scholarly journals Signaling through CD5 Activates a Pathway Involving Phosphatidylinositol 3-Kinase, Vav, and Rac1 in Human Mature T Lymphocytes

1998 ◽  
Vol 18 (3) ◽  
pp. 1725-1735 ◽  
Author(s):  
Sonja I. Gringhuis ◽  
Lou F. M. H. de Leij ◽  
Paul J. Coffer ◽  
Edo Vellenga
Keyword(s):  
T Cell ◽  
T Lymphocytes ◽  
Signaling Pathway ◽  
Tyrosine Phosphorylation ◽  
Interleukin 2 ◽  
Dominant Negative ◽  
Induced Response ◽  
Cam Kinase ◽  
Cam Kinase Iv ◽  
Constitutively Active

ABSTRACT CD5 acts as a coreceptor on T lymphocytes and plays an important role in T-cell signaling and T-cell–B-cell interactions. Costimulation of T lymphocytes with anti-CD5 antibodies results in an increase of the intracellular Ca2+ levels, and subsequently in the activation of Ca2+/calmodulin-dependent (CaM) kinase type IV. In the present study, we have characterized the initial signaling pathway induced by anti-CD5 costimulation. The activation of phosphatidylinositol (PI) 3-kinase through tyrosine phosphorylation of its p85 subunit is a proximal event in the CD5-signaling pathway and leads to the activation of the lipid kinase activity of the p110 subunit. The PI 3-kinase inhibitors wortmannin and LY294002 inhibit the CD5-induced response as assessed in interleukin-2 (IL-2) secretion experiments. The expression of an inactivated Rac1 mutant (Rac1 · N17) in T lymphocytes transfected with an IL-2 promoter-driven reporter construct also abrogates the response to CD5 costimulation, while the expression of a constitutively active Rac1 mutant (Rac1-V12) completely replaces the CD5 costimulatory signal. The Rac1-specific guanine nucleotide exchange factor Vav is heavily phosphorylated on tyrosine residues upon CD5 costimulation, which is a prerequisite for its activation. A role for Vav in the CD5-induced signaling pathway is further supported by the findings that the expression of a dominant negative Vav mutant (Vav-C) completely abolishes the response to CD5 costimulation while the expression of a constitutively active Vav mutant [Vav(Δ1–65)] makes the CD5 costimulation signal superfluous. Wortmannin is unable to block the Vav(Δ1–65)- or Rac1 · V12-induced signals, indicating that both Vav and Rac1 function downstream from PI 3-kinase. Vav and Rac1 both act upstream from the CD5-induced activation of CaM kinase IV, since KN-62, an inhibitor of CaM kinases, and a dominant negative CaM kinase IV mutant block the Vav(Δ1–65)-and Rac1 · V12-mediated signals. We propose a model for the CD5-induced signaling pathway in which the PI 3-kinase lipid products, together with tyrosine phosphorylation, activate Vav, resulting in the activation of Rac1 by the Vav-mediated exchange of GDP for GTP.

scholarly journals Interleukin-2-induced tyrosine phosphorylation of p52shc in T lymphocytes.

1993 ◽  
Vol 268 (24) ◽  
pp. 17659-17661 ◽  
Author(s):  
L.A. Burns ◽  
L.M. Karnitz ◽  
S.L. Sutor ◽  
R.T. Abraham
Keyword(s):  
T Lymphocytes ◽  
Tyrosine Phosphorylation ◽  
Interleukin 2

scholarly journals Maturation of acute T-lymphoblastic leukemia cells after CD2 ligation and subsequent treatment with interleukin-2

Blood ◽  
1994 ◽  
Vol 84 (4) ◽  
pp. 1182-1192 ◽  
Author(s):  
F Mentz ◽  
F Ouaaz ◽  
A Michel ◽  
C Blanc ◽  
P Herve ◽  
...  
Keyword(s):  
T Cell ◽  
Tyrosine Phosphorylation ◽  
Lymphoblastic Leukemia ◽  
Interleukin 1 ◽  
Interleukin 2 ◽  
Subsequent Treatment ◽  
Leukemic Cells ◽  
Cell Phenotype ◽  
Cell Functions

