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Results for “Interleukin-2 Receptor alpha Subunit”

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PubMed1992

The augmentation of lymphokine-activated killer activity following pulsing of human peripheral blood mononuclear cells with recombinant human interleukin-2.

The short-term exposure of peripheral blood mononuclear cells (PBMC) to recombinant human interleukin-2 (rhIL-2) at 37 degrees C leads to the generation of lymphokine-activated killer (LAK) activity similar in magnitude to that obtained by the exposure of PBMC to rhIL-2 continuously for 3-5 days. In order to investigate whether the required signal for LAK induction occurred during the short exposure to rhIL-2 or at a later point in the induction phase, PBMC were exposed to rhIL-2 for 1 h at 4 degrees C and then exposed to a low-pH wash to remove bound IL-2 from its receptor. PBMC treated in such a way showed increased LAK activity and proliferation compared to cells exposed to rhIL-2 alone. Expression of the p55 (alpha) subunit of the IL-2 receptor was also increased. In order to cause the augmentation, a lowering of the pH below 4.0 was necessary, and exposure of PBMC to low pH alone (in the absence of rhIL-2) failed to cause activation. Another relevant feature was a transient increase in the expression of the p75 subunit of the IL-2 receptor (beta chain) immediately following the exposure to low pH and the release of interferon gamma, tumour necrosis factor alpha and IL-6; activation was blocked by the inclusion of neutralising antisera raised against rhIL-2 and interferon gamma, thus demonstrating that the endogenous release of these cytokines is important for activation.

Cytokines↗

PubMed1992

Detection of regulatory factors of lymphokine-activated killer cell activity in head and neck cancer patients treated with interleukin-2 and interferon alpha.

Interleukin-2 (IL-2) and interferon-alpha (INF-alpha) are biologic modifiers that have met with limited clinical success in the treatment of human malignancies. We conducted a phase 2 trial of IL-2-IFN-alpha in patients with advanced or unresectable squamous cell carcinoma of the upper aerodigestive tract. Two patients were analyzed sequentially for serum induction phase-blocking factors of lymphokine-activated killer (LAK) cell activity in their therapy. Serum also modulated LAK activity independent of autologous or allogeneic effector cells. Significantly inhibitory serum samples were stable in multiple freezings and thawings. Heat-treating the inhibitory serum, at 56 degrees C for 30 minutes, only partially removed the serum inhibitory capacity. Sequential analysis of p55 and p75, subunits of IL-2 receptors, showed that absence of effector cell lytic activity was associated with markedly decreased fluorescence of the IL-2Rp75 subunit only. No significant alteration of the IL-2Rp55 subunit occurred with therapy. These studies support the theory that lymphocyte and multiple serum factors, developing during IL-2-IFN-alpha therapy, regulate the induction of in vitro LAK activity. Further understanding of these factors may lead to improvements in biologic modifier therapy.

Aged↗

PubMed1992

Mechanism of action of maternal serum on the interleukin2 receptor expression.

Neoplastic Jurkat cells were submitted to a PHA stimulation test after a preincubation in maternal or nulliparous serum (10% dilution). The Il2R expression was significantly downregulated among maternal serum treated cells. Retroplacental serum was significantly more inhibitive than peripheral maternal serum (P less than 0.01). The maternal IgG fractions and mostly the retroplacental IgG fraction proved to contain a factor mainly responsible for the Il2R expression inhibitive property. The molecular mechanism of this phenomenon was further studied. It was shown that H7 (acting as a protein kinase inhibitor) could not influence the Il2R modulation. E.G.T.A., a calcium chelator, was not able to interfere with the inhibitive influence of maternal serum. It was suggested that the maternal serum mediated inhibition of the IL2R expression is not influenced by the hydrolysis of membrane bound phosphatidyl inositol. In contrast, pertussis toxin markedly enhanced, in a dose dependent way, the suppressive influence of maternal serum as compared to nulliparous serum. At low concentrations, pertussis toxin lost its stimulating property and retained its ability to ADP ribosylate the alpha subunit of G proteins inducing a release of adenylcyclase mediating cAMP synthesis. This mechanism has been further studied by the addition of dbc AMP or dbc GMP to Jurkat cells preparations stimulated by PHA. dbc AMP, in a dose-related way, induced a downregulation of the IL2R expression of stimulated neoplastic cells preincubated in nulliparous or maternal serum. dbc GMP did not influence the IL2R expression in the same experimental conditions. The maternal serum mediated cells showed the most pronounced IL2R inhibition. Finally, it was shown that the cAMP synthesis by PHA stimulated Jurkat cells was upregulated in a dose dependent way, after a previous cellular incubation in progressive concentrations of maternal serum. In contrast, among nulliparous serum pretreated cells, cAMP synthesis remained significantly lower, after a lectin stimulation, as compared to the cAMP production derived from retroplacental serum treated and stimulated cells. Taken together, these experiments suggest that the maternal serum dependent suppression of the IL2R expression is related to a protein G stimulation followed by an enhanced cAMP synthesis.

