IP3 receptor-mediated Ca2+ release in naive CD4 T cells dictates their cytokine program (original) (raw)
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Science Signaling, 2014
The guanine nucleotide exchange factor SLAT (SWAP-70-like adaptor of T cells) regulates T cell activation and differentiation by enabling Ca 2+ release from intracellular stores in response to stimulation of the T cell receptor (TCR). We found a TCR-induced association between SLAT and inositol 1,4,5-trisphosphate (IP 3) receptor type 1 (IP 3 R1). The N-terminal region of SLAT, which contains two EF-hand motifs that we determined bound Ca 2+ , and the SLAT pleckstrin homology (PH) domain independently bound to IP 3 R1 by associating with a conserved motif within the IP 3 R1 ligand-binding domain. Disruption of the SLAT-IP 3 R1 interaction with cell-permeable, IP 3 R1-based fusion peptides inhibited TCR-stimulated Ca 2+ signaling, activation of the transcription factor NFAT (nuclear factor of activated T cells), and production of cytokines, suggesting that this interaction is required for optimal T cell activation. The finding that SLAT is an IP 3 R1-interacting protein required for T cell activation suggests that this interaction could be a potential target for a selective immunosuppressive drug. Ca 2+ signaling plays a crucial role in immune responses by regulating many aspects of lymphocyte biology, including development, activation, and effector functions (1). Impaired Ca 2+ signaling in T lymphocytes is linked to pathophysiological processes in several autoimmune and inflammatory diseases (2). Antigen recognition through the T cell receptor (TCR) results in the activation and recruitment of several tyrosine kinases and substrates to
Frontiers in immunology, 2013
the potential involvement of Ca v 1.4 in non-excitable cells as mast cells (McRory et al., 2004) and more recently in mouse T-lymphocytes (Omilusik et al., 2011). CalCium in T-lymphoCyTes: prominenT role of The sTim-orai paThway In T-lymphocytes, Ca 2+ ions are important for the activation of many enzymes including phospholipase C gamma (PLCγ), classical protein kinases C, for proper protein folding, for the accessibility of key enzymes in T-cell transduction, and as a second messenger (Vig and Kinet, 2009). Variations in the intracellular calcium concentration ([Ca] i) are responsible for modulating the transcription of more than 75% of genes induced or down-regulated by T-cell receptor engagement in T-lymphocytes (Feske et al., 2001). The intracellular [Ca] i that decides the cellular fate is tightly regulated in both resting and activated conditions. The calcium concentration in the external medium is about 1-2 mM, whereas the [Ca] i is about 50-100 nM and depends on the calcium channels expressed at both the cell and endoplasmic reticulum (ER) membranes, on exchangers, pumps, … Activation of potassium channels that extrude the potassium from the cell is needed for supporting the electrochemical driving force allowing the calcium influx. In T-lymphocytes, TCR engagement results in a cascade of tyrosine kinase activation, the constitution of a platform transducing the signal with the recruitment of adapters and enzymes such as PLCγ that generates inositol trisphosphate (IP3) and diacylglycerol. IP3 binds to its receptors on the ER membrane leading to the release of ER Ca 2+ stores, which induces a conformational change of STIM1, an ER Ca 2+ sensor. STIM1 then localizes near the cell membrane, and activates the SOCC
Nature immunology, 2008
Store-operated Ca 2+ entry through calcium release-activated calcium channels is the chief mechanism for increasing intracellular Ca 2+ in immune cells. Here we show that mouse T cells and fibroblasts lacking the calcium sensor STIM1 had severely impaired store-operated Ca 2+ influx, whereas deficiency in the calcium sensor STIM2 had a smaller effect. However, T cells lacking either STIM1 or STIM2 had much less cytokine production and nuclear translocation of the transcription factor NFAT. T cell-specific ablation of both STIM1 and STIM2 resulted in a notable lymphoproliferative phenotype and a selective decrease in regulatory T cell numbers. We conclude that both STIM1 and STIM2 promote store-operated Ca 2+ entry into T cells and fibroblasts and that STIM proteins are required for the development and function of regulatory T cells.
