scholarly journals Two essential regulatory elements in the human interferon gamma promoter confer activation specific expression in T cells.

1993 ◽  
Vol 178 (5) ◽  
pp. 1483-1496 ◽  
Author(s):  
L Penix ◽  
W M Weaver ◽  
Y Pang ◽  
H A Young ◽  
C B Wilson
Keyword(s):  
T Cells ◽  
Interferon Gamma ◽  
Regulatory Elements ◽  
T Cell Subsets ◽  
Jurkat T Cells ◽  
Intact Cells ◽  
Ifn Gamma ◽  
Promoter Function ◽  
Gm Csf ◽  
Nuclear Extracts

Like interleukin 2 (IL-2), interferon gamma (IFN-gamma) is an early response gene in T cells and both are prototypical T helper cell type 1 (Th-1) lymphokines. Yet IL-2 and IFN-gamma production are independently regulated, as demonstrated by their differential expression in certain T cell subsets, suggesting that the regulatory elements in these two genes must differ. To explore this possibility, the 5' flank of the human IFN-gamma gene was analyzed. Expression of IFN-gamma promoter-driven beta-galactosidase reporter constructs containing 538 bp of 5' flank was similar to that by constructs driven by the IL-2 promoter in activated Jurkat T cells; expression nearly as great was observed with the construct containing only 108 bp of IFN-gamma 5' flank. These IFN-gamma promoter constructs faithfully mirrored expression of the endogenous gene, in that expression required activation both with ionomycin and PMA, was inhibited by cyclosporin A, and was not observed in U937 or THP-1 cells. The region between -108 and -40 bp in the IFN-gamma promoter was required for promoter function and contained two elements that are conserved across species. Deletion of 10 bp within either element reduced promoter function by 70%, whereas deletions in nonconserved portions of this region had little effect on promoter function. The distal conserved element (-96 to -80 bp) contained a consensus GATA motif and a potential regulatory motif found in the promoter regions of the GM-CSF and macrophage inflammatory protein (MIP) genes. Factors binding to this element, including GATA-3, were found in Jurkat nuclear extracts by electromobility shift assays and two of the three complexes observed were altered in response to activation. One or both of these motifs are present in the 5' flank of multiple, other lymphokine genes, including IL-3, IL-4, IL-5, and GM-CSF, but neither is present in the promoter of the IL-2 gene. The proximal conserved element (-73 to -48 bp) shares homology with the NFIL-2A element in the IL-2 promoter; these elements compete for binding of factors in Jurkat nuclear extracts, although the NFIL-2A element but not the IFN-gamma element binds Oct-1. Factors binding to this element in the IFN-gamma gene were present in extracts from resting and activated Jurkat T cells. However, by in vivo footprinting of intact cells, this element was protected from methylation only with activation.(ABSTRACT TRUNCATED AT 400 WORDS)

scholarly journals Natural killer cells are a source of interferon gamma that drives differentiation of CD4+ T cell subsets and induces early resistance to Leishmania major in mice.

1993 ◽  
Vol 178 (2) ◽  
pp. 567-577 ◽  
Author(s):  
T M Scharton ◽  
P Scott
Keyword(s):  
T Cells ◽  
Nk Cells ◽  
Natural Killer ◽  
Interferon Gamma ◽  
Leishmania Major ◽  
Nk Cell ◽  
T Cell Subsets ◽  
Th2 Cells ◽  
Th1 Cell ◽  
Ifn Gamma

Infection of mice with the protozoan Leishmania major provides an excellent model to define the factors involved in T helper (Th) subset development, since Th1 cells confer protection in resistant strains of mice, whereas Th2 cells are associated with the fatal outcome of susceptible mice. We previously found that interferon gamma (IFN-gamma) was required for Th1 cell development after infection of mice with L. major. In this report, we evaluate the contribution of natural killer (NK) cells to IFN-gamma levels early in L. major infection. NK cell activity was higher in resistant C3H/HeN mice than in susceptible BALB/c mice during the first week of infection, and removal of NK cells significantly decreased IFN-gamma levels and promoted interleukin 4 (IL-4) production in both the draining lymph nodes and spleen. IFN-gamma production by NK cells required the presence of CD4+ T cells or IL-2, but not CD8+ T cells. Enhanced disease, as measured by parasite numbers and lesion development, was observed in NK cell-depleted mice. Furthermore, a comparison of the NK cell response and the subsequent parasite burden in several inbred strains of mice demonstrated that NK cells mediate early resistance to L. major. Together, these data indicate that the stimulation of NK cells, through the production of IFN-gamma, plays an important role in initiating Th1 cell differentiation in leishmaniasis and in controlling early resistance to L. major.

scholarly journals Interleukin 12 induces stable priming for interferon gamma (IFN-gamma) production during differentiation of human T helper (Th) cells and transient IFN-gamma production in established Th2 cell clones.

