Arun Kannan

Tec family kinases: Itk signaling and the development of NKT αβ and γδ T cells

The Tec family tyrosine kinase interleukin‐2 inducible T‐cell kinase (Itk) is predominantly expressed in T cells and has been shown to be critical for the development, function and differentiation of conventional αβ T cells. However, less is known about its role in nonconventional T cells such as NKT and γδ T cells. In this minireview, we discuss evidence for a role for Itk in the development of invariant NKT αβ cells, as well as a smaller population NKT‐like γδ T cells. We discuss how these cells take what could be the same signaling pathway regulated by Itk, and interpret it to give different outcomes with regards to development and function.

Signal Transduction via the T cell Antigen Receptor in naïve and effector/memory T cells

T cells play an indispensable role in immune defense against infectious agents, but can also be pathogenic. These T cells develop in the thymus, are exported into the periphery as naïve cells and participate in immune responses. Upon recognition of antigen, they are activated and differentiate into effector and memory T cells. While effector T cells carry out the function of the immune response, memory T cells can last up to the life time of the individual, and are activated by subsequent antigenic exposure. Throughout this life cycle, the T cell uses the same receptor for antigen, the T cell Receptor, a complex multi-subunit receptor. Recognition of antigen presented by peptide/MHC complexes on antigen presenting cells unleashes signaling pathways that control T cell activation at each stage. In this review, we discuss the signals regulated by the T cell receptor in naïve and effector/memory T cells.

IL-2–Inducible T Cell Kinase Tunes T Regulatory Cell Development and Is Required for Suppressive Function

IL-2–inducible T cell kinase (ITK) is a key signaling mediator downstream of TCR, mediating T cell positive selection, as well as innate T cell and CD4+ Th2/Th17 differentiation. In this article, we show that ITK also negatively tunes IL-2–induced expansion of conventional Foxp3-expressing regulatory T cells (Tregs). In vivo, Treg abundance is inversely correlated with ITK expression, and inducible Treg development is inversely dependent on ITK kinase activity. While Treg development normally requires both hematopoietic and thymic MHC class 2 (MHC2) expression, the absence of ITK allows Treg development with MHC2 expression in either compartment, with preference for selection by thymic MHC2, suggesting a gatekeeper role for ITK in ensuring that only Tregs selected by both thymic and hematopoietic MHC2 survive selection. Although ITK suppresses Treg development and is not required for maintenance of neuropilin-1–positive natural Tregs in the periphery, it is indispensable for Treg functional suppression of naive CD4+ T cell–induced colitis in Rag−/− recipients. ITK thus regulates the development and function of Tregs.

IL-2–inducible T-cell kinase modulates TH2-mediated allergic airway inflammation by suppressing IFN-γ in naive CD4+ T cells

BACKGROUND Asthma is a predominantly TH2 cell-dominated inflammatory disease characterized by airway inflammation and a major public health concern affecting millions of persons. The Tec family tyrosine kinase IL-2-inducible T-cell kinase (Itk) is primarily expressed in T cells and critical for the function and differentiation of TH cells. Itk(-/-) mice have a defective TH2 response and are not susceptible to allergic asthma. OBJECTIVE We sought to better understand the role of Itk signaling in TH differentiation programs and in the development and molecular pathology of allergic asthma. METHODS Using a murine model of allergic airway inflammation, we dissected the role of Itk in regulating TH cell differentiation through genetic ablation of critical genes, chromatin immunoprecipitation assays, and house dust mite-driven allergic airway inflammation. RESULTS Peripheral naive Itk(-/-) CD4(+) T cells have substantially increased transcripts and expression of the prototypic TH1 genes Eomesodermin, IFN-γ, T-box transcription factor (T-bet), and IL-12Rβ1. Removal of IFN-γ on the Itk(-/-) background rescues expression of TH2-related genes in TH cells and allergic airway inflammation in Itk(-/-) mice. Furthermore, small hairpin RNA-mediated knockdown of Itk in human peripheral blood T cells results in increased expression of mRNA for IFN-γ and T-bet and reduction in expression of IL-4. CONCLUSION Our results indicate that Itk signals suppress the expression of IFN-γ in naive CD4(+) T cells, which in a positive feed-forward loop regulates the expression of TH1 factors, such as T-bet and Eomesodermin, and suppress development of TH2 cells and allergic airway inflammation.

ITK tunes IL-4-induced development of innate memory CD8+ T cells in a γδ T and invariant NKT cell-independent manner

True memory CD8+ T cells develop post antigenic exposure and can provide life‐long immune protection. More recently, other types of memory CD8+ T cells have been described, such as the memory‐like CD8+ T cells (IMP; CD44hiCD122+) that arise spontaneously in Itk−/− mice, which are suggested to develop as a result of IL‐4 secreted by NKT‐like γδ T or PLZF+ NKT cells found in Itk−/− mice. However, we report here that whereas IMP CD8+ T cell development in Itk−/− mice is dependent on IL‐4/STAT6 signaling, it is not dependent on any γδ T or iNKT cells. Our experiments suggest that the IMP develops as a result of tuning of the CD8+ T cell response to exogenous IL‐4 and TCR triggering by ITK and challenge the current model of IMP CD8+ T cell development as a result of NKT‐like γδ T or iNKT cells. These findings suggest that some naive CD8+ T cells may be preprogrammed by weak homeostatic TCR signals in the presence of IL‐4 to become memory phenotype cells with the ability to elaborate effector function rapidly. The role of ITK in this process suggests a mechanism by which IMP CD8+ T cells can be generated rapidly in response to infection.

