Biomedical subjects

K Sugamura

Publications and source records attributed to K Sugamura.

At least 127 records · Page 7Linked to original sources

PubMed1994

Expression of the IL-2 receptor gamma chain on various populations in human peripheral blood.

We have established two rat mAbs, TUGh4 and TUGh5, specific for the human gamma chain of the IL-2 receptor (IL-2R), which is known to be shared among receptors for IL-2, IL-4 and IL-7. The antibodies bound to cell lines transfected with the human gamma chain gene but not to their parental cell lines, and precipitated 65-70 and 80-90 kDa cell surface molecules from lysates of human T cells surface-labeled with Na125I and chemically cross-linked with [125I]IL-2 respectively. Flow cytometry with TUGh4 and TUGh5 detected the gamma chain in a wide variety of peripheral blood cell populations including CD4+ T cells, CD8+ T cells, CD20+ B cells, CD56+ natural killer cells, CD14+ monocytes and granulocytes, contrasting with expression of the alpha and beta chains of IL-2R.

Antibodies, Monoclonal↗

PubMed1994

Implication of the common gamma chain of the IL-7 receptor in intrathymic development of pro-T cells.

The effects of IL-7 on the growth and differentiation of thymocytes were analyzed using murine fetal thymus organ cultures (FTOC) in the presence of mAbs specific for the conventional IL-7 receptor (IL-7R) and for the common gamma (gamma c) chain. In FTOC, the development of CD4-CD8- double-negative thymocytes to CD4+CD8+ double-positive (DP) and CD4+ or CD8+ single-positive (SP) cells was not completely blocked by adding these mAbs, although cell growth was reduced by the treatment. To define a developing stage sensitive to the mAbs, most immature thymocytes, Pgp-1+ c-kit+ cells, were cultured in 2-deoxyguanosine treated fetal thymus. In the presence of both mAbs in the culture, neither DP nor SP thymocytes developed whereas either of the mAbs partially blocked their development. These results indicate that the gamma c chain is involved in early T cell development as an indispensable subunit of the functional IL-7 receptor complex.

Animals↗

PubMed1994

Immunological characterization of human decidual mononuclear cells: natural killer activity, response to interleukin-2 and distribution of interleukin-2 receptor subunits.

We investigated the immunological character of mononuclear cells obtained from the decidua in the first trimester of normal pregnancy. These cells showed in vitro cytotoxicity against NK cell targets, although with lower activity than that of the peripheral blood mononuclear cells. Response to IL-2 was dose-dependent. Since decidual CD56-positive cells express two IL-2 receptor subunits, p55 and p75, it is concluded that these cells have high-affinity receptors to IL-2. Peripheral blood CD56-positive cells, which express p75 alone, might have intermediate-affinity IL-2 receptors. These results indicate that the decidual mononuclear cells have a function resembling that of the peripheral blood NK cells in vitro, moreover, that even low levels of IL-2 can affect the character of the decidual mononuclear cells through the high-affinity IL-2 receptor. It is considered that the NK activity of decidual mononuclear cells is suppressed in conditions of low IL-2 levels to permit the maintenance of pregnancy, but can be rapidly elicited by intrauterine infections or abortion, both of which elicit the secretion of IL-2.

Cytotoxicity, Immunologic↗

PubMed1994

The frequency of human parvovirus B19 infection in nonimmune hydrops fetalis.

Intrauterine infection of human parvovirus B19 has recently been identified as an etiology for nonimmune hydrops fetalis (NIHF) and fetal death. To examine the frequency of B19 infection in cases of NIHF, forty two cases of NIHF of unknown cause were tested for the presence of B19 antibodies by enzyme-linked immunosorbent assay (ELISA) and B19 DNA by polymerase chain reaction (PCR). Three maternal sera were positive for B19 IgM antibody and one fetal serum was positive for B19 DNA. Overall, four (10%) of the 42 cases were positive for B19 infection. All of the four cases positive for B19 infection were found during or just before an outbreak of erythema infectiosum and the incidence was calculated at 36% if only cases found during the outbreak were collected. In all four cases of B19 infection, NIHF was diagnosed at 20 to 23 weeks of gestation, which suggested that B19 infection might be a particular threat to the fetus during this stage of gestation. In conclusion, B19 infection may contribute to 10% of cause unknown NIHF and the incidence may be higher if cases during outbreaks are exclusively collected.