Abstract In this study, we have investigated the ability of various cytokines to induce the maturation of acute lymphoblastic leukemia (T-ALL) cells with early T-cell phenotype. Leukemic blasts from 17 untreated T-ALL patients were assayed for their ability to acquire mature T-cell markers, CD3/T-cell receptor (TCR) in particular, after incubation with one or a combination of recombinant human interleukin-1 (IL-1), IL-2, IL-4, IL-7, and CD2-specific monoclonal antibody (MoAb). IL-7 or IL-2 induced the proliferation of some leukemic cells, whereas sequential cell treatment with CD2-MoAb and then IL-2 promoted CD3/TCR expression on nearly all CD2+ cells (15 of 16), except for 1 T-ALL that developed into CD3-CD16+CD56+ cells. Differentiation of T-ALL cells was also evidenced through the downregulation of CD34 precursor cell antigen, the generation of CD4+ and CD8+ cells from CD4+ CD8+ precursors, and the acquisition of mature T-cell functions. CD2 ligation induced a progressive increase of surface expression of IL-2 receptor alpha (IL- 2R alpha) and IL-2R beta and an accelerated in vitro death of leukemic cells. The ligation of IL-2R by IL-2 rescued T-ALL cells from death and promoted their progression toward more mature cells expressing extracellular CD3/TCR alpha beta complexes. Intracellular analysis indicates that TCR alpha transcription and membrane translocation of both TCR alpha and TCR beta were promoted in these conditions. Analysis of intracellular signals transduced during T-ALL differentiation indicated that CD2-ligation induced Ca2+ influx and that the ligation of CD2 and IL-2R induced distinct tyrosine phosphorylation patterns. The addition of inhibitors of tyrosine phosphorylation abolished T-ALL cell differentiation, which suggests the involvement of tyrosine kinases in this phenomenon. Together, we showed the constant maturation of leukemic early T cells after stimulation of surface CD2 and the high- affinity IL-2R.

scholarly journals Human Platelet Lysate Media Supplement Supports Lentiviral Transduction and Expansion of Human T Lymphocytes While Maintaining Memory Phenotype

2019 ◽  
Vol 2019 ◽  
pp. 1-11 ◽  
Author(s):  
Emanuele Canestrari ◽  
Hayley R. Steidinger ◽  
Brianna McSwain ◽  
Steven J. Charlebois ◽  
Christina Tenenhaus Dann
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Lymphocytes ◽  
Human Platelet ◽  
Interleukin 2 ◽  
Platelet Lysate ◽  
Central Memory ◽  
Protein Profiling ◽  
Memory Phenotype ◽  
Human Platelet Lysate

Immune cell therapy has emerged as a promising approach to treat malignancies that were up until recently only treated on a palliative basis. Chimeric antigen receptor- (CAR-) modified T lymphocytes (T cells) in particular have proven to be very effective for certain hematological malignancies. The production of CAR T cells usually involves viral transduction andex vivoculture of T cells. The aim of this study was to explore the use of human platelet lysate (HPL) compared to two commonly used supplements, human AB serum (ABS) and fetal bovine serum (FBS), for modified T cell production. For studying transduction, activated T cells were transduced with lentivirus to deliver GFP transgenes with three different promoters. Transduction efficiency (percent GFP) was similar among the supplements, and a modest increase in the transgene product (mean fluorescence intensity) was observed when HPL was used as a supplement compared to ABS. To study the effect of supplements on expansion, peripheral blood mononuclear cells (PBMCs) were activated and expanded in the presence of interleukin 2 (IL2) for fourteen days. T cell expansions using HPL and ABS were comparable and slightly less than the expansion obtained with FBS. Interestingly, cells expanded in media supplemented with HPL showed a higher percentage of T cells with a central memory phenotype compared to those expanded in ABS or FBS. Protein profiling revealed that the phenotypic differences may be explained by elevated levels of several cytokines in HPL, including IL7. The results suggest that the use of HPL as a cell culture supplement during the production of modified T cells is a reasonable alternative to ABS. Furthermore, the use of HPL may enhancein vivoperformance of the final product by enriching for central memory T cells that are associated with long-term persistence following adoptive transfer.

scholarly journals The role of Raf-1 in the regulation of extracellular signal-regulated kinase 2 by the T cell antigen receptor.

