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G Bismuth

Publications and source records attributed to G Bismuth.

At least 37 records · Page 2Linked to original sources
PubMed1996

Coordinated regulation in human T cells of nucleotide-hydrolyzing ecto-enzymatic activities, including CD38 and PC-1. Possible role in the recycling of nicotinamide adenine dinucleotide metabolites.

The human leukocyte surface Ag CD38 was recently identified as a nicotinamide adenine dinucleotide (NAD)(+)-glycohydrolase ecto-enzyme, degrading NAD into nicotinamide and ADP-ribose. We show here that expression of CD38 is increased in the Jurkat T cell line after treatment with agents that augment intracellular cAMP, with the permeant cAMP analogue dibutyryl-cAMP (db-cAMP), and also with PMA, which activates protein kinase C. Treatment of human PBL T cells with db-cAMP or submitogenic doses of PMA also increased CD38 expression. Two other nucleotide-hydrolyzing activities were induced on the T cell surface concomitantly with CD38: the human PC-1 molecule, a nucleotide phosphodiesterase/pyrophosphatase that produces AMP from NAD or ADP-ribose, and a nucleotidase that produces adenosine from AMP, but which may be distinct from the CD73 5'-nucleotidase. All three enzymes were up-regulated after stimulation of human peripheral blood T cells with PHA. The coordinated regulation of these ecto-enzymes suggested that, besides a possible signaling function, they may recycle extracellular NAD by degrading it to adenosine and nicotinamide, which can be taken up by cells. In support of this hypothesis, db-cAMP-treated Jurkat cells could degrade extracellular NAD for de novo synthesis of purines, while untreated cells could not. Activated lymphocytes are often located in tissues in which cell death is common. It is suggested that the coordinated expression of these enzymes may allow activated T cells to re-use NAD and nucleotides from dead cells.

ADP-ribosyl Cyclase↗
PubMed1995

Triggering of a sustained calcium response through a p56lck-dependent pathway by exogenous ganglioside GM1 in human T lymphocytes.

Various biologic effects induced by free external gangliosides, including cell-signaling events, have been described in several cell systems. We show in this report that free monosialoganglioside GM1, following its rapid and saturable binding to the cell membrane of human Jurkat T cells, triggers in a few seconds a sustained elevation of the intracellular free calcium concentration. It also induces in parallel the early tyrosine phosphorylation of numerous proteins, including phospholipase C gamma-1. Parallel experiments performed with asialo-GM1 or the ceramide part of the molecule do not reproduce these effects, demonstrating the prominent role played by the sialylated part of the ganglioside. A marked conversion of the T cell-specific tyrosine kinase p56lck to a slow migrating 60-kDa form is also found following GM1 addition. It is accompanied in the same time by an increased kinase activity in p56lck immunoprecipitates. Finally, the marked calcium response and tyrosine phosphorylations triggered by GM1 cannot be observed in a p56lck-negative T cell variant. Together these results demonstrate that the monosialoganglioside GM1 can behave as an authentic activation molecule on human T lymphocytes, likely through a p56lck tyrosine kinase-dependent pathway.

Calcium↗
PubMed1995

The proline-rich region of Vav binds to Grb2 and Grb3-3.

Vav has structural features found in signaling proteins and is expressed only in hematopoietic cells. The recent development of mice Vav -/- has confirmed a major role of Vav in early blood cell development. We previously showed that Vav constitutively interacts with glutathione-S-transferase-Grb2. Coimmunoprecipitation experiments supported the idea of a complex formed by Vav-Grb2 in vivo. This complex is of potential interest in signaling of hematopoietic cells. In this work we localize the domains of Vav and Grb2 involved in this interaction. By the use of an in vivo genetic approach (the double hybrid system) and through in vitro experiments (glutathione-S-transferase fusion proteins) we furnish evidence that the interaction between Vav and Grb2 involves the C-SH3 domain of Grb2 and the proline-rich region located in the N-SH3 of Vav. Furthermore this was confirmed by the use of both Vav and Sos derived proline-rich peptides which blocked the binding. In addition we show that Vav also interacts with Grb3-3, a naturally occurring Grb2 isoform wich lacks functional SH2 domain.