Antibodies, Viral↗

PubMed1994

[Structure and function of IL-2 receptor subunits].

IL-2 regulates growth and differentiation of various types of cells in the immune system via its interaction with IL-2 receptor (IL-2R). The high and intermediate-affinity IL-2Rs, which consist of the alpha beta gamma heterotrimer complex and the beta gamma heterodimer complex, respectively, harbor the function of the intracellular signal transduction, indicating that the beta and gamma chains are indispensable for the signal transduction but not the alpha chain. The reconstitution studies of IL-2Rs with alpha, beta and gamma chain genes demonstrated that each subunit has potential for altering the affinity of the receptor, and the cytoplasmic domains of the beta and gamma chains participate in signal transduction in terms of cell growth, activation of alpha tyrosine kinase and enhancement of c-myc, c-fos and c-jun transcription. The region containing the SH2 homologous sequence of the gamma chain should have a critical function for signal transduction. On the other hand, common subunits are known to be shared among receptors for IL-3, IL-5 and GM-CSF, and receptors for IL-6, LIF, OSM and LIF. We have demonstrated that the monoclonal antibody specific for the IL-2R gamma chain completely inhibited not only IL-2-dependent cell growth but also IL-4-dependent, IL-7-dependent, and IL-9-dependent cell growth, suggesting that the gamma chain is possibly shared among receptors for IL-2, IL-4, IL-7 and IL-9. Impairment of the gamma chain function is considered to be closely related to human XSCID characterized by profound T cell defect.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1993

Sharing of the interleukin-2 (IL-2) receptor gamma chain between receptors for IL-2 and IL-4.

The gamma chain of the interleukin-2 (IL-2) receptor is an indispensable subunit for IL-2 binding and intracellular signal transduction. A monoclonal antibody to the gamma chain, TUGm2, inhibited IL-2 binding to the functional IL-2 receptors and also inhibited IL-4-induced cell growth and the high-affinity binding of IL-4 to the CTLL-2 mouse T cell line. Another monoclonal antibody, TUGm3, which reacted with the gamma chain cross-linked with IL-2, also immunoprecipitated the gamma chain when cross-linked with IL-4. These results suggest that the IL-2 receptor gamma chain is functionally involved in the IL-4 receptor complex.

Animals↗

PubMed1993

A novel human tyrosine kinase gene inducible in T cells by interleukin 2.

We have cloned a novel human protein tyrosine kinase gene specific to T cells by the polymerase chain reaction method. This gene encodes a 620 amino-acid polypeptide including a catalytic domain for tyrosine kinase, an SH2 domain and an SH3 domain, seemingly belonging to the src family. However, characteristics of a long unique N-terminal stretch and lack of a myristylation site at the N-terminus and of a kinase regulatory tyrosine residue in the C-terminus classify this molecule into a new subfamily comprising recently cloned mouse tec, itk/tsk and human atk/bpk genes. This gene was transcriptionally induced in normal T cells by interleukin 2 stimulation. These results suggest the crucial roles of this gene in T cell proliferation and differentiation.

Amino Acid Sequence↗

PubMed1993

Cloning of the mouse interleukin 2 receptor gamma chain: demonstration of functional differences between the mouse and human receptors.

We isolated a cDNA clone for the gamma chain of the mouse interleukin 2 receptor. Introduction of the mouse gamma chain cDNA clone into a mouse fibroblast cell line, L929, expressing the mouse alpha beta heterodimer IL-2 receptor converted pseudo-high affinity of the IL-2 receptor into functional high, resulting in internalization of IL-2 and induction of the c-myc, c-fos and c-jun genes. The mouse beta gamma heterodimer, however, failed to bind IL-2 unlike the human beta gamma heterodimer intermediate-affinity receptor. These results indicate that the mouse functional IL-2 receptor is a complex comprising three distinct subunits, alpha, beta and gamma chains, but the beta gamma heterodimer is not functional and different from the human heterodimer.

Amino Acid Sequence↗

PubMed1993

Reconstitution of functional interleukin 2 receptor complexes on fibroblastoid cells: involvement of the cytoplasmic domain of the gamma chain in two distinct signaling pathways.

