The IL-2 receptor alpha-chain alters the binding of IL-2 to the beta-chain

The binding of IL-2 to its high affinity receptor results in the formation of the ternary complex consisting of IL-2, alpha-chain (p55, Tac) and beta-chain (p75). We studied the role of alpha-chain in IL-2 binding to the high affinity receptor using IL-2 analog Lys20 which was made by the substitution of Lys for Asp20 of wild-type rIL-2. Lys20 bound to MT-1 cells solely expressing alpha-chain at low affinity, but did not bind to YT-2C2 cells which solely expressed beta-chain. However, direct binding of radiolabeled Lys20 to ED515-D cells, an HTLV-I-infected and IL-2-dependent T cell line, revealed both high affinity and low affinity binding although the Kd value of high affinity binding was 50 to 100 times higher than that of the high affinity binding of wild-type rIL-2. High affinity binding of Lys20 was completely blocked by 2R-B mAb recognizing IL-2R beta-chain. Anti-Tac mAb recognizing IL-2R alpha-chain abolished all of the specific Lys20 bindings. In contrast to the replacement of cell bound 2R-B mAb with wild-type rIL-2 at 37 degrees C, the addition of an excess of Lys20 did not cause the detachment of cell-bound radiolabeled or FITC-labeled 2R-B mAb. Consistent with the results of binding studies, Lys20 induced the proliferation of ED515-D cells, but not large granular lymphocyte leukemic cells. The growth of ED-515D cells was completely suppressed by either anti-Tac mAb or 2R-B mAb. These results strongly suggest that coexpression of the IL-2R alpha- and beta-chains alters the binding affinity of Lys20 and that the interaction between IL-2 and the alpha-chain is a key event in the formation of the IL-2/IL-2R ternary complex.     

Reconstitution of a functional IL-2 receptor by the beta-chain cDNA. A newly acquired receptor transduces negative signal

The high-affinity IL-2R results from the noncovalent association between at least two subunits; alpha (p55) and beta (p70), both of which are capable of binding IL-2 with a low and intermediate affinity, respectively. Although the alpha-chain itself has been shown to be nonfunctional, suggestions have been made that the beta-chain mediates an IL-2 signal. To directly study the role of the beta-chain in the signal transduction, we transfected with the cDNA encoding the IL-2R beta-chain a human T lymphotropic virus-I-transformed T cell line, MT-1 originally expressing low-affinity alpha-chain alone, and established a stable transformant (designated MT-beta 7) which expressed both alpha- and beta-chains simultaneously. We showed 1) MT-beta 7 manifested the high-affinity IL-2 binding, which was completely disrupted by the anti-beta chain mAb (Mik-beta 1), 2) the 125I-IL-2 crosslinking patterns of MT-beta 7 were indistinguishable from those of cells expressing the native high-affinity IL-2R, 3) MT-beta 7, but not parental MT-1, internalized the bound IL-2 and responded to IL-2 with a negative signal, i.e., inhibition of the de novo DNA synthesis. These results clearly demonstrate that the beta-chain not only participates in forming the high-affinity IL-2R with the alpha-chain but also is directly involved in the IL-2 signal transduction.     

ATL-derived factor (ADF), an IL-2 receptor/Tac inducer homologous to thioredoxin; possible involvement of dithiol-reduction in the IL-2 receptor induction.

HTLV-I transformed T cells not only express a large number of interleukin-2 receptors [IL-2R/p55(Tac)], but also produce a factor named ATL-derived factor (ADF) that augments the expression of IL-2R/p55(Tac). Based on a partial N-terminal amino acid sequence, complementary DNA (cDNA) clones for human and mouse ADF were isolated and sequenced. Recombinant ADF produced by COS-7 monkey kidney cells showed IL-2R/Tac inducing activity on YT cells, which are sensitive for ADF. ADF mRNA was strongly expressed in HTLV-I(+) T cells lines, but not in inactivated cells (THP-1, unstimulated PBMC). Furthermore, in normal human peripheral blood mononuclear cells, the expression of ADF mRNA was enhanced by mitogens or phorbol myristate acetate, suggesting a possible involvement of ADF in the lymphocyte activation. Homology analysis revealed an unexpected relationship between ADF and dithiol-reducing enzyme, thioredoxin, involved in many important biological reactions such as the conversion of ribonucleotides into deoxyribonucleotides, or the stabilization of glucocorticoid receptors. The biological significance of the generation of a redox potential in lymphocyte activation, and the possible involvement of dithiol reduction in the induction of IL-2R/Tac are discussed.     