1994 ◽  
Vol 180 (1) ◽  
pp. 401-406 ◽  
Author(s):  
M Izquierdo ◽  
S Bowden ◽  
D Cantrell
Keyword(s):  
T Cells ◽  
T Cell ◽  
Map Kinases ◽  
Interleukin 2 ◽  
Antigen Receptor ◽  
T Cell Antigen Receptor ◽  
Cell Antigen Receptor ◽  
Cell Antigen ◽  
Constitutively Active

Triggering of the T cell antigen receptor (TCR) complex activates the serine/threonine kinase Raf-1 whose function is necessary for TCR induction of the interleukin 2 gene. Raf-1 has been identified as a candidate mitogen-activated protein (MAP) kinase kinase kinase (MKKK) and thus has the potential to couple the TCR to the activation of the MAP kinases such as ERK2. In the present study, the role of Raf-1 in ERK2 regulation of ERK2 in T cells has been explored. A constitutively active Raf-1 kinase, v-raf, or a dominant inhibitory Raf-1 mutant were expressed transiently from the pEF BOS vector in Jurkat cells and the effects of these Raf-1 mutants on a coexpressed ERK2 reporter was assessed. The action of the constitutively active Raf-1 was to stimulate the ERK2 kinase, whereas the dominant negative version of Raf-1 inhibited the ERK2 activation induced by triggering of the TCR. These data indicate a role for Raf-1 in the regulation of ERK2 in T cells.

Interferon- Activates Multiple STAT Proteins and Upregulates Proliferation-Associated IL-2R, c-myc, and pim-1 Genes in Human T Cells

Blood ◽  
1999 ◽  
Vol 93 (6) ◽  
pp. 1980-1991 ◽  
Author(s):  
Sampsa Matikainen ◽  
Timo Sareneva ◽  
Tapani Ronni ◽  
Anne Lehtonen ◽  
Päivi J. Koskinen ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Lymphocytes ◽  
Cell Biology ◽  
Interleukin 2 ◽  
T Cell Proliferation ◽  
Stat Proteins ◽  
Human T Cells ◽  
T Cell Biology ◽  
Cytokine Stimulation

Interferon- (IFN-) is a pleiotropic cytokine that has antiviral, antiproliferative, and immunoregulatory functions. There is increasing evidence that IFN- has an important role in T-cell biology. We have analyzed the expression ofIL-2R, c-myc, and pim-1 genes in anti-CD3–activated human T lymphocytes. The induction of these genes is associated with interleukin-2 (IL-2)–induced T-cell proliferation. Treatment of T lymphocytes with IFN-, IL-2, IL-12, and IL-15 upregulated IL-2R, c-myc, andpim-1 gene expression. IFN- also sensitized T cells to IL-2–induced proliferation, further suggesting that IFN- may be involved in the regulation of T-cell mitogenesis. When we analyzed the nature of STAT proteins capable of binding to IL-2R,pim-1, and IRF-1 GAS elements after cytokine stimulation, we observed IFN-–induced binding of STAT1, STAT3, and STAT4, but not STAT5 to all of these elements. Yet, IFN- was able to activate binding of STAT5 to the high-affinity IFP53 GAS site. IFN- enhanced tyrosine phosphorylation of STAT1, STAT3, STAT4, STAT5a, and STAT5b. IL-12 induced STAT4 and IL-2 and IL-15 induced STAT5 binding to the GAS elements. Taken together, our results suggest that IFN-, IL-2, IL-12, and IL-15 have overlapping activities on human T cells. These findings thus emphasize the importance of IFN- as a T-cell regulatory cytokine.

