A mutation within the SH2 domain of slp‐76 regulates the tissue distribution and cytokine production of iNKT cells in mice

Abstract

TCR ligation is critical for the selection, activation, and integrin expression of T lymphocytes. Here, we explored the role of the TCR adaptor protein slp-76 on iNKT-cell biology. Compared to B6 controls, slp-76 ace/ace mice carrying a missense mutation (Thr428Ile) within the SH2-domain of slp-76 showed an increase in iNKT cells in the thymus and lymph nodes, but a decrease in iNKT cells in spleens and livers, along with reduced ADAP expression and cytokine response. A comparable reduction in iNKT cells was observed in the livers and spleens of ADAP-deficient mice. Like ADAP −/− iNKT cells, slp-76 ace/ace iNKT cells were characterized by enhanced CD11b expression, correlating with an impaired induction of the TCR immediate-early gene Nur77 and a decreased adhesion to ICAM-1. Furthermore, CD11b-intrinsic effects inhibited cytokine release, concanavalin A-mediated inflammation, and iNKT-cell accumulation in the liver. Unlike B6 and ADAP −/− mice, the expression of the transcription factors Id3 and PLZF was reduced, whereas NP-1-expression was enhanced in slp-76 ace/ace mice. Blockade of NP-1 decreased the recovery of iNKT cells from peripheral lymph nodes, identifying NP-1 as an iNKT-cell-specific adhesion factor. Thus, slp-76 contributes to the regulation of the tissue distribution, PLZF, and cytokine expression of iNKT cells via ADAP-dependent and -independent mechanisms.Keywords: ADAP r Cytokine r iNKT cell r Integrin r slp-76Additional supporting information may be found in the online version of this article at the publisher's web-site Eur. J. Immunol. 2016Immunol. . 46: 2121Immunol. -2136 Introduction iNKT cells express a panoply of NK-cell receptors [1] and a canonical TCR through which they recognize (glyco-)lipid antigens [2]. iNKT cells activate similar signaling cascades after TCR ligation like other T lymphocytes [3], but utilize unique transcription factors for their development such as the promyelocytic leukemia zinc finger (PLZF) [4][5][6]. According to two different developmental models PLZF characterizes distinct maturation stages and polarized subsets. The sequential lineage model suggests a gradual decrease of PLZFexpression following selection of iNKT cells which show a Th2-dominated cytokine profile during earlier and a Th1-dominated cytokine profile during later stages of intrathymic maturation [1,7]. The second model describes lineage diversification and simultaneous differentiation into Th1-, Th2-, or Th17-polarized subsets that are defined by the level of PLZF-expression [8]. Although many iNKT cells release both Th1 and Th2 cytokines on a single cell level [9], the production of IL-17 and IFN-γ is mutually exclusive within NK1.1 − cells [10][11][12]. Several transcription factors, such as Egr2, T-bet, ThPOK, Id2, Id3, and the Tec kinases Itk and Rlk have been implicated in the differentiation of iNKT cell subsets [6,8,[13][14][15][16][17] which home to distinct tissues. Specifically, liver and spleen constitute the main source for the Th1-polarized sublineage which is PLZF low . Th2-or ...

The Role of Adaptor Proteins in the Biology of Natural Killer T (NKT) Cells

“…SAP transfers SLAM receptor signals, propagates the thymic selection of iNKT cells and induces the iNKT cell effector program (33). The SH2 domain of slp-76 influences the tissue distribution and phenotype of iNKT cells in the periphery (58). AP-3 interferes with the presentation of glycolipid antigens by CD1d (59).…”

“…For example, one of the pivotal molecules engaged upon TCR ligation is the intracellular adaptor protein slp-76. While the complete absence of slp-76 (54–56) or of its N-terminal region (57) leads to a lack of all peripheral T cell populations, selective mutations in the SH2 domain of slp-76 affect in particular iNKT cells (58). Most importantly and in strict contrast to conventional T cells, the selection of iNKT cells requires co-stimulation via SLAM (signaling lymphocyte-activation molecule) family members (20–24).…”

“…A missense mutation within the SH2-domain of slp-76 led to an accumulation of iNKT cells in the thymus and in peripheral lymph nodes. In contrast, iNKT cells were selectively reduced in the spleens and livers of mice with the same mutation, along with a reduced cytokine response, decreased levels of ADAP protein and altered integrin and NKR expression patterns (58). Although TCR signals were affected by these mutations, NKRs might contribute to the observed phenotype as this mutation affected also synapse formation and elimination of missing-self targets by natural killer (NK) cells (91).…”

“…In addition, the specificity of this mutation for iNKT cells needs to be characterized in further detail by assessing the alterations in subsequent signaling pathways and by screening additional slp-76 mutations. Interestingly, despite exhibiting an NKR distribution that has been associated with enhanced Th1 polarization (7, 38), a simultaneous reduction of both IL-4- and IFN-γ-expression along with a reduced TCR-reactivity was observed in iNKT cells carrying this missense mutation within the SH2-domain of slp-76 (58). Thus, variations in the tissue distribution rather than the cytokine polarization are to be considered in patients with allelic mutations in TCR signaling molecules before pursuing vaccination strategies involving α-GalCer, the prototypical iNKT cell ligand as an adjuvant.…”