Presence of a p70 IL-2-binding peptide on leukemic cells from various hemopoietic lineages

Recent studies have shown that IL-2R are composed of at least two polypeptide chains of 55 kDa (Tac or alpha-chain) and 70 to 75 kDa (p70 or beta-chain). The association of both chains forms high affinity IL-2R, whereas each chain alone binds IL-2 with a low (alpha-chain) or intermediate (beta-chain) affinity. So far, the p70 peptide has been found, in the absence of the Tac peptide, on the surface of lymphoid cells of T, B, or NK lineage. In this study, we investigated whether leukemic cells of various hemopoietic lineages expressed the p70 IL-2-binding protein. We found that both fresh leukemic cells obtained from patients, and cells from established leukemic lines of T cells, B cell, and myeloid origin constitutively expressed a p70 IL-2-binding protein on their surface, as detected by affinity cross-linking of radioiodinated IL-2. IL-2 binding and cross-linking to these cells was completely inhibited in the presence of an excess unlabeled rIL-2, but not with an anti-Tac mAb. Binding experiments on pre-B and myeloid cell lines revealed intermediate affinity IL-2R, whereas both high and intermediate affinity IL-2R were detected in T leukemic cells. The intermediate affinity binding of 125I-rIL-2 to the leukemic cell lines MOLT4 and Reh6 was inhibited by the TU27 mAb, which recognized the p75 chain of IL-2R. Moreover, the TU27 mAb could stain the K562, KM3, and MOLT4 (weakly) cell lines by indirect immunofluorescence. A high dose of rIL-2 (400 U/ml) enhanced the proliferation of cells from one out of three patients with acute myeloblastic leukemia, but it did not induce differentiation of the cells in any of three cases. Thus the finding of p70 IL-2-binding molecules on immature lymphoid and nonlymphoid hemopoietic cells should disclose new biologic functions for IL-2.     

IL-7 regulation of cytotoxic lymphocytes: pore-forming protein gene expression, interferon-gamma production, and cytotoxicity of human peripheral blood lymphocytes subsets.

The effects of IL-7 on the generation of cytolytic human peripheral blood lymphocytes (PBL) were investigated. Induction of T-cell pore-forming protein (PFP) mRNA and cytotoxic potential by IL-7 was both slow and minor compared with that observed in IL-2-cultured T cells. IL-7 and suboptimal doses of IL-2 (10 U/ml) were found to costimulate PFP mRNA expression and cytotoxic potential in T cells. Clearly, however, both IL-7 and IL-2/IL-7 induced the PFP gene expression and cytotoxic potential of CD8+ T cells and not CD4+ T cells. In addition, neither monoclonal antibodies (mAb) to the p55 or p75 IL-2-receptor subunits had any effect upon IL-7 induction of CD8+ T-cell cytotoxicity, indicating that IL-7 induction of cytotoxic CD8+ T cells was IL-2 independent. IL-7 induction of CD3- large granular lymphocyte (LGL) and PB gamma delta T-cell cytotoxicity was also delayed and reduced compared with that effected by IL-2. IL-7 (10 or 1000 U/ml, 72 hr) enhanced the NK and LAK cytotoxic of LGL and PB gamma delta T cells. By contrast IL-7 or IL-2 augmented the redirected cytotoxic potential of PB gamma delta T cells, but not that of LGL, and neither lymphokine had any effect on constitutive PFP mRNA expression in either lymphocyte subset. In addition, IL-7 induction of LGL IFN-gamma production was weak and delayed compared with that effected by IL-2 and neither IL-2 nor IL-7 stimulated IFN-gamma production in PB gamma delta T cells. Therefore, overall the effects of IL-2 and IL-7 on various cytotoxic human PBL were qualitatively similar, but quantitatively and kinetically different.     