Calcium↗

PubMed1992

TcR-alpha mRNA accumulation does not dictate cell surface TcR/CD3 expression.

The TcR-alpha chain is the last subunit of the TcR/CD3 complex to be expressed during thymic ontogeny. Since the presence of all subunits are required for efficient expression of this complex on the cell surface, this has lead to the hypothesis that the TcR-alpha chain is the limiting subunit which controls cell surface TcR/CD3 expression during thymocyte differentiation. We have examined this issue using a T-lymphoma cell clone, RS4.2, which has a CD4- CD8- Thyl+ J11d+ IL2R+ phenotype, identical with immature thymocytes which have the capacity to generate all major T cell subsets. The RS4.2 cell clone accumulates abundant amounts of TcR-beta, CD3-gamma, -delta, -epsilon and -zeta transcripts, but expresses trace levels of TcR-alpha transcripts and cell surface TcR/CD3. TcR-alpha mRNA levels can be dramatically augmented in RS4.2 cells by three distinct mechanisms: in response to treatment with either phorbol myristate acetate (PMA), calcium ionophore (A23187), or cycloheximide (CHX). However, the expression of TcR/CD3 on the cell surface fails to be increased by any of these agents. In fact, PMA induces a rapid down-regulation of cell surface TcR/CD3 expression. In contrast, these agents trigger an increase in cell surface IL2R expression which coincides with augmented IL2R-alpha mRNA levels. Thus, in RS4.2 cells, IL2R surface expression is controlled by transcript levels, while TcR/CD3 surface expression is regulated by post-transcriptional events, independent of TcR-alpha mRNA accumulation.

Antigens, Differentiation, T-Lymphocyte↗

PubMed1992

Four interleukin-2 surface binding proteins detected in rat spleen cells.

Four specific interleukin-2 (IL-2) surface binding proteins can be detected by covalent cross-linking of [125I]IL-2 to rat spleen cells that have been activated with various stimuli including concanavalin A (Con A), phytohaemagglutinin (PHA), calcium ionophore, and phorbol dibutyrate (PDB) with or without calcium ionophore. These four cross-linked proteins could not be demonstrated in either unstimulated T cells or in activated T cells when binding was performed in the presence of a 20-100-fold excess of unlabelled IL-2. The molecular weights of the four cross-linked proteins, after subtraction of the molecular weight contribution of IL-2 are: 53,000, 70,000, 90,000 and 118,000. The 53,000 MW protein was identified as the rat IL-2 receptor (IL-2R) alpha-chain by immune precipitation. Additionally, results suggest that the rat IL-2R alpha-chain is tightly complexed to both the 118,000 and 90,000 MW IL-2 binding proteins. Purification of surface labelled proteins from activated cells using IL-2 affinity chromatography yields four proteins with similar molecular weight to those identified by cross-linking plus an additional non-ligand cross-linked protein of 46,000 MW. The 46,000 MW band may be a non-binding associated protein since it was not seen following [125I]IL-2 binding cross-linking. Tryptic digests and two-dimensional separation of the affinity-isolated proteins indicate that unique peptide maps are generated for the 46,000, 53,000 and 70,000 MW proteins and excludes the possibility that the bands identified by cross-linking represents cross-linking of multiple ligands to the 53,000 MW subunit. However, the 90,000 and 118,000 MW bands yield peptide maps that closely resemble each other suggesting that these binding proteins may be related. These results suggest that at least four IL-2 surface binding proteins may constitute the rat IL-2R system.