Cell Calcium, 2005
Inositol 1,4,5-trisphosphate (IP 3 ) has long been recognized as a second messenger for intracellular Ca 2+ mobilization. Recently, sphingosine 1-phosphate (S1P) has been shown to be involved in Ca 2+ release from the endoplasmic reticulum (ER). Here, we investigated the role of S1P and IP 3 in antigen (Ag)-induced intracellular Ca 2+ mobilization in RBL-2H3 mast cells. Antigen-induced intracellular Ca 2+ mobilization was only partially inhibited by the sphingosine kinase inhibitor dl-threo-dihydrosphingosine (DHS) or the IP 3 receptor inhibitor 2-aminoethoxydiphenyl borate (2-APB), whereas preincubation with both inhibitors led to complete inhibition. In contrast, stimulation of A 3 adenosine receptors with N 5 -ethylcarboxamidoadenosine (NECA) caused intracellular Ca 2+ mobilization that was completely abolished by 2-APB but not by DHS, suggesting that NECA required only the IP 3 pathway, while antigen used both the IP 3 and S1P pathways. Interestingly, however, inhibition of IP 3 production with the phospholipase C inhibitor U73122 completely abolished Ca 2+ release from the ER induced by either stimulant. This suggested that S1P alone, without concomitant production of IP 3 , would not cause intracellular Ca 2+ mobilization. This was further demonstrated in some clones of RBL-2H3 cells excessively overexpressing a  isoform of Class II phosphatidylinositol 3-kinase (PI3KC2). In such clones including clone 5A4C, PI3KC2 was overexpressed throughout the cell, although endogenous PI3KC2 was normally expressed only in the ER. Overexpression of PI3KC2 in the cytosol and the PM led to depletion of phosphatidylinositol 4,5-bisphosphate (PI(4,5)P 2 ), resulting in a marked reduction in IP 3 production. This could explain the abolishment of intracellular Ca 2+ mobilization in clone 5A4C. Supporting this hypothesis, the Ca 2+ mobilization was reconstituted by the addition of exogenous PI(4,5)P 2 in these cells. Our results suggest that both IP 3 and S1P contribute to FcRI-induced Ca 2+ release from the ER and production of IP 3 is necessary for S1P to cause Ca 2+ mobilization from the ER.
Nature Immunology, 2009
Survival of T lymphocytes requires sustained Ca 2+ influx-dependent gene expression. The molecular mechanism, which governs sustained Ca 2+ influx in naive T lymphocytes, is unknown. Here we report an essential role for the β3 regulatory subunit of Ca v channels in the maintenance of naive CD8 + T cells. β3 deficiency resulted in a profound survival defect of CD8 + T cells. This defect correlated with depletion of the pore-forming subunit Ca v 1.4 and attenuation of T cell receptor-mediated global Ca 2+ entry in the absence of β3 in CD8 + T cells. Ca v 1.4 and β3 associated with T cell signaling machinery and Ca v 1.4 localized in lipid rafts. Our data demonstrate a mechanism by which Ca 2+ entry is controlled by a Ca v 1.4-β3 channel complex in T cells. Keywords Calcium; Cacnb3; T lymphocyte; Ca v channel; Apoptosis Calcium (Ca 2+) signaling plays a pivotal role in adaptive immune responses. Sustained Ca 2+ influx across the plasma membrane of T cell leads to the activation of NFAT transcription factors. NFATs regulate survival, activation, proliferation and effector functions of T lymphocytes1-4. Despite the established role of Ca 2+ in T lymphocyte development and functions, the mechanism of sustained Ca 2+ entry in naive T lymphocytes remains elusive to date. A well-characterized mode of Ca 2+ entry in T cells is the CRAC (Calcium Release Activated Ca 2+) pathway, in which, two key players, STIM1 and ORAI1 (also known as Correspondence should be addressed to R.A.F.