Protein phosphorylation mediated by IL-2/IL-2 receptor beta-chain interaction

We previously found that IL-2 rapidly induced protein phosphorylation of a 67-kDa (pp67) and four 63-kDa (pp63s) cellular proteins in various T cells. Here, we show that the IL-2-stimulated phosphorylation is mediated by the IL-2R beta-chain composed of the high affinity IL-2R, and induced by activation of Ca2+/phospholipid-dependent protein kinase C (PKC). The IL-2-stimulated phosphorylation was always observed in various T cell lines bearing high affinity IL-2R, but never observed in cells which express only low affinity IL-2R consisted of alpha-chain alone. When the expression of high affinity IL-2R was modified by anti-IL-2R mAb for reducing the affinity to 8- to 10-fold lower without affecting the sites of IL-2R, the effective dose of IL-2 on phosphorylation of pp67 increased 8 to 10 times. When cells were treated with pronase, approximately 95% sites of low affinity IL-2R were selectively decreased, but the IL-2 dose dependency for pp67 phosphorylation was little affected. These data exactly suggest that protein phosphorylation in response to IL-2 such as pp67 and pp63s, is mediated by high affinity but not low affinity IL-2R. Furthermore, the IL-2-stimulated phosphorylation of these proteins was also observed in MLA 144 cells which express only low affinity IL-2R consisting of beta-chain alone. In addition, various phorbol esters and tumor promoters, which activate PKC, were also demonstrated to induce the phosphorylation of a pp67 and pp63s in these T cell lines. Therefore, the present study suggests that IL-2/IL-2R beta-chain interaction triggers the phosphorylation of pp67 and pp63s, where the PKC may have an important role.     

A major 50-kDa human B-cell growth factor-II induces both Tac antigen expression and proliferation by several types of lymphocytes

Many cytokines have been documented to have a multiplicity of biological effects by acting on a variety of cells. In order to determine whether human BCGF-II acts on any cells in addition to normal B cells, the effect of human BCGF-II on murine thymocytes, human peripheral blood T cells, a human natural killer-like cell line, YT, and Epstein-Barr virus (EBV)-transformed B-cell lines was further examined. BCGF-II augmented incorporation of [3H]thymidine by murine thymocytes in combination with suboptimal doses (0.5 microgram/ml) of concanavalin A (Con A) but not at lower doses (0.1 microgram/ml) of Con A, a concentration usually used for interleukin 1 (IL-1) assays. BCGF-II could not induce proliferation or Tac antigen (Ag) expression on normal peripheral blood T cells stimulated with OKT3 antibody. Both proliferation and Tac Ag expression on YT cells were also augmented by BCGF-II. BCGF-II induced both high- and low-affinity IL-2 receptor (IL-2R) on YT cells as determined by 125I-IL-2-binding assay. Two of seven EBV-transformed B-cell lines tested (ORSON and AUM cells) in response to BCGF-II exhibited augmentation of proliferation and cell surface Tac Ag expression. BCGF-II in the presence of low doses (0.1 microgram/ml) of phorbol myristate acetate (PMA) also induced Tac Ag mRNA (3.5 and 1.5 kb) in these B-cell lines. The IL-2R induced on these B-cell lines, however, consisted mostly of low-affinity receptors. Both Tac Ag and its mRNA in these B-cell lines were not induced by Forskolin but by PMA, suggesting that this induction may involve protein kinase C. The present study shows that human BCGF-II can stimulate YT cells, murine thymocytes, and some EBV-transformed B-cell lines but not peripheral blood T cells. Consequently, BCGF-II can induce the growth and differentiation of a number of cell types in addition to normal B cells.     

Selective availability of IL-2 is a major determinant controlling the production of CD4+CD25+Foxp3+ T regulatory cells

The development and maintenance of T regulatory (Treg) cells critically depend on IL-2. This requirement for IL-2 might be due to specificity associated with IL-2R signal transduction or because IL-2 was uniquely present in the niche in which Treg cells reside. To address this issue, we examined the capacity of IL-7R-dependent signaling to support Treg cell production and prevent autoimmunity in IL-2Rbeta(-/-) mice. Expression of transgenic wild-type IL-7R or a chimeric receptor that consisted of the extracytoplasmic domain of the IL-7R alpha-chain and the cytoplasmic domain of IL-2R beta-chain in IL-2Rbeta(-/-) mice did not prevent autoimmunity. Importantly, expression of a chimeric receptor that consisted of the extracytoplasmic domain of the IL-2R beta-chain and the cytoplasmic domain of IL-7R alpha-chain in IL-2Rbeta(-/-) mice led to Treg cells production in the thymus and periphery and prevented autoimmunity. Signaling through the IL-2R or chimeric IL-2Rbeta/IL-7Ralpha in vivo or the culture of thymocytes from IL-2Rbeta(-/-) mice with IL-7 led to up-regulation of Foxp3 and CD25 on Treg cells. These findings indicate that IL-7R signal transduction is competent to promote Treg cell production, but this signaling requires triggering through IL-2 by binding to the extracytoplasmic portion of the IL-2R via this chimeric receptor. Thus, a major factor controlling the nonredundant activity of the IL-2R is selective compartmentalization of IL-2-producing cells with Treg cells in vivo.     