Animals↗

PubMed1992

Cytochalasans and PMA induce IL-2 receptors on CD8+ lymphocytes.

The cytochalasans, fungal metabolites that interact with actin, can affect lymphocyte proliferation; high concentrations inhibit lectin-induced proliferation and low concentrations augment it. The phorbol ester tumor promoter, PMA, alone is not mitogenic for primary lymphocytes but enhances the activity of mitogenic lectins. Because the cytochalasans have been reported to increase intracellular Ca2+ and because PMA activates protein kinase C, lymphocytes were treated with PMA and cytochalasin B (CyB) to determine if this combination would induce DNA synthesis. While this treatment by itself did not cause proliferation, lymphocytes cultured with PMA and CyB overnight, washed, and recultured with IL-2 proliferated to the same degree as lymphocytes stimulated with Con A. Three different cytochalasans, cytochalasin B, cytochalasin D, and chaetoglobosin C, all of which bind to cellular actin with different affinities and only one of which affects glucose transport, induced IL-2 receptors in combination with PMA. Flow cytometric analysis with an antibody to the IL-2 receptor alpha subunit confirmed the induction of receptors on CD8+ cells. However, no IL-2 was produced after the exposure of lymphocytes to the combination of cytochalasans and PMA. Therefore, there was sufficient signal to induce IL-2 receptor expression but not to induce IL-2.

Animals↗

PubMed1992

T helper function of CD4+ cells specific for defined epitopes on the acetylcholine receptor in congenic mouse strains.

We previously identified sequence segments of Torpedo acetylcholine receptor (TAChR) alpha subunit recognized by CD4+ cells of congenic mouse strains of different H-2 haplotypes, susceptible to experimental autoimmune myasthenia gravis. CD4+ cells from BALB/c and CB17 mice (H-2d) recognized the peptide sequences alpha 1-20 and alpha 304-322, while C57BL/6 and BALB/b mice (H-2b) recognized alpha 150-169 and alpha 360-378. C57BL/6 mice recognized to a lesser extent also peptide alpha 181-200. In the present study we demonstrate that CD4+ cells which recognize these epitopes have T-helper function. CD4+ cells from TAChR immunized mice, stimulated in vitro with synthetic epitope peptides, induced proliferation in vitro of B cells via soluble factors which were not strain specific, and induced secretion in vitro of anti-AChR antibodies. Upon in vitro stimulation with T-epitope peptides, they secreted interleukin-2. Immunization of mice with synthetic T-epitope peptides caused sensitization of CD4+ cells, which responded in vitro both to the immunizing peptides and to TAChR, and appearance of anti-AChR antibodies in vivo, further identifying the epitope-specific CD4+ cells as AChR-specific T-helper cells.

Animals↗

PubMed1992

Role of lysophosphatidylcholine in T-lymphocyte activation: involvement of phospholipase A2 in signal transduction through protein kinase C.

2-Lysophosphatidylcholine (lysoPtdCho), a product of the hydrolysis of phosphatidylcholine catalyzed by phospholipase A2, greatly potentiates the activation of human resting T lymphocytes that is induced by a membrane-permeant diacylglycerol plus a calcium ionophore, as determined by the expression of the alpha subunit of the interleukin 2 receptor and thymidine incorporation into DNA. LysoPtdCho per se is inactive unless both diacylglycerol and a calcium ionophore are present. This effect of lysoPtdCho is also observed when diacylglycerol is replaced by a tumor-promoting phorbol ester. Other lysophosphatides including lysophosphatidylserine, lysophosphatidylinositol, and lysophosphatidic acid are inert except for lysophosphatidylethanolamine, which is far less effective than lysoPtdCho. Tracer experiments with radioactive choline indicate that, when T lymphocytes are stimulated with an antigenic signal, lysoPtdCho is indeed produced in a time-dependent fashion, although the concentration of this lysophospholipid accumulated remains to be quantitated. It suggests that phospholipase A2 is directly involved in the signal transduction pathway through protein kinase C to induce long-term cellular responses.

Calcium↗

PubMed1992

The interleukin 2 receptor (IL-2R): the IL-2R alpha subunit alters the function of the IL-2R beta subunit to enhance IL-2 binding and signaling by mechanisms that do not require binding of IL-2 to IL-2R alpha subunit.