Cutting Edge: Drebrin-Regulated Actin Dynamics Regulate IgE-Dependent Mast Cell Activation and Allergic Responses

Mast cells play critical roles in allergic responses. Calcium signaling controls the function of these cells, and a role for actin in regulating calcium influx into cells has been suggested. We have previously identified the actin reorganizing protein Drebrin as a target of the immunosuppressant 3,5-bistrifluoromethyl pyrazole, which inhibits calcium influx into cells. In this study, we show that Drebrin−/− mice exhibit reduced IgE-mediated histamine release and passive systemic anaphylaxis, and Drebrin−/− mast cells also exhibit defects in FcεRI-mediated degranulation. Drebrin−/− mast cells exhibit defects in actin cytoskeleton organization and calcium responses downstream of the FcεRI, and agents that relieve actin reorganization rescue mast cell FcεRI-induced degranulation. Our results indicate that Drebrin regulates the actin cytoskeleton and calcium responses in mast cells, thus regulating mast cell function in vivo.

Itk signals promote neuroinflammation by regulating CD4+ T-cell activation and trafficking.

Here we demonstrate that interleukin-2-inducible T-cell kinase (Itk) signaling in cluster of differentiation 4-positive (CD4+) T cells promotes experimental autoimmune encephalomyelitis (EAE), the animal model of multiple sclerosis (MS). We show that Itk−/− mice exhibit reduced disease severity, and transfer of Itk−/− CD4+ T cells into T cell-deficient recipients results in lower disease severity. We observed a significant reduction of CD4+ T cells in the CNS of Itk−/− mice or recipients of Itk−/− CD4+ T cells during EAE, which is consistent with attenuated disease. Itk−/− CD4+ T cells exhibit defective response to myelin antigen stimulation attributable to displacement of filamentous actin from the CD4+ coreceptor. This results in inadequate transmigration of Itk−/− CD4+ T cells into the CNS and across brain endothelial barriers in vitro. Finally, Itk−/− CD4+ T cells show significant reduction in production of T-helper 1 (Th1) and Th17 cytokines and exhibit skewed T effector/T regulatory cell ratios. These results indicate that signaling by Itk promotes autoimmunity and CNS inflammation, suggesting that it may be a viable target for treatment of MS.

Allele-sensitive mutant, Itkas, reveals that Itk kinase activity is required for Th1, Th2, Th17, and iNKT-cell cytokine production.

Itk−/− mice exhibit defects in the activation, development, and function of CD4+ and CD8+ T cells and iNKT cells. These and other defects in these mice make it difficult to uncouple the developmental versus functional requirement of Itk signaling. Here, we report an allele‐sensitive mutant of Itk (Itkas) whose catalytic activity can be selectively inhibited by analogs of the PP1 kinase inhibitor. We show that Itkas behaves like WT Itk in the absence of the inhibitor and can rescue the development of Itk−/− T cells in mice. Using mice carrying Itkas, we show using its inhibitor that Itk activity is required not only for Th2, Th17, and iNKT‐cell cytokine production, but also surprisingly, for Th1 cytokine production. This work has important implications for understanding the role of Itk signaling in the development versus function of iNKT cells, Th1, Th2, and Th17 cells.

T-Bet independent development of IFNγ secreting natural T helper 1 cell population in the absence of Itk

Th1, Th2, Th9 and Th17 cells are conventional CD4+ effector T cells identified as secretors of prototypical cytokines IFNγ, IL4, IL9, and IL-17A respectively. Recently, populations of natural Th17 and Th1 cells (nTh17 and nTh1) with innate-like phenotype have been identified in the thymus that are distinct from conventional Th17 and Th1 cells. The absence of the Tec family kinase Interleukin-2 inducible T cell kinase (Itk) results in T cell immunodeficiency in mice and humans. Here we show that Itk negatively regulates the development of nTh1 cells that express IFNγ in a Tbet independent manner, and whose expansion can be enhanced by IL4. Furthermore, we show that robust induction of IL4 responses during Trichinella spiralis infection enhance the presence of nTh1 cells. We conclude T cell receptor signaling via Itk controls the development of natural Th1 cells, which are expanded by the presence of IL4.

Structure and function of Tec family kinase Itk

Abstract Itk is a member of the Tec family of kinases that is expressed predominantly in T cells. Itk regulates the T cell receptor signaling pathway to modulate T cell development and T helper cell differentiation, particularly Th2 differentiation. Itk is also important for the development and function of iNKT cells. In this review we discuss current progress on our understanding of the structure, activation and signaling pathway of Itk, in addition to inhibitors that have been developed, which target this kinase. We also place in context the function of Itk, available inhibitors and potential use in treating disease.