Natural, proteolytic release of a soluble form of human IL-15 receptor alpha-chain that behaves as a specific, high affinity IL-15 antagonist

IL-15 and IL-2 are two structurally and functionally related cytokines whose high affinity receptors share the IL-2R beta-chain and gamma-chain in association with IL-15R alpha-chain (IL-15R alpha) or IL-2R alpha-chain, respectively. Whereas IL-2 action seems restricted to the adaptative T cells, IL-15 appears to be crucial for the function of the innate immune responses, and the pleiotropic expression of IL-15 and IL-15R alpha hints at a much broader role for the IL-15 system in multiple cell types and tissues. In this report, using a highly sensitive radioimmunoassay, we show the existence of a soluble form of human IL-15R alpha (sIL-15R alpha) that arises from proteolytic shedding of the membrane-anchored receptor. This soluble receptor is spontaneously released from IL-15R alpha-expressing human cell lines as well as from IL-15R alpha transfected COS-7 cells. This release is strongly induced by PMA and ionomycin, and to a lesser extent by IL-1 beta and TNF-alpha. The size of sIL-15R alpha (42 kDa), together with the analysis of deletion mutants in the ectodomain of IL-15R alpha, indicates the existence of cleavage sites that are proximal to the plasma membrane. Whereas shedding induced by PMA was abrogated by the synthetic matrix metalloproteinases inhibitor GM6001, the spontaneous shedding was not, indicating the occurrence of at least two distinct proteolytic mechanisms. The sIL-15R alpha displayed high affinity for IL-15 and behaved as a potent and specific inhibitor of IL-15 binding to the membrane receptor, and of IL-15-induced cell proliferation (IC(50) in the range from 3 to 20 pM). These results suggest that IL-15R alpha shedding may play important immunoregulatory functions.     

IL-7 receptor expression levels do not identify CD8+ memory T lymphocyte precursors following peptide immunization

Identification of the mechanisms underlying the survival of effector T cells and their differentiation into memory T lymphocytes are critically important to understanding memory development. Because cytokines regulate proliferation, differentiation, and survival of T lymphocytes, we hypothesized that cytokine signaling dictates the fate of effector T cells. To follow cytokine receptor expression during T cell responses, we transferred murine TCR transgenic T cells into naive recipients followed by immunization with peptide emulsified in adjuvant or pulsed on dendritic cells. Our findings did not correlate IL-7R alpha-chain and IL-2R beta-chain expression on effector CD8+ cells with the generation of memory T lymphocytes. However, we could correlate the extent of IL-7R alpha expression down-regulation on effector T cells with the level of inflammation generated by the immunization. Furthermore, our findings showed that the maintenance of a high level of IL-7R expression by effector T cells at the peak of the response does not preclude their death. This suggests that maintenance of IL-7R expression is not sufficient to prevent T cell contraction. Thus, our results indicate that expression of the IL-7R is not always a good marker for identifying precursors of memory T cells among effectors and that selective expression of the IL-7R by effector T cells should not be used to predict the success of vaccination.     

IL-2 and IL-6 synergize to augment the pore-forming protein gene expression and cytotoxic potential of human peripheral blood T cells

Our previous studies have demonstrated that high dose IL-2 (1000 U/ml) alone can induce human peripheral blood T cell pore-forming protein (PFP) mRNA expression and cytotoxic potential. We now report that the levels of IL-2 needed to induce these effects in T cells can be significantly reduced in the presence of IL-6. IL-6 and suboptimal doses of IL-2 (10 U/ml) were found to costimulate PFP mRNA expression and cytotoxic potential in resting human peripheral blood T cells, whereas IL-6 or low dose IL-2 alone had no effect. The induction of T cell PFP mRNA by IL-2/IL-6 was extremely rapid and increases in both PFP mRNA expression and cytotoxic potential were IL-6 dose dependent. The costimulatory effect of IL-6 did not appear to involve the IL-2/IL-2R pathway in as much as IL-6 did not induce IL-2 production or detectably increase IL-2R surface expression in T cells. These findings, in addition to the rapid induction of PFP mRNA by IL-2/IL-6, suggested that IL-6 can directly and independently provide an additional signal to augment the differentiation of CTL. In contrast to the results observed in T cells, IL-6 and IL-2 could enhance CD3- large granular lymphocyte (LGL) NK activity, but IL-6 either alone or in combination with IL-2 had no effect on constitutive PFP mRNA expression in resting LGL. These data further confirm that different mechanisms may be responsible for lymphokine activation of CTL and LGL in human peripheral blood. In particular it appears that IL-6 acts as a costimulatory signal with IL-2 in generating CTL and that IL-6 functions in part by acting in synergy with IL-2 to induce PFP, a major lytic protein involved in lymphocyte cytotoxicity.     

