scholarly journals IL-2 receptors preassemble and signal in the ER/Golgi causing resistance to antiproliferative anti–IL-2Rα therapies

2019 ◽  
Vol 116 (42) ◽  
pp. 21120-21130 ◽  
Author(s):  
Julianna Volkó ◽  
Ádám Kenesei ◽  
Meili Zhang ◽  
Péter Várnai ◽  
Gábor Mocsár ◽  
...  
Keyword(s):  
T Cell ◽  
Cell Surface ◽  
T Cell Activation ◽  
Cell Activation ◽  
Secretory Pathway ◽  
Interleukin 2 ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Correlation Spectroscopy ◽  
Autocrine Signaling

Interleukin-2 (IL-2) and IL-15 play pivotal roles in T cell activation, apoptosis, and survival, and are implicated in leukemias and autoimmune diseases. Their heterotrimeric receptors share their β- and γc-chains, but have distinct α-chains. Anti–IL-2Rα (daclizumab) therapy targeting cell surface-expressed receptor subunits to inhibit T cell proliferation has only brought limited success in adult T cell leukemia/lymphoma (ATL) and in multiple sclerosis. We asked whether IL-2R subunits could already preassemble and signal efficiently in the endoplasmic reticulum (ER) and the Golgi. A combination of daclizumab and anti–IL-2 efficiently blocked IL-2–induced proliferation of IL-2–dependent wild-type (WT) ATL cells but not cells transfected with IL-2, suggesting that in IL-2–producing cells signaling may already take place before receptors reach the cell surface. In the Golgi fraction isolated from IL-2–producing ATL cells, we detected by Western blot phosphorylated Jak1, Jak3, and a phosphotyrosine signal attributed to the γc-chain, which occurred at much lower levels in the Golgi of WT ATL cells. We expressed EGFP- and mCherry-tagged receptor chains in HeLa cells to study their assembly along the secretory pathway. Confocal microscopy, Förster resonance energy transfer, and imaging fluorescence cross-correlation spectroscopy analysis revealed partial colocalization and molecular association of IL-2 (and IL-15) receptor chains in the ER/Golgi, which became more complete in the plasma membrane, further confirming our hypothesis. Our results define a paradigm of intracellular autocrine signaling and may explain resistance to antagonistic antibody therapies targeting receptors at the cell surface.

scholarly journals ORAI1, STIM1/2, and RYR1 shape subsecond Ca2+microdomains upon T cell activation

2018 ◽  
Vol 11 (561) ◽  
pp. eaat0358 ◽  
Author(s):  
Björn-Philipp Diercks ◽  
René Werner ◽  
Paula Weidemüller ◽  
Frederik Czarniak ◽  
Lola Hernandez ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Ryanodine Receptors ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Super Resolution ◽  
Cell Receptor ◽  
Intracellular Signals

The earliest intracellular signals that occur after T cell activation are local, subsecond Ca2+microdomains. Here, we identified a Ca2+entry component involved in Ca2+microdomain formation in both unstimulated and stimulated T cells. In unstimulated T cells, spontaneously generated small Ca2+microdomains required ORAI1, STIM1, and STIM2. Super-resolution microscopy of unstimulated T cells identified a circular subplasmalemmal region with a diameter of about 300 nm with preformed patches of colocalized ORAI1, ryanodine receptors (RYRs), and STIM1. Preformed complexes of STIM1 and ORAI1 in unstimulated cells were confirmed by coimmunoprecipitation and Förster resonance energy transfer studies. Furthermore, within the first second after T cell receptor (TCR) stimulation, the number of Ca2+microdomains increased in the subplasmalemmal space, an effect that required ORAI1, STIM2, RYR1, and the Ca2+mobilizing second messenger NAADP (nicotinic acid adenine dinucleotide phosphate). These results indicate that preformed clusters of STIM and ORAI1 enable local Ca2+entry events in unstimulated cells. Upon TCR activation, NAADP-evoked Ca2+release through RYR1, in coordination with Ca2+entry through ORAI1 and STIM, rapidly increases the number of Ca2+microdomains, thereby initiating spread of Ca2+signals deeper into the cytoplasm to promote full T cell activation.

