Post-Transcriptional Regulation of Interleukin-7 Receptor Expression

Division Of Basic Sciences - Nci

Investigators

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Abstract

In this project, our main goal has been to address the role of cytokine receptor expression in the development and differentiation of T cells, specifically focusing on the role of gc family cytokine receptors. In the thymus, gc expression is critical for thymopoiesis and for CD8 lineage commitment, whereby IL-7 signaling provides pro-survival and pro-metabolic cues but also induces the expression of Runx3d that specifies CD8 lineage differentiation. In addition to these effects, we recently identified a new role for gc expression in the generation of innate-like CD8 T cells in the thymus. Innate-like CD8 T cells correspond to a small population of terminally differentiated cytotoxic effector T cells that phenotypically appear as antigen-experienced memory cells, expressing large amounts of CD44, and that functionally resemble proinflammatory CD8 T cells, expressing copious amounts of IFNg. However, innate-like CD8 T cells are distinct from conventional effector-memory CD8 T cells as they acquire phenotypic and functional maturity during their generation in the thymus, and prior to antigen exposure. Moreover, innate-like CD8 T cells are thymus-resident while conventional effector memory cells are found in peripheral tissues. Understanding the molecular mechanisms that drive the thymic development and differentiation of innate-like CD8 T cells is an intensely studied subject in T cell immunity. However, the selecting ligand and the environmental cues that promote their generation remain poorly understood. In our recent studies, we identified the cytokine receptor gc as a critical mediator of innate CD8 T cell generation that, surprisingly, promotes their positive selection of innate CD8 T cells even in the absence of the classical MHC-I molecules, H-2 KbDb. Classical MHC-I molecules are critical for the generation of conventional CD8 T cells in the thymus so that KbDb-deficiency results in severe paucity of naive CD8 T cells. Surprisingly, however, we found that the generation of innate CD8 T cells was independent of classical MHC-I molecules. Specifically, we found that overexpression of gc resulted in a dramatic increase of innate CD8 T cells, not only in WT mice, but also in KbDb-deficient mice (Won HY et al., 2023, J. Autoimmunity). These results indicated that the TCR-mediated selection of innate CD8 T cells is independent of classical MHC-I molecules, at least, when driven by the increased abundance of gc protein in thymocytes. We further assessed and mapped its underlying mechanism to the expansion of IL-4-producing invariant NKT (iNKT) cells, so that it is the increased availability of intrathymic IL-4 that augments the selection of innate CD8 T cells. Collectively, these results unravel the selection of innate CD8 T cells being mediated by non-classical MHC-I molecules and being modulated by the abundance of the gc cytokine, IL-4. Because we found that gc expression can substantially alter the subset composition of thymic iNKT cells, we expanded our studies into assessing the effect of other cytokine receptors in iNKT cell differentiation. iNKT cells correspond to a population of thymus-generated T cells with innate-like characteristics and effector functions. Depending on their effector cytokine production, three major subsets of iNKT cells have been identified, namely IFNg-producing NKT1, IL-4-producing NKT2, and IL-17-producing NKT17 cells. Importantly, NKT17 is the only subset that produces the proinflammatory cytokine IL-17. But, how NKT17 cells acquire this ability and what would selectively trigger their activation remain incompletely understood. Because we previously identified the cytokine receptor CD122, a critical component of the IL-15 receptor, as a non-redundant factor in NKT1 cell differentiation, we questioned whether cytokine receptors could be also involved in the differentiation of NKT17 cells. Be screening the expression of a series of gc and non-gc family cytokines, we recently identified the TNF superfamily cytokine receptor Death Receptor 3 (DR3) being specifically expressed on thymic NKT17 cells and mostly absent on other thymic iNKT subsets (Luo S et al., 2023, Cell Mol Life Sci). Importantly, ligation of DR3 molecules using agonistic anti-DR antibodies, but also by stimulation with its natural ligand TL-1A, we showed that DR3 stimulation promoted the in vivo activation of thymic NKT17 cells and provided costimulatory effects upon agonistic alpha-GalCer stimulation. Thus, these results identified a specific surface marker for thymic NKT17 cells that can trigger their activation and augments their effector functions both in vivo and in vitro. We consider these findings important as they expand the scope and relevance of cytokine receptor signaling in iNKT cell generation and differentiation. Thes result further provide new insights for deciphering the role and function of murine NKT17 cells and for understanding the development and activation mechanisms of iNKT cells in general (Liman N and Park JH, 2023, Exp. Mol Med). Whether other cytokine receptors, specifically of the gc family, contribute to the subset differentiation remains a question that we are currently addressing with other genetically engineered mouse models. However, our preliminary results indicate that the role and requirement for cytokines substantially differ depending on the specific T cell type as well as on the tissue environment, and that these factors need to be considered when assessing the individual role of a given cytokine. In this regard, we have recently documented that the survival requirement for thymic and peripheral iNKT cells dramatically differ, so that iNKT cells in the thymus critically depend on IL-15 but iNKT cells in the periphery require IL-7 for their survival (Park JY., 2022, Cell Rep). These results suggested that the tissue-specific cytokine requirement could have been linked to selective or preferential expression of individual cytokine receptors, but these postulations need to be further investigated. As such, we are continuing our efforts to interrogate the regulatory mechanisms of IL-7 and gc receptor expression whereby we address both the transcriptional and epigenetic aspects of cytokine expression, based on tissue origin and cell type. A major part of these studies is addressing the role of the chromatin remodeling zinc finger transcription factor Ikaros (Ikfz1), which we propose as a novel regulator of controlling the accessibility of IL-7Ra gene locus. Our data showed that Ikfz1 controls the cell surface IL-7Ra expression during the early T cell development in the thymus, and affected multiple cytokine receptor expression in the survival/homeostasis of peripheral T cells. To understand how Ikaros contributes to cytokine receptor regulation in general, and specifically to IL-7Ra expression, we are currently analyzing a series of Ikfz1-conditional knock out mice, whereby Ikfz1-floxed mice were crossed with CD2-Cre recombinase transgenic mice or CD4-Cre transgenic mice, among others. In parallel to these studies, we also generated Ikfz1 transgenic mice to test the effect of Ikfz1 overexpression on T cell generation and homeostasis in the context of IL-7R signaling. Analyzing the T cell development and differentiation of these mice in context of IL-7R and other cytokine receptor expression is currently under investigation.