Journal of Experimental Medicine, 2007
S ustained elevation of intracellular calcium by Ca 2+ release-activated Ca 2+ channels is required for lymphocyte activation. Sustained Ca 2+ entry requires endoplasmic reticulum (ER) Ca 2+ depletion and prolonged activation of inositol 1,4,5-trisphosphate receptor (IP 3 R)/ Ca 2+ release channels. However, a major isoform in lymphocyte ER, IP3R1, is inhibited by elevated levels of cytosolic Ca 2+ , and the mechanism that enables the prolonged activation of IP 3 R1 required for lymphocyte activation is unclear. We show that IP 3 R1 binds to the scaffolding protein linker of activated T cells and colocalizes with the T cell receptor during activation, resulting in persistent phosphorylation of IP 3 R1 at Tyr353. This phosphorylation increases the sensitivity of the channel to activation by IP 3 and renders the channel less sensitive to Ca 2+-induced inactivation. Expression of a mutant IP3R1-Y353F channel in lymphocytes causes defective Ca 2+ signaling and decreased nuclear factor of activated T cells activation. Thus, tyrosine phosphorylation of IP3R1-Y353 may have an important function in maintaining elevated cytosolic Ca 2+ levels during lymphocyte activation.
Proceedings of the National Academy of Sciences, 1984
The human T-cell leukemia, Jurkat, and a T3-negative mutant of Jurkat (S.5) were used to study the role of T3 in human T-cell activation. Incubation of Jurkat with phytohemagglutinin (PHA) resulted in the production of interleukin 2, which was markedly increased by the addition of phorbol 12-myristate 13-acetate (PMA). Antibodies reactive with T3 could activate Jurkat only if added together with PMA. However, S.5 cells failed to produce interleukin 2 in response to PHA and produced 1/16th the interleukin 2 activity that Jurkat produced in response to PHA and PMA. Incubation of S.5 cells with the calcium ionophore A23187 and PMA resulted in the production of interleukin 2 activity comparable to that produced by Jurkat. Like antibodies reactive with T3, A23187 demonstrated an obligate requirement for PMA in order to activate Jurkat or S.5. These observations suggested that T3 might participate in T-cell activation through mechanisms that increase intracellular Ca2+. This was examined ...
Biochimica et Biophysica Acta (BBA) - Molecular Cell Research, 2018
Inositol 1,4,5-trisphosphate receptors (IP 3 R) are the most widely expressed intracellular Ca 2+ release channels. Their activation by IP 3 and Ca 2+ allows Ca 2+ to pass rapidly from the ER lumen to the cytosol. The resulting increase in cytosolic [Ca 2+ ] may directly regulate cytosolic effectors or fuel Ca 2+ uptake by other organelles, while the decrease in ER luminal [Ca 2+ ] stimulates store-operated Ca 2+ entry (SOCE). We are close to understanding the structural basis of both IP 3 R activation, and the interactions between the ER Ca 2+-sensor, STIM, and the plasma membrane Ca 2+ channel, Orai, that lead to SOCE. IP 3 Rs are the usual means through which extracellular stimuli, through ER Ca 2+ release, stimulate SOCE. Here, we review evidence that the IP 3 Rs most likely to respond to IP 3 are optimally placed to allow regulation of SOCE. We also consider evidence that IP 3 Rs may regulate SOCE downstream of their ability to deplete ER Ca 2+ stores. Finally, we review evidence that IP 3 Rs in the plasma membrane can also directly mediate Ca 2+ entry in some cells.
The FASEB Journal, 2001
Signaling events induced by T-cell receptor (TCR) engagement involve a cascade of tyrosine phosphorylation events leading to activation of several downstream pathways and resulting in cytokine production. TCR-dependent interferon γ (IFN-γ) production by Th1 cells has been shown to require tyrosine phosphorylation of numerous proteins, intracellular Ca 2+ mobilization, and mitogen-activated protein kinase activation. In contrast, the signaling pathways responsible for TCR-dependent interleukin (IL) 4 production remain poorly understood. By using a T-cell hybridoma that displays a hierarchized production of IL-4 and IFN-γ following TCR engagement (IL-4 being produced at a lower threshold of activation than IFN-γ), we showed that IL-4 can be produced in spite of the absence of tyrosine phosphorylation of phospholipase Cγ1. However, protein kinase C (PKC) was found to be translocated to the cell membrane, and an increase in intracellular Ca 2+ concentration was observed. The PKC-dependent Ca 2+ response and IL-4 expression were accounted for by a dihydropyridine-sensitive Ca 2+ entry, which could occur through L-type calcium channels. This pathway was also functional in the D10G4.1 Th2 clone. The fact that this pathway, allowing IL-4 production, did not require optimal activation might explain why low doses of peptides or altered peptide ligands favor Th2 responses.