Adaptor Proteins, Signal Transducing↗
PubMed1995

B cell superstimulatory influenza virus (H2-subtype) induces B cell proliferation by a PKC-activating, Ca(2+)-independent mechanism.

The influenza virus hemagglutinin glycoprotein (HA) induces a vigorous B cell proliferation and Ig-synthesis by an unknown activation mechanism, which is susceptible to the inhibitory effects of anti-Ig and anti-class II mAbs. To gain further insight into the activation mode of this T cell-independent, B cell "superstimulatory" virus, we analyzed the sensitivity of H2-subtype virus-mediated B cell activation to the inhibitory effects of various signal transduction-blocking agents and compared it to the well characterized anti-mu-mediated and the LPS-employed pathway. Cyclic-AMP agonists (cAMP-analogues, pentoxifylline, cholera toxin, and forskolin) blocked HA-mediated activation of B cells only at concentrations at least 50-fold higher than required for blocking of anti-mu-induced activation. However, HA-treatment failed to induce an increase in intracellular cAMP levels in responding B cells. The B cell response to HA was highly resistant to calcineurin-inhibitory cyclosporin-A treatment and did not result in a measurable Ca2+ influx. Similarly, HA failed to induce an increase in tyrosine phosphorylations, including phosphorylation of phospholipase C gamma 2. HA-activated B cells showed an increase in membrane-associated protein kinase C activity, and depletion of protein kinase C by pretreatment of B cells with phorbol esters inhibited a subsequent activation by HA. Collectively, our results provide a new example of B cell stimulation by multivalent type-2 Ags, which seems to be mediated by a phosphatidylinositol- and Ca(2+)-independent signaling pathway.

Animals↗
PubMed1995

HIV-1 glycoprotein gp120 disrupts CD4-p56lck/CD3-T cell receptor interactions and inhibits CD3 signaling.

Using the CD4+ human T cell clone P28, we demonstrated that the HIV-1 glycoprotein gp120 inhibited CD3-induced inositol trisphosphate production, calcium influx and T cell proliferation. Additionally, gp120 was shown to dissociate the tyrosine kinase p56lck from CD4 in CEM cells, with a concommittant inhibition of CD4-linked kinase activity. We have addressed the question whether disruption of CD4/p56lck or CD4/CD3-T cell receptor interactions, or both, could account for the inhibitory effect of gp120 in P28 cells. By comparing the effects of various anti-CD4 monoclonal antibodies (mAb) with those of gp120, we show that gp120 and IOT4a modulate CD4 expression, and decrease CD4-associated p56lck and CD4-linked kinase activity at the plasma membrane. In contrast, OKT4A and OKT4 anti-CD4 mAb have no inhibitory effect. Interestingly, gp120 also inhibits CD3-induced Lck activation and cellular tyrosine phosphorylation, particularly of phosphoinositide-specific phospholipase C-gamma-1. Kinetic experiments reveal that the inhibitory effect of gp120 on CD3-induced tyrosine phosphorylation appears as early as 30 min, but culminate when CD4-p56lck complexes disappear from the cell surface after 4 h. These results suggest that a negative signal is triggered by gp120 that results, after a few hours, in down-modulation of CD4-p56lck complexes and the impairment of CD3 signaling. Supporting this hypothesis, gp120 inhibits CD3-linked kinase activity as shown by the inhibition of the phosphorylation of CD3 chains, leading to the inhibition of subsequent signal transduction.

Antibodies, Monoclonal↗
PubMed1995

The proto-oncogene Vav product is constitutively tyrosine-phosphorylated in normal human immature T cells.

The proto-oncogene Vav product is markedly tyrosine-phosphorylated after the recruitment of various receptors of cells of hematopoietic origin, including mature T cells. Recent studies on Vav-deficient mice have clearly implicated the product of the proto-oncogene Vav in intrathymic T cell development. Vav tyrosine phosphorylation is probably crucial to connect early tyrosine kinase(s) to downstream molecular events leading to cell division and maturation that occur in the thymus. We investigated the tyrosine phosphorylation of Vav in human thymocytes. Immunoblotting experiments demonstrate that, as in mature T cells, tyrosine phosphorylation of Vav is induced following thymocyte stimulation through the T cell receptor. The main observation, however, is that an important fraction of cellular Vav is constitutively tyrosine-phosphorylated in freshly isolated cells. This phenomenon takes place apparently both in the CD4+CD8+ and the more mature CD4+CD8- and CD4-CD8+ thymocyte cell subsets. Co-immunoprecipitation experiments showed, moreover, that a small amount of Vav is engaged in the multimolecular complex that includes elements of the T cell receptor and the T cell specific ZAP-70 tyrosine kinase. Altogether, these data suggest that a critical pathway for T cell development in the human thymus likely involves the permanent activation of Vav in vivo.