Comparison of the effect of IL-2 and IL-6 on the lytic activity of purified human peripheral blood large granular lymphocytes

The effects of IL-6 and IL-2 on highly purified, human peripheral blood large granular lymphocytes (LGL) were investigated and compared. IL-6 enhanced LGL NK activity in a dose-dependent manner against K562, however IL-2 was a more potent stimulus of LGL NK function. Neither IL-2 nor IL-6 increased LGL cytotoxic potential in a parallel estimation of heteroconjugated antibody (anti-CD16 x anti-nitrophenyl mAb)-dependent cytotoxicity against nitrophenyl-modified YAC. Unlike IL-2, IL-6 did not significantly induce LGL lymphokine-activated killer activity, LGL proliferation, or LGL lymphokine production. In particular, IL-6 did not stimulate detectable LGL IL-2 production or IL-2R modulation, and mAb to the p75 IL-2R had no effect on IL-6 induction of LGL NK activity. Therefore, in the absence of T cells, IL-6 provided an IL-2-independent signal to LGL that resulted in augmentation of their NK activity without stimulating their proliferation or other LGL functions.     

Immature, double negative (CD4-,CD8-) rat thymocytes do not express IL-2 receptors

IL-2R alpha-chain is expressed on a subset of mouse CD4- and CD8-, double negative (DN) thymocytes. This expression of IL-2R alpha-chain on some DN thymocytes in the mouse has led to the proposal that IL-2 might serve as a principal growth and/or differentiation factor for immature thymocytes. However, previous histologic observations have indicated that IL-2R alpha-chain is not expressed on the subcapsular thymic blasts (an area rich in DN cells) in either huma or rat thymus, whereas all three species display IL-2R expression on a few cells in the thymic medulla. Therefore, we characterized rat DN thymocytes to determine whether they contained an IL-2R+ population. The results show that rat thymic DN cells share several characteristics with mouse DN cells. However, most of the rat strains do not express the IL-2R on DN cells as shown either by immunofluorescence or by IL-2 binding and receptor cross-linking. Thus, the rare medullary IL-2R+ cells were not found in the DN cells. Only in the exceptional F344 rat strain is the IL-2R alpha-chain expressed on a major proportion of thymocytes, including both DN cells and small cortical-type thymocytes. Furthermore, rat DN cells do not contain detectable IL-2 mRNA or cytoplasmic IL-2 activity, thus supporting the conclusion that it is unlikely that IL-2 and IL-2R serve to maintain the proliferation of rat DN thymocytes in vivo. The possible significance of in vivo expression of IL-2R alpha-chain on immature thymocytes in the mouse and in a single rat strain is discussed.     

IL-2 receptor(p55)/Tac-inducing factor. Purification and characterization of adult T cell leukemia-derived factor

Many human T cell lymphotropic virus-I (HTLV-I) transformed T cells from adult T cell leukemia (ATL) patients continuously produce a humoral factor called ATL-derived factor (ADF) which induces IL-2R/Tac expression on T and NK cells. Using gel filtration, procion red Sepharose, DEAE, and reverse phase chromatography, we have purified ADF protein to homogeneity from 15 liters of serum-free culture supernatant of an HTLV-I(+) T cell line ATL-2. Purified ADF protein had the m.w. of 14,000 by SDS-PAGE and gel filtration, and its isoelectric point is around 5.0. ADF did not react with heteroantibodies against IL-1 alpha and IL-1 beta, which have also IL-2R/Tac-inducing activity on suitable target cells. Partial N-terminal amino acid sequence of ADF is different from other cytokines such as, IFN, BSF-2, and various IL whose cDNA has been cloned. Western blot analysis using rabbit antibodies against N-terminal 10mer synthetic peptide of ADF showed that IL-1 alpha and ADF are different proteins. ADF had its IL-2R/Tac-inducing activity not only on human NK-like cell line YT, but also on HTLV-I(+) T cells, such as ED. In contrast, macrophage-derived IL-1 lacked IL-2R/Tac-inducing activity on ED cells despite their IL-2R/Tac induction on YT, indicating that ADF and IL-1 have their effect via different receptors.     