scholarly journals Delivery of IL-2 to the T Cell Surface Through Phosphatidylserine Permits Robust Expansion of CD8 T Cells

2021 ◽  
Vol 12 ◽  
Author(s):  
Alana MacDonald ◽  
Brandon Lam ◽  
John Lin ◽  
Louise Ferrall ◽  
Yu Jui Kung ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Surface ◽  
T Cell Activation ◽  
Cd8 T Cells ◽  
Cell Activation ◽  
Interleukin 2 ◽  
Annexin V ◽  
Cell Immunity

The phospholipid phosphatidylserine (PS) is naturally maintained on the cytoplasmic side of the plasma membrane. Independent of apoptosis, PS is redistributed to the surface of CD8 T cells in response to TCR-mediated activation. Annexin V (AnnV) is a protein known to bind PS with high affinity and has been effectively utilized to anchor antigen to the surface of CD8 T cells. To expand these studies, we aimed to exploit TCR activation driven PS exposure as a target to deliver cytokine, namely interleukin-2 (IL-2), to the surface of CD8 T cells. This was accomplished using a novel chimeric fusion protein of annexin V and interleukin 2 (AnnV-IL2). In vitro analysis revealed that AnnV-IL2 is able to specifically bind PS on the T cell surface following TCR stimulation. Consequently, AnnV-IL2 proved to be significantly more effective at enhancing T cell activation compared to recombinant IL-2. In vivo, AnnV-IL2 promotes robust expansion of antigen-specific cells capable of interferon gamma (IFNγ) production when administered following peptide vaccination. Importantly, upon antigen rechallenge, AnnV-IL2 treatment mice demonstrated a stronger secondary expansion, indicating durability of AnnV-IL2 mediated responses. Our data supports the use of AnnV-IL2 to modulate antigen-specific T cell immunity and demonstrates that the PS-AnnV axis is a feasible mechanism to target diverse cargo to CD8 T cells.

scholarly journals CD28-B7 interactions allow the induction of CD8+ cytotoxic T lymphocytes in the absence of exogenous help.

1993 ◽  
Vol 177 (6) ◽  
pp. 1791-1796 ◽  
Author(s):  
F A Harding ◽  
J P Allison
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Cytotoxic T Cells ◽  
Interleukin 2 ◽  
Cell Receptor ◽  
Histocompatibility Complex ◽  
Necessary And Sufficient ◽  
Additional Antigen

The activation requirements for the generation of CD8+ cytotoxic T cells (CTL) are poorly understood. Here we demonstrate that in the absence of exogenous help, a CD28-B7 interaction is necessary and sufficient for generation of class I major histocompatibility complex-specific CTL. Costimulation is required only during the inductive phase of the response, and not during the effector phase. Transfection of the CD28 counter receptor, B7, into nonstimulatory P815 cells confers the ability to elicit P815-specific CTL, and this response can be inhibited by anti-CD28 Fab or by the chimeric B7-binding protein CTLA4Ig. Anti-CD28 monoclonal antibody (mAb) can provide a costimulatory signal to CD8+ T cells when the costimulatory capacity of splenic stimulators is destroyed by chemical fixation. CD28-mediated signaling provokes the release of interleukin 2 (IL-2) from the CD8+ CTL precursors, as anti-CD28 mAb could be substituted for by the addition of IL-2, and an anti-IL-2 mAb can block the generation of anti-CD28-induced CTL. CD4+ cells are not involved in the costimulatory response in the systems examined. We conclude that CD8+ T cell activation requires two signals: an antigen-specific signal mediated by the T cell receptor, and an additional antigen nonspecific signal provided via a CD28-B7 interaction.

scholarly journals SLFN2 protection of tRNAs from stress-induced cleavage is essential for T cell–mediated immunity