We have previously shown that the interleukin 2 (IL-2) receptor gamma chain is a member of the cytokine receptor superfamily and is indispensable for the formation of receptor complexes with high and intermediate affinities for IL-2. The present study demonstrates that the alpha beta gamma heterotrimer and beta gamma heterodimer complexes of IL-2 receptor reconstituted on murine fibroblast L929 cells can transduce IL-2-mediated signals for activation of tyrosine kinase and for induction of c-myc, c-fos, and c-jun expression. A mutant of the gamma chain lacking the C-terminal 68 amino acids in its cytoplasmic region showed a loss of such signal-transducing ability when incorporated into the IL-2 receptor complexes but brought no effect on IL-2 binding and IL-2 internalization. Another mutant, with a C-terminal deletion of 30 amino acids, retained the ability to activate a tyrosine kinase and to induce c-myc expression but lost the ability to induce c-fos and c-jun expression. These results suggest that at least two distinct signals, one for c-myc induction, which parallels tyrosine kinase activation, and the other for c-fos and c-jun induction, can be transduced from the IL-2 receptor complexes reconstituted on fibroblastoid cells.

Amino Acid Sequence↗

PubMed1993

Kinetic study of interleukin-2 binding on the reconstituted interleukin-2 receptor complexes including the human gamma chain.

To gain an insight into roles of the interleukin-2 (IL-2) receptor gamma chain in IL-2 binding mechanisms, we examined association and dissociation rate constants for IL-2 binding with a series of fibroblastoid L929 cell lines expressing IL-2 receptor complexes reconstituted by transfection with the alpha, beta and gamma genes. The association rate constant (k) with the alpha beta gamma complex on L cells was fourfold larger than that with the alpha beta complex on L cells, and the dissociation rate constant (k') was one fifth of that with the alpha beta complex. These results indicate that the gamma chain is involved in both mechanisms by which IL-2 associates with and dissociates from receptors, resulting in the generation of the high-affinity IL-2 receptor along with the alpha and beta chains. During the course of this study, we found that the IL-2 dissociation from alpha beta gamma complex on lymphoid cells was a lot slower than that from the alpha beta gamma complex on fibroblast cells. A similar difference was observed with the beta gamma complex. These observations may indicate functional and constitutional differences of the high- and intermediate-affinity IL-2 receptor complexes between lymphoid and fibroblastoid cells.

Animals↗

PubMed1993

Interleukin-2 receptor heterotrimer complex and intracellular signaling.

Identification of a third component of the IL-2 receptor complex, gamma-chain, has established that the high-affinity complex consists of at least three distinct subunits, alpha-, beta- and gamma-chains. The alpha-chain specifies the low-affinity IL-2 binding. The beta- or gamma-chains alone do not show any appreciable IL-2 binding activity, however simultaneous existence of both chains generates a functional receptor complex that is suggested to associate with a non-receptor type protein tyrosine kinase that may deliver IL-2-induced signals further downstream. Mutation studies have revealed that discrete cytoplasmic regions of the beta- and gamma-chains transduce at least two independent signaling pathways.

Animals↗

PubMed1992

Expression of calpain II gene in human hematopoietic system cells infected with human T-cell leukemia virus type I.

We examined the distribution of calpains I and II in human hematopoietic system cell lines by Western and Northern blot analyses and enzyme activity assay. Expression of calpain I, a low Ca(2+)-requiring cysteine protease, was observed in all human T-cell lines tested. By contrast, expression of calpain II, a high Ca(2+)-requiring form, in human T-cells was closely correlated with human T-cell leukemia virus type I (HTLV-I) infection, which is known to result in the expression of adult T-cell leukemia-associated antigens, interleukin-2 (IL-2) receptor alpha, and Ca(2+)-dependent cell proliferation. Specific expression of calpain II in HTLV-I-infected cells occurred at the mRNA level. Furthermore, expression of calpain II in human natural killer-like cells was augmented by HTLV-I pX gene transfection. In HTLV-I-infected cells, the trans-acting transcriptional activation of the long terminal repeat and control elements for the IL-2 receptor alpha, c-fos, and granulocyte-macrophage colony-stimulating factor genes by the Tax from the pX gene is already known. Our results suggest that the similar trans-activation occurs to the calpain II gene in HTLV-I-infected hematopoietic system cells.

Blotting, Western↗

PubMed1992

Cell type-specific tyrosine phosphorylation of IL-2 receptor beta chain in response to IL-2.