H-Ras Is Involved in the Inside-out Signaling Pathway of Interleukin-3–Induced Integrin Activation

Blood ◽  
1999 ◽  
Vol 93 (5) ◽  
pp. 1540-1548 ◽  
Author(s):  
Hirohiko Shibayama ◽  
Naoyuki Anzai ◽  
Stephen E. Braun ◽  
Seiji Fukuda ◽  
Charlie Mantel ◽  
...  
Keyword(s):  
Signaling Pathway ◽  
Dominant Negative ◽  
The Other ◽  
Wild Type ◽  
Integrin Activation ◽  
Mapk Kinase ◽  
Interleukin 3 ◽  
Other Hand ◽  
Inside Out ◽  
Constitutively Active

Abstract The proto-oncogene product, p21ras, has been implicated in the cellular mechanism of adhesion, although its precise role has been controversial. Numerous cytokines and growth-factors activate Ras, which is an important component of their growth-promoting signaling pathways. On the other hand, the role of Ras in cytokine-induced adhesion has not been elucidated. We therefore investigated the function of H-Ras in the inside-out signaling pathway of interleukin-3 (IL-3)–induced integrin activation in the murine Baf3 cell line after transfection of cells with either constitutively active, dominant-negative, or wild-type H-Ras cDNAs. Adhesion of Baf3 cells to fibronectin was induced by IL-3 in a dose-dependent manner via very late antigen-4 (VLA-4; 4β1 integrins) and VLA-5 (5β1 integrins) activation. On the other hand, IL-4 did not induce the adhesion of Baf3 cells to fibronectin, although IL-4 did stimulate the cell proliferation of Baf3 cells. Constitutively active H-Ras–transfected Baf3 cells adhered to fibronectin without IL-3 stimulation through VLA-4 and VLA-5, whereas dominant-negative H-Ras–transfected Baf3 cells showed significantly less adhesion induced by IL-3 compared with wild-type and constitutively active H-Ras–transfected Baf3 cells. Anti-β1 integrin antibody (clone; 9EG7), which is known to change integrin conformation and activate integrins, induced the adhesion of dominant-negative H-Ras–transfected Baf3 cells as much as the other types of H-Ras–transfected Baf3 cells. 8-Br-cAMP, Dibutyryl-cAMP, Ras-Raf-1 pathway inhibitors, and PD98059, a MAPK kinase inhibitor, suppressed proliferation and phosphorylation of MAPK detected by Western blotting with anti–phospho-MAPK antibody, but not adhesion of any type of H-Ras–transfected Baf3 cells, whereas U-73122, a phospholipase C (PLC) inhibitor, suppressed adhesion of these cells completely. These data indicate that H-Ras and PLC, but not Raf-1, MAPK kinase, or the MAPK pathway, are involved in the inside-out signaling pathway of IL-3–induced VLA-4 and VLA-5 activation in Baf3 cells.

scholarly journals Partial signaling by CD8+ T cells in response to antagonist ligands.

1996 ◽  
Vol 184 (1) ◽  
pp. 149-157 ◽  
Author(s):  
C Reis e Sousa ◽  
E H Levine ◽  
R N Germain
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tyrosine Phosphorylation ◽  
Cd8 T Cells ◽  
Inositol Phosphates ◽  
Partial Agonist ◽  
Interleukin 2 ◽  
Cell Receptor ◽  
Cd4 Cells ◽  
Partial Agonists