Biosynthesis and posttranslational regulation of human IL-12

IL-12 is a heterodimeric proinflammatory cytokine consisting of a light alpha-chain, formerly defined as p35, disulfide-linked to a heavier beta-chain, formerly defined as p40. The beta-chain is also produced in large excess in a free form, and disulfide-linked beta-chain homodimers with anti-inflammatory effects are produced in the mouse. We analyzed the biosynthesis and glycosylation of IL-12 in human monocytes, and in a cell line stably transfected with IL-12 alpha and beta genes (P5-0.1). The IL-12 heterodimer and free beta-chain were immunoprecipitated from supernatants and cell lysates of metabolically labeled cells and resolved in SDS-PAGE. Whereas the beta-chain showed similar pI pattern whether in the free form or associated in the heterodimer, either in the secreted or intracellular form, the alpha-chain in the secreted heterodimer was much more acidic than that present in the intracellular heterodimer. Deglycosylation experiments with neuraminidase and Endo-F combined with two-dimensional PAGE of single bands of the intracellular vs extracellular IL-12 heterodimer revealed that the alpha-chain was extensively modified with sialic acid adducts to N-linked oligosaccharides before secretion. N-glycosylation inhibition by tunicamycin (TM) did not alter free beta-chain secretion, while preventing the IL-12 heterodimer assembling and secretion. Pulse-chase experiments indicated that IL-12 persists intracellularly for a long period as an immature heterodimer, and that glycosylation is the regulatory step that determines its secretion. beta-chain disulfide-linked homodimers were observed in TM-treated P5-0.1 cells, but in neither TM-treated nor untreated monocytes.     

Synergistic induction of interleukin 2 receptor (TAC) expression on YT cells by simultaneous activation of distinct signal transduction pathways

The human NK-like leukemic cell line YT was used to study interleukin 2 receptor (IL-2R; Tac) expression induced by activators of distinct signal transduction pathways. Tac expression was induced by active phorbol esters (12-O-tetradecanoylphorbol 13-acetate [TPA] and 4 beta-phorbol 12,13-didecanoate), which directly activate protein kinase C (PKC), as well as forskolin (FK), a stimulator of adenylate cyclase. A synergistic effect on Tac expression was obtained by simultaneous stimulation with optimal concentrations of phorbol esters and FK. Inactive phorbol esters (4 beta-phorbol, 4 alpha-phorbol 12,13-didecanoate) and the inactive analog of FK (1,9-dideoxyforskolin) had no effect on Tac expression. The active phorbol esters synergized also with interleukin 1 (IL-1) and tumor necrosis factor alpha (TNF alpha) in Tac expression. Staurosporine, a potent inhibitor of PKC in vitro, inhibited Tac expression marginally in YT cells stimulated with FK, and enhanced Tac expression in cultures treated with TPA, TNF alpha, or IL-1. Based on the assumption that synergistic effects are observed when two agonists use different signaling pathways, these findings provide evidence that IL-1, TNF, and TPA use different pathways/regulatory elements to regulate Tac expression on the cell surface. Synergistic upregulation of Tac expression by simultaneous activation of distinct pathways may be an important mechanism to modulate the immune response.     

Induction of interleukin 2 receptor (TAC) by tumor necrosis factor in YT cells

The effect of human recombinant tumor necrosis factor alpha (TNF-alpha) and interleukin 1 beta (IL-1 beta) on interleukin 2 receptor (IL-2R) expression on YT cells was examined. IL-2R expression was assessed by flow cytometric analysis with a monoclonal antibody to IL-2R (anti-TAC). TNF-alpha, like IL-1 beta, induced increased levels of IL-2R expression on YT cells with similar kinetics of induction. Maximum induction occurred at 20 to 30 hr. On a molar basis. TNF was less active than IL-1 beta. RNA isolated from TNF-alpha- or IL-1 beta-treated YT cells contained increased levels of IL-2R-specific mRNA as indicated by slot blot analysis by using an IL-2R-specific mRNA probe. Kinetic and IL-1 beta mRNA expression studies indicated that the TNF effect was a direct one. Because IL-2R expression is known to be associated with lymphocyte activation, the present results suggest that TNF-alpha may play a role in the regulation of immune responses.