Interleukin 2 (IL-2)-mediated signaling through its high-affinity receptor involves a complex interrelationship between IL-2 and two IL-2-binding chains, IL-2R alpha and beta chains. Previously with the reagents available it was difficult to define functional interactions between these two IL-2R subunits involved in IL-2 binding and signal transduction. To extend our understanding of the interplay between the two binding subunits we have done studies with the monoclonal antibody HIEI, which interferes with interaction of IL-2R alpha and beta chains (IL-2R alpha and IL-2R beta, respectively). Furthermore, we used two forms of IL-2, recombinant native IL-2 and F42A, an IL-2 analog (Phe-42----Ala substitution) that binds only to IL-2R beta. Analog F42A manifested 75-100% of the bioactivity of wild-type IL-2. This observation is inconsistent with the strict hierarchical IL-2-binding affinity conversion model previously proposed by Saito and coworkers [Saito Y., Sabe, H., Suzuki, N., Kondo, S., Ogura, T., Shimizu, A. & Honjo, T. (1988) J. Exp. Med. 168, 1563-1572] that predicted an ordered sequence of events in which IL-2 must first bind to IL-2R alpha before its interaction with IL-2R beta. Previous investigations using IL-2 variants were interpreted to show that IL-2R alpha merely acts to concentrate IL-2 to the cell surface and that no other meaningful interaction occurred between IL-2R alpha and IL-2R beta. However, our data are inconsistent with this view. We draw this conclusion on the basis of our observation that antibody HIEI, which reacts with an epitope of IL-2R alpha and interferes with interaction of this chain and IL-2R beta, inhibits the IL-2-dependent proliferative effects mediated by analog F42A. Furthermore, by blocking interaction of IL-2R alpha and IL-2R beta with the antibody HIEI, a decrease in the affinity of radiolabeled analog F42A for IL-2R beta was seen. In our proposed model IL-2R alpha contributes several functions to IL-2-mediated signaling through the high-affinity IL-2R. These functions include concentration of IL-2 within the two-dimensional surface of the plasma membrane as well as alteration of the functional capacity of IL-2R beta, an effect that does not require prior binding of IL-2R to IL-2R alpha. The IL-2R alpha-mediated augmentation of IL-2R beta functions involves affinity conversion of IL-2R beta, increasing its affinity for IL-2, and may involve facilitation of Il-2-mediated signaling after binding of IL-2 to this IL-2R beta.

Amino Acid Sequence↗

PubMed1992

Characterization of the interleukin 2 receptors (IL-2R) expressed on human natural killer cells activated in vivo by IL-2: association of the p64 IL-2R gamma chain with the IL-2R beta chain in functional intermediate-affinity IL-2R.

Interleukin 2 (IL-2) receptors expressed on the surface of activated T cells and natural killer (NK) cells exhibit a variety of affinity states depending on their subunit composition. Low-affinity binding is associated with a 55-kDa alpha chain, intermediate-affinity binding with a 70-75-kD beta chain, and high-affinity binding with a bimolecular complex of the alpha and beta subunits. In a previous study of the IL-2 receptors expressed on NK cells obtained from cancer patients after in vivo IL-2 therapy, we documented a discrepancy between the level of beta chain and the level of intermediate-affinity IL-2 binding sites expressed on the cell surface. Based on this result, we postulated that formation of intermediate-affinity receptor sites required a component in addition to the beta chain, and that this component was present at limiting levels in the patient NK cells. In the present study we have examined the structure of the intermediate-affinity receptor complex using monoclonal antibodies that recognize the beta chain, but that do not interfere with its ability to bind IL-2. Evidence is presented establishing the physical association of a novel protein of 64 kD with the beta chain in intermediate-affinity IL-2 binding sites. This molecule, termed IL-2R gamma chain, coprecipitated with beta chains prepared from cells that had been incubated with IL-2, but was undetectable in immunoprecipitates prepared in the absence of IL-2. Examination of gamma chain expression in post-IL-2 therapy NK cells, where only low levels of intermediate-affinity IL-2 binding were detectable, revealed that the gamma chain was associated with, on average, only 10-12% of the beta chains expressed on such cells. This contrasted with approximately equal levels of beta and gamma chain expression on YT cells, a cell line that has both high levels of cell surface beta chain expression and high levels of IL-2 binding. Thus, the ratio of gamma chain to beta chain present in the immunoprecipitates roughly correlated with the proportion of beta chain involved in intermediate-affinity receptor sites. This result suggests that the 64-kD gamma chain is the component responsible for regulating the affinity of IL-2 association with the beta subunit. By further defining the structural components necessary for IL-2 receptor formation, these studies provide additional insight into mechanisms whereby lymphocytes might regulate their responsiveness to IL-2.

