytokine expression in dendritic cells (DCs). Ripk3(-/-) bone-marrow-derived dendritic cells (BMDCs) were highly defective in lipopolysaccharide (LPS)-induced expression of inflammatory cytokines. These effects were caused by impaired NF-κB subunit RelB and p50 activation and by impaired caspase 1-mediated processing of interleukin-1β (IL-1β). This DC-specific function of RIPK3 was critical for injury-induced inflammation and tissue repair in response to dextran sodium sulfate (DSS). Ripk3(-/-) mice exhibited an impaired axis of injury-induced IL-1β, IL-23, and IL-22 cytokine cascade, which was partially corrected by adoptive transfer of wild-type DCs, but not Ripk3(-/-) DCs. These results reveal an unexpected function of RIPK3 in NF-κB activation, DC biology, innate inflammatory-cytokine expression, and injury-induced tissue repair.

Shared and distinct transcriptional programs underlie the hybrid nature of iNKT cells

Invariant natural killer T cells (iNKT cells) are innate-like T lymphocytes that act as critical regulators of the immune response. To better characterize this population, we profiled gene expression in iNKT cells during ontogeny and in peripheral subsets as part of the Immunological Genome Project. High-resolution comparative transcriptional analyses defined developmental and subset-specific programs of gene expression by iNKT cells. In addition, we found that iNKT cells shared an extensive transcriptional program with NK cells, similar in magnitude to that shared with major histocompatibility complex (MHC)-restricted T cells. Notably, the program shared by NK cells and iNKT cells also operated constitutively in γδ T cells and in adaptive T cells after activation. Together our findings highlight a core effector program regulated distinctly in innate and adaptive lymphocytes.

Deletion of p37Ing1 in Mice Reveals a p53-Independent Role for Ing1 in the Suppression of Cell Proliferation, Apoptosis, and Tumorigenesis

ING proteins have been proposed to alter chromatin structure and gene transcription to regulate numerous aspects of cell physiology, including cell growth, senescence, stress response, apoptosis, and transformation. ING1, the founding member of the inhibitor of growth family, encodes p37(Ing1), a plant homeodomain (PHD) protein that interacts with the p53 tumor suppressor protein and seems to be a critical cofactor in p53-mediated regulation of cell growth and apoptosis. In this study, we have generated and analyzed p37(Ing1)-deficient mice and primary cells to further explore the role of Ing1 in the regulation of cell growth and p53 activity. The results show that endogenous levels of p37(Ing1) inhibit the proliferation of p53-wild-type and p53-deficient fibroblasts, and that p53 functions are unperturbed in p37(Ing1)-deficient cells. In addition, loss of p37(Ing1) induces Bax expression and increases DNA damage-induced apoptosis in primary cells and mice irrespective of p53 status. Finally, p37(Ing1) suppresses the formation of spontaneous follicular B-cell lymphomas in mice. These results indicate that p53 does not require p37(Ing1) to negatively regulate cell growth and offers genetic proof that Ing1 suppresses cell growth and tumorigenesis. Furthermore, these data reveal that p37(Ing1) can negatively regulate cell growth and apoptosis in a p53-independent manner.

Immunological Genome Project and systems immunology

Immunological studies of single proteins in a single cell type have been complemented in recent years by larger studies, enabled by emerging high-throughput technologies. This trend has recently been exemplified by the discovery of gene networks controlling regulatory and effector αβ T cell subset development and human hematopoiesis. The Immunological Genome Project (ImmGen) aims to decipher the gene networks underpinning mouse hematopoiesis. The first phase, completed in 2012, profiled the transcriptome of 249 immune cell types. We discuss the utilities of the datasets in high-resolution mapping of the hematopoietic system. The immune transcriptome compendium has revealed unsuspected cell lineage relations and the network reconstruction has identified novel regulatory factors of hematopoiesis.

Molecular determinants of TCR expression and selection.

The process of T cell development in the thymus is tightly regulated, being dependent on the integration of signals required for thymocyte maturation and survival. Rearrangements, expression and signaling of TCR genes play an indispensable role in this developmental program. Recent advances have provided insights into the molecular mechanisms that regulate TCR repertoire formation at the level of alphabeta versus gammadelta T cell fate and CD4(+) versus CD8(+) lineage determination.