Science ◽  
2021 ◽  
Vol 372 (6543) ◽  
pp. eaba4220 ◽  
Author(s):  
Tao Yue ◽  
Xiaoming Zhan ◽  
Duanwu Zhang ◽  
Ruchi Jain ◽  
Kuan-wen Wang ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Interleukin 2 ◽  
Cell Receptor ◽  
Cell Mediated Immunity ◽  
Activated T Cells ◽  
Granule Formation

Reactive oxygen species (ROS) increase in activated T cells because of metabolic activity induced to support T cell proliferation and differentiation. We show that these ROS trigger an oxidative stress response that leads to translation repression. This response is countered by Schlafen 2 (SLFN2), which directly binds transfer RNAs (tRNAs) to protect them from cleavage by the ribonuclease angiogenin. T cell–specific SLFN2 deficiency results in the accumulation of tRNA fragments, which inhibit translation and promote stress-granule formation. Interleukin-2 receptor β (IL-2Rβ) and IL-2Rγ fail to be translationally up-regulated after T cell receptor stimulation, rendering SLFN2-deficient T cells insensitive to interleukin-2’s mitogenic effects. SLFN2 confers resistance against the ROS-mediated translation-inhibitory effects of oxidative stress normally induced by T cell activation, permitting the robust protein synthesis necessary for T cell expansion and immunity.

scholarly journals Antigen-independent activation of naive and memory resting T cells by a cytokine combination.

1994 ◽  
Vol 180 (3) ◽  
pp. 1159-1164 ◽  
Author(s):  
D Unutmaz ◽  
P Pileri ◽  
S Abrignani
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Interleukin 2 ◽  
Effector Function ◽  
Necrosis Factor Alpha ◽  
Naive T Cells ◽  
Naïve T Cells

We investigated whether human resting T cells could be activated to proliferate and display effector function in the absence of T cell receptor occupancy. We report that combination of interleukin 2 (IL-2), tumor necrosis factor alpha, and IL-6 activated highly purified naive (CD45RA+) and memory (CD45RO+) resting CD4+ T cells to proliferate. Under this condition, memory resting T cells could also display effector function as measured by lymphokine synthesis and help for immunoglobulin production by B cells. This novel Ag-independent pathway of T cell activation may play an important role in vivo in recruiting effector T cells at the site of immune response and in maintaining the clonal size of memory T cells in the absence of antigenic stimulation. Moreover, cytokines can induce proliferation of naive T cells without switch to memory phenotype and this may help the maintenance of the peripheral pool of naive T cells.

scholarly journals Actin Cytoskeleton Regulates Calcium Dynamics and NFAT Nuclear Duration

2004 ◽  
Vol 24 (4) ◽  
pp. 1628-1639 ◽  
Author(s):  
Fabiola V. Rivas ◽  
James P. O'Keefe ◽  
Maria-Luisa Alegre ◽  
Thomas F. Gajewski
Keyword(s):  
T Cell ◽  
Actin Cytoskeleton ◽  
T Cell Activation ◽  
Actin Polymerization ◽  
Cell Activation ◽  
Cell Function ◽  
Immunological Synapse ◽  
Interleukin 2 ◽  
Actin Dynamics ◽  
Activated T Cells

ABSTRACT T-cell activation by antigen-presenting cells is accompanied by actin polymerization, T-cell receptor (TCR) capping, and formation of the immunological synapse. However, whether actin-dependent events are required for T-cell function is poorly understood. Herein, we provide evidence for an unexpected negative regulatory role of the actin cytoskeleton on TCR-induced cytokine production. Disruption of actin polymerization resulted in prolonged intracellular calcium elevation in response to anti-CD3, thapsigargin, or phorbol myristate acetate plus ionomycin, leading to persistent NFAT (nuclear factor of activated T cells) nuclear duration. These events were dominant, as the net effect of actin blockade was augmented interleukin 2 promoter activity. Increased surface expression of the plasma membrane Ca2+ ATPase was observed upon stimulation, which was inhibited by cytochalasin D, suggesting that actin polymerization contributes to calcium export. Our results imply a novel role for the actin cytoskeleton in modulating the duration of Ca2+-NFAT signaling and indicate that actin dynamics regulate features of T-cell activation downstream of receptor clustering.