Cell Differentiation↗
PubMed1995

The intracellular Ca2+ concentration optimal for T cell activation is quite different after ionomycin or CD3 stimulation.

The relationship between the initial increase of intracellular Ca2+ concentration ([Ca2+]i) (measured at the single-cell level with an imaging system) and the ensuing proliferation was examined in a human T cell clone stimulated by a phorbol ester in combination with ionomycin, thapsigargin or an anti-CD3 mAb (monoclonal antibody against the CD3 molecule, UCHT1). From the responses to various ionomycin concentrations, one can define a range of [Ca2+]i values (400-900 nM) which appears optimal for T cell proliferation; lower [Ca2+]i values are suboptimal, higher values are cytotoxic. It was then examined if the [Ca2+]i requirements were similar following anti-CD3 stimulation. [Ca2+]i oscillations elicited by a concentration of UCHT1 (1/1,000) optimal for mitogenicity fall precisely within the 400-900 nM range. However, very low concentrations of UCHT1 (1/100,000) which evoke barely detectable [Ca2+]i responses still cause the cells to proliferate. The possibility that the lower [Ca2+]i requirements observed following anti-CD3 stimulation was due to [Ca2+]i oscillations was tested under conditions which prevented the appearance of these oscillations. It turns out that an oscillatory Ca2+ signal is not more mitogenic than a sustained augmentation of [Ca2+]i. Finally, it was examined if overstimulation via CD3 could have toxic consequences similar to those elicited after ionomycin overstimulation. Large transient [Ca2+]i responses can be observed following anti-CD3 stimulation in appropriate conditions, and namely in T cells pretreated with interleukin-2. These [Ca2+]i augmentations are not cytotoxic. A role for the plasmalemmal Ca2+ pump in the prevention of cytotoxicity can be demonstrated.(ABSTRACT TRUNCATED AT 250 WORDS)

Antibodies↗
PubMed1995

Activation signals are delivered through two distinct epitopes of CD100, a unique 150 kDa human lymphocyte surface structure previously defined by BB18 mAb.

We have previously reported the identification of a glycoprotein dimer, now termed CD100, at the lymphocyte cell surface using a mAb termed BB18. In the present study we isolated a second mAb, termed BD16, recognizing another epitope of this 150 kDa structure. We show that interaction with individual mAb triggers differential functional effects on peripheral blood lymphocytes (PBL) and purified T cells. In particular and of the utmost interest, BD16 mAb strongly inhibits CD3 induced PBL proliferation while it dramatically increases the CD2 induced proliferation. In contrast, after T cell purification, BD16 mAb increases both CD2 and CD3 induced proliferation. It has to be noted that perturbation of the same 150 kDa surface molecule with BB18 mAb exerts no effect on CD2 and CD3 induced proliferations, while inducing a strong lymphocyte proliferation in the presence of submitogenic concentrations of phorbol myristate acetate.

Antibodies, Monoclonal↗
PubMed1994

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

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.

Antibodies, Monoclonal↗
PubMed1994

Antigen recognition by helper T cells elicits a sequence of distinct changes of their shape and intracellular calcium.