Structural variants of an agonist peptide-major histocompatibility complex (MHC) molecule ligand can show partial agonist and/or antagonist properties. A number of such altered ligands appear to act as pure antagonists. They lack any detectable ability to induce T cell effector function and have been described as unable to induce calcium transients and turnover of inositol phosphates. This has been interpreted as an inability of these ligands to initiate any T cell receptor (TCR)-dependent signal transduction, with their antagonist properties ascribed to competition with offered agonist for TCR occupancy. Yet antagonists for mature CD8+ T cells can induce positive selection of thymocytes, implying active induction of T cell differentiation events, and partial agonists or agonist/antagonist combinations elicit a distinctive pattern of early TCR-associated tyrosine phosphorylation events in CD4+ T cells. We have therefore directly examined proximal TCR signaling in a CD8+ T cell line in response to various related ligands. TCR engagement with natural peptide-MHC class I agonist resulted in the same pattern of early TCR-associated tyrosine phosphorylation events as seen with CD4+ cells, including accumulation of both the p21 and p23 forms of phosphorylated zeta, phosphorylation of CD3 epsilon, and association of phosphorylated ZAP-70 with the TCR. Two antagonists that lacked the ability to induce any detectable CTL effector response (cytolysis, esterase release, gamma interferon secretion, interleukin-2 receptor alpha upregulation) were nevertheless found to also induce TCR-dependent phosphorylation events. In these cases, there was preferential accumulation of the p21 form of phospho-zeta without net phosphorylation of CD3 epsilon, as well as the association of nonphosphorylated ZAP-70 kinase with the receptor. These data show that variant ligands induce similar TCR-dependent phosphorylation events in CD8+ T cells as first observed in CD4+ cells. More importantly, they demonstrate that some putatively pure antagonists are actually a subset of partial agonists able to induce intracellular biochemical changes through the TCR. This delivery of a partial signal by antagonists raises the possibility that antagonism in some cases may result from active interference with stimulation of effector activity by agonist in mature T cells, while the same variant signal could selectively trigger intracellular events that allow positive without negative selection in thymocytes.

scholarly journals Characterization of Avian γδ T-Cell Subsets afterSalmonella entericaSerovar Typhimurium Infection of Chicks

2010 ◽  
Vol 79 (2) ◽  
pp. 822-829 ◽  
Author(s):  
Jana Pieper ◽  
Ulrich Methner ◽  
Angela Berndt
Keyword(s):  
T Cell ◽  
T Lymphocytes ◽  
Fas Ligand ◽  
Interleukin 2 ◽  
Cell Subset ◽  
Immune Defense ◽  
T Cell Subsets ◽  
Mrna Levels ◽  
Γδ T Cell ◽  
Ifn Γ

ABSTRACTAvian γδ T lymphocytes are frequently found in blood and organs and are assumed to be crucial to the immune defense againstSalmonellainfections of chicks. To elucidate the so-far-unknown immunological features of subpopulations of avian γδ T cells in the course of infection, day-old chicks were infected orally withSalmonella entericaserovar Typhimurium. Until 11 days after infection, the occurrence as well as transcription of the CD8 antigen and immunologically relevant protein genes of CD8α−and CD8α+high(CD8αα+CD8αβ+) γδ cells were analyzed using flow cytometry and quantitative real-time reverse transcription-PCR (RT-PCR) with blood, spleen, thymus, and cecum samples. After infection, an increased percentage of CD8α+highγδ T lymphocytes was found in blood, in spleen, and, with the highest values and most rapidly, in cecum. Within the CD8α+highsubset, a significant rise in the number of CD8αα+cells was accompanied by enhanced CD8α antigen expression and reduced gene transcription of the CD8β chain. CD8αα+and CD8αβ+cells showed elevated transcription for Fas, Fas ligand (FasL), interleukin-2 receptor α (IL-2Rα), and gamma interferon (IFN-γ). While the highest fold changes in mRNA levels were observed in CD8αβ+cells, the mRNA expression rates of CD8αβ+cells never significantly exceeded those of the CD8αα+cells. In conclusion, both CD8α+highγδ T-cell subpopulations (CD8αα+and CD8αβ+) might be a potential source of IFN-γ inSalmonella-infected chicks. However, due to their prominent frequency in blood and organs after infection, the avian CD8αα+γδ T-cell subset seems to be unique and of importance in the course ofSalmonellaTyphimurium infection of very young chicks.

scholarly journals Characterization of CD3+, CD4-, CD8- clones expressing the putative T cell receptor gamma gene product. Analysis of the activation pathways leading to interleukin 2 production and triggering of the lytic machinery.