Expression of A2B adenosine receptors in human lymphocytes: their role in T cell activation

1999 ◽  
Vol 112 (4) ◽  
pp. 491-502
Author(s):  
M. Mirabet ◽  
C. Herrera ◽  
O.J. Cordero ◽  
J. Mallol ◽  
C. Lluis ◽  
...  
Keyword(s):  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Human Peripheral Blood ◽  
Human Lymphocytes ◽  
Interleukin 2 ◽  
Cell Receptor ◽  
Physiological Characterization ◽  
A2a Receptors

Extracellular adenosine has a key role in the development and function of the cells of the immune system. Many of the adenosine actions seem to be mediated by specific surface receptors positively coupled to adenylate cyclase: A2A and A2B. Despite the fact that A2A receptors (A2ARs) can be easily studied due to the availability of the specific agonist CGS21680, a pharmacological and physiological characterization of adenosine A2B receptors (A2BRs) in lymphocytes has not been possible due to the lack of suitable reagents. Here we report the generation and characterization of a polyclonal antipeptide antibody raised against the third extracellular loop of the A2BR human clone which is useful for immunocytochemical studies. This antibody has permitted the detection of A2BR+ cells in lymphocyte samples isolated from human peripheral blood. The pharmacology of cAMP-producing compounds is consistent with the presence of functional A2BRs but not of A2A receptors in these human cells. The percentage of A2BR-expressing cells was similar in the CD4(+) or CD8(+) T cell subpopulations. Interestingly activation signals delivered by either phytohemagglutinin or anti-T cell receptor/CD3 complex antibodies led to a significant increase in both the percentage of cells expressing the receptor and the intensity of the labeling. These receptors are functional since interleukin-2 production in these cells is reduced by NECA but not by R-PIA or CGS21680. These results show that A2BR expression is regulated in T cell activation and suggest that the role of adenosine in lymphocyte deactivation is mediated by A2BRs.

Effect of CD28 signal transduction on c-Rel in human peripheral blood T cells

1994 ◽  
Vol 14 (12) ◽  
pp. 7933-7942
Author(s):  
R G Bryan ◽  
Y Li ◽  
J H Lai ◽  
M Van ◽  
N R Rice ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Peripheral Blood ◽  
Cell Activation ◽  
Nuclear Translocation ◽  
Human Peripheral Blood ◽  
Interleukin 2 ◽  
Cell Receptor ◽  
Cd28 Costimulation

Optimal T-cell activation requires both an antigen-specific signal delivered through the T-cell receptor and a costimulatory signal which can be delivered through the CD28 molecule. CD28 costimulation induces the expression of multiple lymphokines, including interleukin 2 (IL-2). Because the c-Rel transcription factor bound to and activated the CD28 response element within the IL-2 promoter, we focused our study on the mechanism of CD28-mediated regulation of c-Rel in human peripheral blood T cells. We showed that CD28 costimulation accelerated the kinetics of nuclear translocation of c-Rel (and its phosphorylated form), p50 (NFKB1), and p65 (RelA). The enhanced nuclear translocation of c-Rel correlated with the stimulation of Il-2 production and T-cell proliferation by several distinct anti-CD28 monoclonal antibodies. This is explained at least in part by the long-term downregulation of I kappa B alpha following CD28 signalling as opposed to phorbol myristate acetate alone. Furthermore, we showed that the c-Rel-containing CD28-responsive complex is enhanced by, but not specific to, CD28 costimulation. Our results indicate that c-Rel is one of the transcription factors targeted by CD28 signalling.

scholarly journals Diacylglycerol kinase ζ limits IL-2-dependent control of PD-1 expression in tumor-infiltrating T lymphocytes