Animals↗

PubMed1992

Prolongation of graft survival in primate allograft transplantation by yttrium-90-labeled anti-Tac in conjunction with granulocyte colony-stimulating factor.

High-affinity IL-2 receptors are expressed by T cells activated in response to foreign histocompatibility antigens but not by normal resting T cells. To exploit this difference in IL-2R expression, anti-Tac, a murine monoclonal antibody specific for the IL-2R alpha subunit, was used to inhibit organ allograft rejection. To enhance its effector function, anti-Tac was armed by chelation with yttrium-90, a pure beta-emitting radionuclide. Animals received no immunosuppression (n = 5, group I, controls), unmodified anti-Tac (n = 5, 1 mg/kg q.o.d., group II), or 90Y-anti-Tac (n = 5, 1.6 mCi/kg divided into four doses, group III). The animals in group IV (n = 4) were treated identically to those in group III with the exception that 5 micrograms/kg/dose of granulocyte colony-stimulating factor was administered intramuscularly on the days when the yttrium-90 was given and on postoperative days 12 through 35 in order to reduce hematopoietic toxicity. Mean graft survival +/- S.E.M. for the control group was 8.2 +/- 0.5 days as compared with 13.8 +/- 2.1 days (P < 0.05) for those monkeys treated with unmodified anti-Tac. Graft survival was further prolonged in animals of group III that received 90Y-anti-Tac, with a mean graft survival of 45.0 +/- 11.8 days; however, three of the five monkeys retained viable grafts within this group but died secondary to bone marrow suppression. In comparison, the monkeys in group IV that were treated with G-CSF in conjunction with 90Y-anti-Tac had a mean graft survival of 49.2 +/- 2.9 days. In contrast to group III there were no deaths in the group (IV) receiving G-CSF. Furthermore, animals in group IV had a reduced magnitude and shortened duration of irradiation-induced neutropenia when compared with that observed in group III animals that did not receive G-CSF. Thus, treatment with 90Y-anti-Tac in conjunction with G-CSF may have potential applications in organ transplantation and the treatment of IL-2 receptor-expressing neoplastic diseases.

Animals↗

PubMed1992

Migration of recombinant IL-2-activated T and natural killer cells in the intercellular space of human H-2 glioma spheroids in vitro. A study on adhesion molecules involved.

The migration of rIL-2-activated T and NK cells into the intercellular space of glioma tissue was studied using multicellular spheroids grown from the human H-2 glioblastoma cell line as targets. Lymphocytes of all analyzed subtypes migrated into the spheroids, but CD56+ cells were particularly migratory. Lymphocytes and the H-2 tissue expressed adhesion molecule subunits for the following potential cell-cell or cell-matrix interactions: alpha 3 beta 1 (VLA-3) to fibronectin, laminin, and collagen; alpha 4 beta 1 (VLA-4) and alpha 5 beta 1 (VLA-5) to fibronectin; alpha 6 beta 1 (VLA-6) to laminin; alpha 4 beta 1 to VCAM-1; alpha L beta 2 (Leu-CAMa/LFA-1) to CD54 (ICAM-1); CD44 to fibronectin, collagen, laminin, hyaluronate; CD2 to CD58 (LFA-3); and CD56 (N-CAM) to CD56. In the H-2 tissue, CD54 and VCAM-1 were expressed as a gradient. The expression of CD54 was weak in the peripheral zone and the expression was stronger in the quiescent deeper zone, whereas the distribution of VCAM-1 showed an inversed pattern. The low expression of CD54 was up-regulated along the frontier of migrating lymphocytes. The migration was almost totally prevented by the anti-CD18 (beta 2) mAb IB4 and TS1/18, and also strongly inhibited by the anti-CD54 mAb LB-2. Instead, mAb known to inhibit the binding of beta 1 integrins to fibronectin were not significantly inhibitory. However, a combination of the GPEILDVPST and GRGDS peptides, which compete for the binding of alpha 4 beta 1 and alpha 5 beta 1 to fibronectin and may also affect other adhesion systems, partially prevented migration.