BACKGROUND: Helper T-cell activation is initiated in vivo when the T-cell receptor complex recognizes an antigen fragment associated with MHC class II molecules on the surface of an antigen-presenting cell. In most previous studies of this phenomenon, T cells were stimulated not with antigen-presenting cells, but with CD3-specific antibodies. This approach provided considerable understanding of the cascade of molecular events triggered by T-cell receptor stimulation. However, the specific consequences of cell-cell interactions are still poorly understood. We therefore used a dual imaging system that provides simultaneous transmission and fluorescence images to study the morphological changes and variations of intracellular calcium concentration ([Ca2+]i) triggered in a human CD4+ antigen-specific T-cell clone in response to antigen presented by a class II-transfected murine fibroblast. RESULTS: T cells loaded with the Ca(2+)-sensitive fluorescent dye Fura-2 were individually monitored for half an hour following their contact with a monolayer of antigen-pulsed antigen-presenting cells. The response was found to have three distinct phases. During the first few minutes after contact, the T cell moves over the antigen-presenting cells, as if 'scanning' them. After several minutes, an oscillating [Ca2+]i response begins, accompanied by the immobilization of the cell and the retraction of pseudopodia. This rounding-up was probably Ca(2+)-dependent, as it could also be triggered by ionomycin or thapsigargin. Later during the [Ca2+]i response, the T cell becomes flattened and further elongated, suggesting increased adhesion to antigen-presenting cells. CONCLUSIONS: The physiological signal for T-cell activation, antigen recognition, is a three-step process reminiscent of the three steps previously observed in the interaction between neutrophils and endothelial cells. During these successive steps, a mobile, weakly interacting T cell is transformed into an immobile cell fully engaged in the activation pathway. Thus, antigenic recognition is not instantaneous, but evolves slowly by progressive amplification of the signal given by a few antigen molecules, eventually resulting in T-cell activation.

Animals↗
PubMed1994

IL-1 beta amplifies bradykinin-induced prostaglandin E2 production via a phospholipase D-linked mechanism.

The proinflammatory cytokine IL-1 was shown to increase the responsiveness of synovial cells to the potent inflammatory peptide bradykinin (BK). We have investigated the biochemical events linked to this amplifying action of IL-1. Stimulation of synoviocytes with only BK elicited a rapid increase in inositol phosphates and a concomitant accumulation of diacylglycerol (DAG), monoacylglycerol, and free arachidonic acid (AA). In contrast, IL-1 did not stimulate any of these events. Thus, BK can induce AA release via the hydrolysis of phosphatidylinositols by a phospholipase C (PLC). BK also activated a phospholipase D (PLD) to cleave phosphatidylcholine (PC), because it caused an increase in phosphatidic acid (PA) content and a sustained DAG formation, which both were inhibited by ethanol in [3H]myristic acid-labeled cells. Moreover, the addition of ethanol diverted PLD into the formation of phosphatidylethanol (PEt) thus inhibiting the amounts of PA and DAG formed. Priming of synovial cells with rIL-1 beta 24 h before exposure to BK in the presence of ethanol further enhanced the BK-induced formation of PEt. Conversely, preincubation with IL-1 did not influence the BK-induced PLC activation nor did it alter the liberation of AA. Finally, we demonstrated that the IL-1-mediated amplification of PGE2 release in response to BK was reduced by the presence of ethanol in the culture medium, suggesting that part of the synergistic action of IL-1 and BK on prostanoid production was dependent on the activation of the PC-specific PLD pathway.

Bradykinin↗
PubMed1994

Cell calcium signaling via GM1 cell surface gangliosides in the human Jurkat T cell line.

The cell surface ganglioside GM1 is the specific receptor for the B subunit of cholera toxin. We show here that in the human Jurkat T cell line an increase in intracellular free Ca2+ concentration can be elicited by using B subunits to ligate GM1 molecules. This Ca2+ signaling effect is clearly mediated through GM1 because it can be observed after direct insertion of exogenous GM1 in a Jurkat cell variant deficient in GM1 expression. The observed Ca2+ response clearly involves both the release of Ca2+ from intracellular stores and a Ca2+ influx from extracellular spaces. It is sustained in the presence of 1 mM extracellular Ca2+, whereas it becomes transient in Ca(2+)-free medium. We show that the GM1-mediated stimulation partially empties the CD3-dependent and inositol 1,4,5-trisphosphate-sensitive intracellular Ca2+ pool suggesting a dependence of the Ca2+ response from activation of phospholipase C (PLC) metabolism. Accordingly, tyrosine phosphorylation of PLC gamma-1 can be evidenced but only in Jurkat cells highly expressing GM1. GM1 stimulation results in an IL-2 production comparable to that obtained after CD3 activation. Finally, the GM1-linked cell Ca2+ activation pathway is also observed in a Jurkat cell clone lacking Ag-specific receptor expression suggesting that the presence of functional CD3/TCR molecules is not essential for GM1-induced cell Ca2+ response. Altogether, these data show that cell surface gangliosides GM1 may act as a signaling molecule in Jurkat T cells possibly by a new pathway, a finding of importance when considering a possible function for ubiquitous membrane carbohydrate structures in T cell recognition systems.