Functional activities of the IL-2 receptor (IL-2R) beta chain exogenously expressed on lymphoid and non-lymphoid cells were examined in terms of phosphorylation of IL-2R beta and cell growth. Lymphoid MOLT-4 and its transfectants expressing IL-2R beta either alone or with IL-2R alpha chain were found to be rapidly phosphorylated predominantly at tyrosine residues of IL-2R beta and to be affected in their growth in an IL-2-dependent manner. In contrast, IL-2 induced neither phosphorylation of IL-2R beta nor cell growth in non-lymphoid transfectants derived from COS7, HeLa and L929, even though they acquired the IL-2 binding ability when coexpressed as IL-2R beta and IL-2R alpha. These results suggest that IL-2 induces activation of a tyrosine kinase possibly associated with IL-2R beta in a cell type-specific manner.

Cell Division↗

PubMed1992

Cloning of the gamma chain of the human IL-2 receptor.

A third subunit, the gamma chain, of the human interleukin-2 receptor (IL-2R) was identified, and a complementary DNA clone encoding this member of the cytokine receptor family was isolated. The gamma chain is necessary for the formation of the high- and intermediate-affinity receptors, which consists of alpha beta gamma heterotrimers and beta gamma heterodimers, respectively. The IL-2R on murine fibroblastoid cells can be internalized after binding IL-2 only if the gamma chain is present; alpha and beta are insufficient for internalization. Thus, the gamma chain is an indispensable component of the functional IL-2R.

Amino Acid Sequence↗

PubMed1992

IL-2-dependent in vivo and in vitro tyrosine phosphorylation of IL-2 receptor gamma chain.

We previously reported a molecule, p64, which was tentatively named the gamma chain, coprecipitable with the beta chain of human interleukin-2 receptor (IL-2R). The present study demonstrated that the gamma chain, as well as the beta chain expressed on IL-2-responsive cells, is phosphorylated on tyrosine residues in an IL-2-dependent manner in vivo and in vitro. The in vivo tyrosine phosphorylation of both chains was similarly induced within 1 min after IL-2 stimulation, and their in vitro tyrosine phosphorylation with the anti-IL-2R beta antibody-directed immunocomplex was also increased by treatment of cells with IL-2. These results suggest that a tyrosine kinase is associated with the beta gamma subunit complex, of which activation by IL-2 may result in transduction of intracellular signals.

Amino Acid Sequence↗

PubMed1992

An associated molecule, p64, with IL-2 receptor beta chain. Its possible involvement in the formation of the functional intermediate-affinity IL-2 receptor complex.

We identified previously a membrane molecule, p64, which co-precipitates with the IL-2R beta-chain in human T cells. We have now investigated the biologic significance of p64 in the formation of the functional IL-2R complex with cell lines transfected with cDNA of IL-2R alpha- and/or beta-chains. Two functional parameters associated with IL-2R, IL-2 binding ability and association of p64 with the beta-chain, were examined. Two subclones, MOLT beta-11 and MOLT beta-12, of an IL-2R beta cDNA-transfected MOLT4 clone expressed similar numbers of IL-2R beta molecules on cell surfaces and bound to IL-2 with intermediate affinity. However, the numbers of IL-2 binding sites were significantly lower than those of IL-2R beta molecules and considerably different between the two subclones. The amount of p64 co-precipitated with IL-2R beta was proportional to numbers of the IL-2 binding sites in the two subclones. In addition, neither p64 co-precipitation nor IL-2 binding was detected in HeLa and COS7 cells transfected with IL-2R beta, and no p64 precipitation was seen even in those transfectants with both IL-2R alpha and beta cDNAs, which bind to IL-2 with high affinity but are not able to transduce intracellular signals. These results suggest the possibility that p64 associates with IL-2R beta and has an important role in formation of the functional IL-2R complex.

Binding Sites↗

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

Localization of an immunodominant domain on baculovirus-produced parvovirus B19 capsids: correlation to a major surface region on the native virus particle.

An immunodominant region on baculovirus-produced parvovirus B19 VP2 capsids was localized between amino acids 259 and 426 by mapping the binding sites of a panel of monoclonal antibodies which recognize determinants on the particles. The binding sites of three monoclonal antibodies were fine-mapped within this antigenic domain. Six VP2-specific monoclonal antibodies recognized determinants common to both the empty capsids and native parvovirus. The defined antigenic region is most probably exposed on the native B19 virion and corresponds to part of the threefold spike on the surface of canine parvovirus particles.

Amino Acid Sequence↗