Amino Acid Sequence↗

PubMed1992

Regulation of lymphokine-activated killer cell induction by human recombinant IL-1 receptor antagonist. Obligate paracrine pathway of IL-1 during lymphokine-activated killer cell induction.

IL-2-stimulated human lymphocytes, referred to as lymphokine-activated killer (LAK) cells, can develop a broad range of lytic activity against fresh tumor cells and cultured tumor cell lines. IL-1, a pleiotropic cytokine shown to synergize with IL-2 on LAK induction, is endogenously synthesized and secreted by LAK cells. Immunoblot analysis demonstrated that IL-2-stimulated PBL produced the 31- to 34-kDa pro-molecules of IL-1 within 24 h and maintained their expression for at least 96 h. The role of secreted IL-1 has been examined using rIL-1R antagonist (IL-1ra). The addition of IL-1ra to LAK activation culture resulted in dose-dependent inhibited lytic activity, which was more apparent in LAK cells cultured with higher doses of IL-2. However, IL-1ra had no effect on proliferative responses elicited in LAK cells by IL-2. Moreover, when IL-1 binding was blocked by IL-1ra, the expression of the IL-2R p55 subunit was reduced compared with control LAK cells. The effect of IL-1 binding blockade on expression of other cytokine mRNA was further examined by polymerase chain reaction analysis, and, specifically, inhibition of both TNF-alpha and TNF-beta mRNA expression by IL-1ra was observed in PBL stimulated with IL-2. The reduced biologic activity of TNF in culture supernatants correlated well with the inhibition of mRNA expression. These findings suggest that autocrine/paracrine IL-1 is involved in the initial generation of LAK activity and, in particular, that TNF expression could be induced via an IL-1 autocrine pathway.

Base Sequence↗

PubMed1992

Functional reconstitution of the human interleukin-3 receptor.

The high-affinity receptors for human interleukin-3 (IL-3), GM-CSF, and IL-5 are composed of alpha and beta subunits. The alpha subunits are primary ligand binding proteins specific for each ligand, whereas the three human receptors share a common beta subunit (beta c). In contrast to humans mice have two closely related genes, AIC2A and AIC2B, which are homologous to human beta c. The AIC2A gene encodes a low-affinity murine IL-3 binding protein, and the AIC2B protein is the beta subunit shared between murine GM-CSF receptors (mGMR) and IL-5 receptors (mIL-5R). To examine the function of these receptor components, we established various stable transfectants of murine IL-2-dependent CTLL-2 cells. CTLL-2 transfectants expressing both the alpha and beta subunits of the human IL-3 receptor (hIL-3R) proliferated in response to physiologic concentrations of hIL-3. Coexpression of hIL-3R alpha with AIC2B but not with AIC2A in CTLL-2 cells conferred a growth response to hIL-3. Although CTLL-2 transfectants expressing hIL-3R alpha alone did not proliferate in the presence of hIL-3, hIL-3-responsive sublines were repeatedly isolated. These sublines expressed endogenous AIC2B but not AIC2A. These results indicate that human beta c is essential for hIL-3 signaling and that AIC2B is a murine equivalent of human beta c. We also showed that hIL-3 and hGM-CSF induced tyrosine phosphorylation of several proteins in CTLL transfectants, similar to those observed in human factor-dependent TF-1 cells stimulated with hIL-3 and hGM-CSF.

Animals↗

PubMed1992

Interferon-gamma increases the expression of the gene encoding the beta subunit of the granulocyte-macrophage colony-stimulating factor receptor.