1987 ◽  
Vol 166 (1) ◽  
pp. 277-282 ◽  
Author(s):  
S Ferrini ◽  
C Bottino ◽  
R Biassoni ◽  
A Poggi ◽  
R P Sekaly ◽  
...  
Keyword(s):  
T Cell ◽  
T Lymphocytes ◽  
T Cell Receptor ◽  
Cell Activation ◽  
Human Peripheral Blood ◽  
Interleukin 2 ◽  
Cell Receptor ◽  
Target Cells ◽  
Product Analysis ◽  
Alpha Beta

Four clones were derived from human peripheral blood T lymphocytes from which CD4+ and CD8+ cells had been removed by treatment with specific mAbs and complement. All expressed the CD2+, 3+, 4-, 8-, T44- phenotype, and did not react with the WT31 mAb, which is specific for a framework determinant of the CD3-associated alpha/beta heterodimer which serves as receptor for antigen on most human T lymphocytes. Surface iodination followed by crosslinking with dithiobis-succinimidyl propionate (DSP) and immunoprecipitation with anti-CD3 mAbs indicated that, in all four clones, the CD3-associated molecules consisted of a major 45 kD band and a minor band of 43 kD. Northern blot analysis showed that mRNA for the gamma chain was expressed at high levels, whereas mRNA for the alpha chain was missing; beta chain mRNA was present in a defective form (1 kb instead of 1.3 kb). These data support the concept that these clones may express, in association with CD3, the molecular product of the T cell receptor gamma genes instead of the typical alpha/beta heterodimer. CD3+, WT31- clones lysed the NK-sensitive K562 target cells and produced IL-2 upon stimulation with PHA. In addition, they released IL-2 after triggering with soluble anti-CD3 mAbs or with an appropriate combination of anti-CD2 mAbs (in the presence of adherent cells). When CD3+, WT31- clones were incubated with an anti-CD3 producing hybridoma as triggering target, the latter was efficiently lysed. Target cell lysis also occurred when a suitable combination of anti-CD2 mAbs-producing hybridomas was used. Therefore, CD3+, WT31- cells appear to use two pathways of cell activation that function also in conventional CD3+, WT31+ T cells, but they lack a third putative pathway initiated by T44 surface molecules.

Long-term effects of intermittent interleukin 2 therapy in patients with HIV infection: characterization of a novel subset of CD4+/CD25+ T cells

Blood ◽  
2002 ◽  
Vol 100 (6) ◽  
pp. 2159-2167 ◽  
Author(s):  
Irini Sereti ◽  
Hector Martinez-Wilson ◽  
Julia A. Metcalf ◽  
Michael W. Baseler ◽  
Claire W. Hallahan ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Lymphocytes ◽  
Cd4 T Cells ◽  
Interleukin 2 ◽  
Hiv Patients ◽  
Activation Markers ◽  
Cd25 Expression ◽  
Cell Counts

The long-term immunologic effects of intermittent interleukin 2 (IL-2) therapy were evaluated in a cross-sectional study by comparing 3 groups: HIV-seronegative volunteers, HIV-infected (HIV+) patients receiving highly active antiretroviral therapy (HAART), and HIV+ patients receiving HAART and intermittent IL-2. Whole-blood immunophenotyping was performed to study expression of the IL-2 receptor chains on T lymphocytes and natural killer cells and to further characterize CD4+/CD25+ T cells. Increased CD25 expression, especially in CD4+ T cells but also in CD8+ T cells, without increases in expression of the β and γ chains of the IL-2 receptor was detected in the IL-2 group. Up to 79% of naive CD4+ T cells (median, 61%) from patients in the IL-2 group expressed CD25, and the number of naive CD4+/CD25+ T cells correlated positively with both the total and naive CD4+ T-cell counts. A discrete population of CD45 double intermediate RA+/RO+CD4+ cells was also preferentially expanded in the IL-2 group, and the number of these cells strongly correlated with the total CD4+ count. Despite increases in CD25 expression, T lymphocytes from patients treated with IL-2 did not have increased expression of early (CD69) or late (CD95) activation markers or evidence of recent proliferation (Ki67). Both CD4+/CD25+ and CD4+/CD25− cells from IL-2–treated HIV+ patients proliferated in response to mitogens, specific antigens, and T-cell-receptor–mediated stimuli. Thus, intermittent administration of IL-2 in HIV+ patients leads to preferential expansion of a unique subset of CD4+ T cells that may represent a critical population in T-cell homeostasis.

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