CD3 Complex↗
PubMed1994

An inhibitor of cytotoxic functions produced by CD8+CD57+ T lymphocytes from patients suffering from AIDS and immunosuppressed bone marrow recipients.

An inhibitor of the cytotoxic functions (ICF) mediated by human immunodeficiency virus (HIV)- or HLA-specific cytotoxic T lymphocytes, natural killer and lymphokine-activated killer (LAK) cells is secreted by CD8+CD57+ T lymphocytes, a subset expanded during infection with HIV and after bone marrow transplantation. We previously showed an apparent molecular mass of 20-30 kDa for this soluble glycosylated concanavalin A-binding inhibitor which is distinct from known cytokines. Here, we report a characterization of the mechanism of action of this CD8+CD57+ ICF. We show that the ICF-induced inhibition of LAK cell cytolytic activity is transient, with a spontaneous recovery of cytolytic potential after 18 h. When testing interactions of ICF with a large set of cytokines we found that the ICF-mediated inhibition of cytotoxic functions is antagonized by two cytokines: recombinant interleukin (rIL)-4 and recombinant interferon (rIFN)-gamma. Finally, we show that ICF acts at the level of cytolytic effector cells, where it induces a significant increase of cyclic AMP (cAMP) level. In contrast, no modification of either cell surface antigen expression or of target/effector cell conjugate formation could be evidenced. Addition of rIL-4 and rIFN-gamma reverses such an increase of cAMP levels and in parallel restores the cytolytic activity. Altogether, these data demonstrate that the glycoprotein ICF produced by CD8+CD57+ cells (1) inhibits cell-mediated cytotoxicity by sensitizing cytolytic effector cells to the cAMP pathway, and (2) is part of a cytokine network controlling cell-mediated cytotoxic functions.

Acquired Immunodeficiency Syndrome↗
PubMed1993

Tyrosine phosphorylation and association with phospholipase C gamma-1 of the GAP-associated 62-kD protein after CD2 stimulation of Jurkat T cell.

Numerous substrates are tyrosine phosphorylated upon CD2 stimulation of human Jurkat T cells using a mitogenic pair of CD2 monoclonal antibodies, including the phospholipase C (PLC)gamma-1-p35/36 complex. Most of these substrates are identically tyrosine phosphorylated after CD3 ligation, suggesting that both stimuli share the same biochemical pathway. We show, however, in this report that a 63-kD protein is specifically phosphorylated on tyrosine residues after ligation of the CD2 molecule. The tyrosine phosphorylation of p63 can be induced independently of other substrates when using a single CD2 mAb recognizing the D66 epitope of the molecule. Importantly, this CD2-induced tyrosine phosphorylation of p63 can also occur in the absence of the CD3 zeta chain membrane expression, and is also distinct from the protein tyrosine kinases p56lck and p59fyn. We demonstrate, moreover, that p63 is physically linked with PLC gamma-1 and p35/36 upon CD2 stimulation. Finally, we also show that a 62-kD protein coimmunoprecipitating with the p21ras GTPase activating protein (GAP) is heavily tyrosine phosphorylated only after CD2 stimulation. This ultimately suggests that p63 may represent in fact the 62-kD protein that associates with GAP after tyrosine phosphorylation. Taken together, these results demonstrate the occurrence in Jurkat cells of a tyrosine kinase pathway specifically coupled to the CD2 molecule. They also suggest a function of the p62-GAP-associated protein as a link between PLC gamma-1 and p21ras activation pathways after CD2 activation.

Antibodies, Monoclonal↗
PubMed1993

Defective calcium response in B-chronic lymphocytic leukemia cells. Alteration of early protein tyrosine phosphorylation and of the mechanism responsible for cell calcium influx.

