RIPK1 blocks T cell senescence mediated by RIPK3 and caspase-8.

Receptor-interacting protein kinase 1 (RIPK1) regulates cell death and inflammation. Here, we show that T cell-specific RIPK1 deficiency in mice leads to the premature senescence of T cells and induces various age-related diseases, resulting in premature death. RIPK1 deficiency causes higher basal activation of mTORC1 (mechanistic target of rapamycin complex 1) that drives enhanced cytokine production, induction of senescence-related genes, and increased activation of caspase-3/7, which are restored by inhibition of mTORC1. Critically, normal aged T cells exhibit similar phenotypes and responses. Mechanistically, a combined deficiency of RIPK3 and caspase-8 inhibition restores the impaired proliferative responses; the elevated activation of Akt, mTORC1, extracellular signal-regulated kinase, and caspase-3/7; and the increased expression of senescence-related genes in RIPK1-deficient CD4 T cells. Last, we revealed that the senescent phenotype of RIPK1-deficient and aged CD4 T cells is restored in the normal tissue environment. Thus, we have clarified the function of RIPK3 and caspase-8 in inducing CD4 T cell senescence, which is modulated by environmental signals.

mTORC1 Signaling Controls TLR2-Mediated T-Cell Activation by Inducing TIRAP Expression.

Effector, but not naïve, T cells are activated by toll-like receptor-2 (TLR2) stimulation, leading to cytokine production and proliferation. We found that the differential response is attributable to the lack of expression of the adaptor protein TIRAP in naive T cells. TIRAP expression is induced upon T-cell receptor (TCR) stimulation and sustained by strong interleukin-2 (IL-2) signals. Expression of TIRAP requires TCR- and IL-2-induced mTORC1 activation. TLR2 stimulation induced the activation of nuclear factor κB (NF-κB) and ERK, leading to much higher production of interferon-γ (IFN-γ) by T helper 1 (Th1) cells cultured in a high concentration of IL-2 than by those cultured in a low concentration of IL-2. In contrast, TLR2 stimulation induces mTORC1 activation through TIRAP, which is essential for TLR2-mediated IFN-γ production. These data demonstrate that the mTORC1 signal confers the response to TLR2 signaling by inducing TIRAP expression and that the TIRAP-mTORC1 axis is critical for TLR2-mediated IFN-γ production by effector T cells.

Reciprocal regulation of STING and TCR signaling by mTORC1 for T-cell activation and function.

Stimulator of interferon genes (STING) plays a key role in detecting cytosolic DNA and induces type I interferon (IFN-I) responses for host defense against pathogens. Although T cells highly express STING, its physiological role remains unknown. Here, we show that costimulation of T cells with the STING ligand cGAMP and TCR leads to IFN-I production and strongly inhibits T-cell growth. TCR-mediated mTORC1 activation and sustained activation of IRF3 are required for cGAMP-induced IFN-I production, and the mTORC1 activity is partially counteracted by cGAMP, thereby blocking proliferation. This mTORC1 inhibition in response to costimulation depends on IRF3 and IRF7. Effector T cells produce much higher IFN-I levels than innate cells in response to cGAMP. Finally, we demonstrated that STING stimulation in T cells is effective in inducing antitumor responses in vivo. Our studies demonstrate that the outputs of STING and TCR signaling pathways are mutually regulated through mTORC1 to modulate T-cell functions.

Bach2 maintains T cells in a naive state by suppressing effector memory-related genes.

The transcriptional repressor BTB and CNC homology 2 (Bach2) is thought to be mainly expressed in B cells with specific functions such as class switch recombination and somatic hypermutation, but its function in T cells is not known. We found equal Bach2 expression in T cells and analyzed its function using Bach2-deficient (-/-) mice. Although T-cell development was normal, numbers of peripheral naive T cells were decreased, which rapidly produced Th2 cytokines after TCR stimulation. Bach2(-/-) naive T cells highly expressed genes related to effector-memory T cells such as CCR4, ST-2 and Blimp-1. Enhanced expression of these genes induced Bach2(-/-) naive T cells to migrate toward CCR4-ligand and respond to IL33. Forced expression of Bach2 restored the expression of these genes. Using Chromatin Immunoprecipitation (ChIP)-seq analysis, we identified S100 calcium binding protein a, Heme oxigenase 1, and prolyl hydroxylase 3 as Bach2 direct target genes, which are highly expressed in effector-memory T cells. These findings indicate that Bach2 suppresses effector memory-related genes to maintain the naive T-cell state and regulates generation of effector-memory T cells.

A critical role for the innate immune signaling molecule IRAK-4 in T cell activation.

IRAK-4 is a protein kinase that is pivotal in mediating signals for innate immune responses. Here, we report that IRAK-4 signaling is also essential for eliciting adaptive immune responses. Thus, in the absence of IRAK-4, in vivo T cell responses were significantly impaired. Upon T cell receptor stimulation, IRAK-4 is recruited to T cell lipid rafts, where it induces downstream signals, including protein kinase C activation through the association with Zap70. This signaling pathway was found to be required for optimal activation of nuclear factor kappaB. Our findings suggest that T cells use this critical regulator of innate immunity for the development of acquired immunity, suggesting that IRAK-4 may be involved in direct signal cross talk between the two systems.

Heterotypic interaction of CRTAM with Necl2 induces cell adhesion on activated NK cells and CD8+ T cells.

NK cells and CD8+ T cells exhibit cytotoxicity and cytokine production upon recognizing target cells through cell-cell interaction. We screened the molecules involved in the recognition and regulation of these cells using cDNA subtraction between naive and activated NK cells. We identified class I-restricted T cell-associated molecule (CRTAM), a two Ig domain-bearing surface receptor, as a molecule rapidly and transiently expressed on NK cells and CD8+ T cells upon activation. CRTAM is expressed as a dimer on the cell surface, and its expression is transcriptionally regulated. Using an expression-cloning system, we then further identified Nectin-like (Necl) molecule 2, a three Ig domain-containing receptor, as a ligand of CRTAM. While Necl2 mediates homotypic interaction, CRTAM interacts with Necl2 but not with CRTAM itself. The heterotypic CRTAM-Necl2 interaction has a higher affinity than the homotypic Necl2 interaction. Although there was no clear alteration in the cytotoxic function of the NK cells and CD8+ T cells against the Necl2-expressing target cells, T cells expressing CRTAM tightly bound to Necl2-expressing cells. CRTAM+ cells did not induce homotypic aggregation but they did exert strong heterotypic binding with Necl2+ cells, which was inhibited by the addition of the CRTAM-Ig fusion protein. These results suggest that the heterotypic interaction between CRTAM and Necl2 plays an important role in the adhesion, interaction or migration of NK cells and CD8+ T cells upon stimulation.