Granulocyte-macrophage colony-stimulating factor (GM-CSF) activates a broad range of myeloid cells through binding to high-affinity receptors (GM-CSF-R) consisting of at least two distinct subunits, GM-CSF-R alpha and GM-CSF-R beta. The genes of these GM-CSF-R subunits have been identified recently, but little is known about the regulation of their expression. In this study, we investigated the expression of the GM-CSF-R subunit genes in normal human monocytes. Out of a panel of various cytokines and factors tested, only interferon-gamma (IFN-gamma) affected the expression of one of the GM-CSF-R subunit genes by increasing the GM-CSF-R beta mRNA expression threefold to sixfold with no effect on GM-CSF-R alpha. Maximal effects occurred 2 to 4 hours after stimulation with 500 to 5,000 U/mL IFN-gamma. Nuclear run-on assays and mRNA half-life studies showed that IFN-gamma modestly enhanced the transcription of the GM-CSF-R beta gene and stabilized the GM-CSF-R beta mRNA, with the latter mechanism predominant. Pretreatment of the monocytes with cycloheximide did not abrogate the increase of GM-CSF-R beta mRNA expression induced by IFN-gamma, indicating that de novo protein synthesis was not required for this activity. When monocytes were exposed to IFN-gamma for 6 to 24 hours, the number of GM-CSF-R per cell was increased 79% as compared with controls, whereas the receptor affinity remained unchanged. These data indicate that the GM-CSF-R expression in monocytes may be upregulated by IFN-gamma via an increased expression of the beta subunit gene, involving both transcriptional and post-transcriptional mechanisms.

Animals↗

PubMed1992

Regulation of IL-2 receptor subunit genes in human monocytes. Differential effects of IL-2 and IFN-gamma.

We investigated the effects of IFN-gamma and IL-2 on IL-2R alpha and beta mRNA expression in human monocytes. Low basal expression of IL-2R beta mRNA was detected in fresh monocytes. Stimulation of monocytes with IL-2 induced a significant increase of IL-2R beta mRNA, but did not induce IL-2R alpha mRNA. In contrast, stimulation of monocytes with IFN-gamma-induced IL-2R alpha mRNA, but did not modify IL-2R beta mRNA. Five U/ml of IFN-gamma induced IL-2R alpha mRNA and 2.2 nM of IL-2 induced IL-2R beta mRNA, both within 3 h. Nuclear run-on experiments demonstrated that the induction of IL-2R alpha mRNA by IFN-gamma is controlled, at least in part, at the transcriptional level. In contrast, the enhancement of IL-2R beta mRNA by IL-2 is controlled at a posttranscriptional level and is associated with an increase in the half-life of IL-2R beta mRNA. The results of studies on the cytotoxic activity and on the expression of c-fms mRNA of monocytes activated by the combination of IFN-gamma and IL-2 show that pretreatment with IFN-gamma renders monocytes more sensitive to activation by IL-2. These results demonstrate that the IL-2R alpha and IL-2R beta subunits are induced by different lymphokines through distinct mechanisms and that both receptor subunits can influence the response of monocytes to IL-2.

Antigens, CD↗

PubMed1992

Activation of NF-kappa B by interleukin 2 in human blood monocytes.

We report here that interleukin 2 (IL-2) acts on human blood monocytes by enhancing binding activity of the transcription factor NF-kappa B to its consensus sequence in the 5' regulatory enhancer region of the IL-2 receptor alpha chain (p55). Similarly, IL-2 activates NF-kappa B in the human monocytic cell line U 937, but not in resting human T-cells. This effect is detectable within 15 min and peaks 1 h after exposure to IL-2. Enhanced NF-kappa B binding activity is followed by functional activation in that inducibility of the IL-2 receptor alpha chain is mediated by enhanced NF-kappa B binding and that a heterologous promoter containing the NF-kappa B consensus sequence (-291 to -245) of the IL-2 receptor alpha chain gene is activated. In addition, IL-2 is capable of increasing transcript levels of the p50 gene coding for the p50 subunit of the NF-kappa B transcription factor, whereas mRNA levels of the p65 NF-kappa B gene remained unchanged.

Base Sequence↗

PubMed1992

[Structural similarity and signal transduction of cytokine receptors].

The advent of expression cloning has established a new group of membrane glycoproteins, called the cytokine receptor superfamily. Members of this family have common structural similarities in the extracellular region and no intrinsic tyrosine kinase activity is seen in the intracellular region. A combination of two distinct subunits (alpha and beta) in the family has been shown to generate high-affinity receptors in which the alpha subunits specify the binding and the beta subunits function as transducers that are presumably associated with tyrosine kinases. The high-affinity receptors for IL-3, IL-5 and GM-CSF all share the same beta subunit. The high-affinity receptor for IL-2 is however unique in that it requires a third component, notably, the gamma subunit, whose gene we have recently cloned.

Animals↗