To study defective signal transduction via the Ag receptor of B-chronic lymphocytic leukemia cells (B-CLL), we examine in this report the Ca2+ response triggered by anti-mu antibody in 23 patients previously classified in three phenotypic groups. Altered Ca2+ changes are essentially found in CLL group II whose leukemic cells are characterized by a marked expression of the CD11b Ag. B-CLL cells from patients with a defective Ca2+ response present an altered pattern of protein tyrosine phosphorylation after anti-mu stimulation in comparison with normal human B cells and B-CLL cells from patients having a normal Ca2+ response. Most of the proteins usually tyrosine phosphorylated after the triggering of cell-surface IgM are concerned. This includes the gamma 1 isoform of phospholipase C, although the protein is normally present in B-CLL cells. These findings suggest that the interruption of the phosphoinositide pathway in B-CLL cells is very proximal, at the level in the signaling cascade between activated surface Ig receptors and protein tyrosine kinases. Simultaneously, we demonstrate that some low responding patients exhibit a decreased Ca2+ response to thapsigargin, an agent known to release intracellular Ca2+ without inositol 1,4,5-trisphosphate production. This suggests that an altered functioning of the mechanism leading to the cell Ca2+ influx in B cells can be also involved in the decreased Ca2+ response observed in B-CLL cells.

Animals↗
PubMed1993

The lymphocyte surface antigen CD38 acts as a nicotinamide adenine dinucleotide glycohydrolase in human T lymphocytes.

The extracellular domain of the lymphocyte surface antigen CD38 has been recently shown to share a high sequence homology with a nicotinamide adenine dinucleotide (NAD+)-specific hydrolyzing enzyme cloned from the ovotestis of the gastropod Aplysia (E. States, D.J., Walseth, T.F., Lee, H. C., Trends Biochem. Sci. 1992. 17:495). In agreement with this finding, we present here evidence that CD38-overexpressing T cells, such as human thymocytes and cells from the human HPB-ALL T cell line, exhibit a NAD(+)-hydrolyzing enzymatic activity present on the outer surface of the cell membrane. In contrast, T lymphocytes with relatively low levels of CD38 marker, such as the human Jurkat cell line, display a lower activity. This suggests a relationship between ecto-NAD+ glycohydrolase activity and CD38 expression, as confirmed here when comparing wild-type Jurkat cells and a Jurkat cell variant overexpressing the CD38 molecule. Moreover, CD38 immunoprecipitates from thymocytes behave as an authentic NAD+ glycohydrolase enzyme: it transforms NAD+ stoichiometrically into nicotinamide plus adenosine 5'-diphosphoribose. Altogether these results strongly support the assumption that CD38 is actually a lymphocyte-specific NAD(+)-hydrolyzing enzyme, a finding that give new prospects to understand the in vivo function of this cell membrane protein.

ADP-ribosyl Cyclase↗
PubMed1992

Calcium fluxes in T lymphocytes.

Mechanisms controlling Ca2+ fluxes through the plasma membrane of lymphocytes have been characterized in a human T-cell clone and in the Jurkat T-cell line. Due to endogenous buffers, about 1/125 of the Ca2+ ions that enter the cell are free. Ca2+ fluxes were estimated from the variations in intracellular Ca2+ concentration ([Ca2+]i) elicited by concentration jumps in extracellular Ca2+ ([Ca2+]o). Thapsigargin was used to inhibit Ca2+ uptake into intracellular stores and to stimulate Ca2+ entry. Ca2+ extrusion was strictly due to the activity of plasma membrane Ca(2+)-ATPases since there was no detectable Na+/Ca2+ exchange activity in these cells. The rate of Ca2+ extrusion was mainly influenced by [Ca2+]i and less by [Ca2+]o but was insensitive to cell depolarization. In depolarized cells, thapsigargin-induced Ca2+ influx was reduced to 10% of the value measured in normally polarized cells, suggesting that depolarization not only reduces the electrochemical gradient for Ca2+ ions, but also inhibits Ca2+ permeation. When Ca2+ ions enter the cell, they bind to a site inside the channel, with a Kd of 3.3 mM. Stimulation of clonal T-cells with low concentrations of either anti-CD3 antibodies or thapsigargin elicited Ca2+ oscillations. Both the amplitude and the frequency of CD3-induced Ca2+ oscillations were sensitive to [Ca2+]o. These oscillations were immediately interrupted when extracellular Ca2+ was removed. The properties of Ca2+ oscillations in T lymphocytes suggest that they are mainly due to variations of Ca2+ influx, modulated by variations in [Ca2+]i.

Antibodies, Monoclonal↗