2020 ◽  
Vol 8 (2) ◽  
pp. e001521
Author(s):  
Javier Arranz-Nicolás ◽  
Miguel Martin-Salgado ◽  
Cristina Rodríguez-Rodríguez ◽  
Rosa Liébana ◽  
Maria C Moreno-Ortiz ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Therapeutic Potential ◽  
Interleukin 2 ◽  
Tumor Rejection ◽  
Cytotoxic T Cell ◽  
Adenocarcinoma Cells ◽  
The Impact

BackgroundThe inhibitory functions triggered by the programmed cell death-1 (PD-1) receptor following binding to its ligand (PD-L1) protect healthy organs from cytotoxic T cells, and neutralize antitumor T cell attack. Antibody-based therapies to block PD-1/PD-L1 interaction have yielded notable results, but most patients eventually develop resistance. This failure is attributed to CD8+ T cells achieving hyporesponsive states from which recovery is hardly feasible. Dysfunctional T cell phenotypes are favored by a sustained imbalance in the diacylglycerol (DAG)- and Ca2+-regulated transcriptional programs. In mice, DAG kinase ζ (DGKζ) facilitates DAG consumption, limiting T cell activation and cytotoxic T cell responses. DGKζ deficiency facilitates tumor rejection in mice without apparent adverse autoimmune effects. Despite its therapeutic potential, little is known about DGKζ function in human T cells, and no known inhibitors target this isoform.MethodsWe used a human triple parameter reporter cell line to examine the consequences of DGKζ depletion on the transcriptional restriction imposed by PD-1 ligation. We studied the effect of DGKζ deficiency on PD-1 expression dynamics, as well as the impact of DGKζ absence on the in vivo growth of MC38 adenocarcinoma cells.ResultsWe demonstrate that DGKζ depletion enhances DAG-regulated transcriptional programs, promoting interleukin-2 production and partially counteracting PD-1 inhibitory functions. DGKζ loss results in limited PD-1 expression and enhanced expansion of cytotoxic CD8+ T cell populations. This is observed even in immunosuppressive milieus, and correlates with the reduced ability of MC38 adenocarcinoma cells to form tumors in DGKζ-deficient mice.ConclusionsOur results, which define a role for DGKζ in the control of PD-1 expression, confirm DGKζ potential as a therapeutic target as well as a biomarker of CD8+ T cell dysfunctional states.

scholarly journals Human T cell activation. II. A new activation pathway used by a major T cell population via a disulfide-bonded dimer of a 44 kilodalton polypeptide (9.3 antigen).

1985 ◽  
Vol 161 (6) ◽  
pp. 1513-1524 ◽  
Author(s):  
T Hara ◽  
S M Fu ◽  
J A Hansen
Keyword(s):  
Cell Proliferation ◽  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Interleukin 2 ◽  
Cell Subset ◽  
Receptor Expression ◽  
T Cell Proliferation ◽  
Activation Pathway

In previous studies (17-21), monoclonal antibody (mAb) 9.3 has been shown to react with a major population of human T cells, which include T4+ helper/inducer T cells and T8+ cytotoxic T cells. In this investigation, mAb 9.3 was shown to precipitate a disulfide-bonded dimer of a 44 kD polypeptide. Comodulation experiments showed that this molecule is not linked to T3/Ti or T11 antigens. mAb 9.3 was capable of inducing T cell proliferation in the presence of 12-o-tetradecanoyl phorbol-13-acetate (TPA). This effect was monocyte-independent. T cell activation with mAb 9.3 and TPA was associated with increases in interleukin 2(IL-2) receptor expression and IL-2 secretion. mAb 9.3 did not activate T cells, even with the addition of IL-1 or IL-2. Modulation of the T3 complex did not abolish mAb 9.3-induced T cell proliferation in the presence of TPA. These results suggest that the 9.3 antigen may serve as a receptor for an activation pathway restricted to a T cell subset.

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