1994 ◽  
Vol 179 (4) ◽  
pp. 1273-1283 ◽  
Author(s):  
R Manetti ◽  
F Gerosa ◽  
M G Giudizi ◽  
R Biagiotti ◽  
P Parronchi ◽  
...  
Keyword(s):  
T Cells ◽  
Interferon Gamma ◽  
Specific Antigen ◽  
Th2 Cells ◽  
Interleukin 12 ◽  
T Helper ◽  
Ifn Gamma ◽  
Selection Of

Interleukin 12 (IL-12) facilitates the generation of a T helper type 1 (Th1) response, with high interferon gamma (IFN-gamma) production, while inhibiting the generation of IL-4-producing Th2 cells in polyclonal cultures of both human and murine T cells and in vivo in the mouse. In this study, we analyzed the effect of IL-12, present during cloning of human T cells, on the cytokine profile of the clones. The culture system used allows growth of clones from virtually every T cell, and thus excludes the possibility that selection of precommitted Th cell precursors plays a role in determining characteristics of the clones. IL-12 present during the cloning procedures endowed both CD4+ and CD8+ clones with the ability to produce IFN-gamma at levels severalfold higher than those observed in clones generated in the absence of IL-12. This priming was stable because the high levels of IFN-gamma production were maintained when the clones were cultured in the absence of IL-12 for 11 d. The CD4+ and some of the CD8+ clones produced variable amounts of IL-4. Unlike IFN-gamma, IL-4 production was not significantly different in clones generated in the presence or absence of IL-12. These data suggest that IL-12 primes the clone progenitors, inducing their differentiation to high IFN-gamma-producing clones. The suppression of IL-4-producing cells observed in polyclonally generated T cells in vivo and in vitro in the presence of IL-12 is not observed in this clonal model, suggesting that the suppression depends more on positive selection of non-IL-4-producing cells than on differentiation of individual clones. However, antigen-specific established Th2 clones that were unable to produce IFN-gamma with any other inducer did produce IFN-gamma at low but significant levels when stimulated with IL-12 in combination with specific antigen or insoluble anti-CD3 antibodies. This induction of IFN-gamma gene expression was transient, because culture of the established clones with IL-12 for up to 1 wk did not convert them into IFN-gamma producers when stimulated in the absence of IL-12. These results suggest that Th clones respond to IL-12 treatment either with a stable priming for IFN-gamma production or with only a transient low level expression of the IFN-gamma gene, depending on their stage of differentiation.

scholarly journals Specific expression of the human CD4 gene in mature CD4+ CD8- and immature CD4+ CD8+ T cells and in macrophages of transgenic mice

1994 ◽  
Vol 14 (2) ◽  
pp. 1084-1094
Author(s):  
Z Hanna ◽  
C Simard ◽  
A Laperrière ◽  
P Jolicoeur
Keyword(s):  
T Cells ◽  
T Cell ◽  
Transgenic Mice ◽  
Cd8 T Cells ◽  
Transcription Initiation ◽  
Critical Role ◽  
Regulatory Elements ◽  
T Cell Subsets ◽  
Double Positive ◽  
Specific Expression

The CD4 protein plays a critical role in the development and function of the immune system. To gain more insight into the mechanism of expression of the human CD4 gene, we cloned 42.2 kbp of genomic sequences comprising the CD4 gene and its surrounding sequences. Studies with transgenic mice revealed that a 12.6-kbp fragment of the human CD4 gene (comprising 2.6 kbp of 5' sequences upstream of the transcription initiation site, the first two exons and introns, and part of exon 3) contains the sequences required to support the appropriate expression in murine mature CD4+ CD8- T cells and macrophages but not in immature double-positive CD4+ CD8+ T cells. Expression in CD4+ CD8+ T cells was found to require additional regulatory elements present in a T-cell enhancer fragment recently identified for the murine CD4 gene (S. Sawada and D. R. Littman, Mol. Cell. Biol. 11:5506-5515, 1991). These results suggest that expression of CD4 in mature and immature T-cell subsets may be controlled by distinct and independent regulatory elements. Alternatively, specific regulatory elements may control the expression of CD4 at different levels in mature and immature T-cell subsets. Our data also indicate that mouse macrophages contain the regulatory factors necessary to transcribe the human CD4 gene.

scholarly journals Agonist-induced inhibition of phosphatidylserine synthesis is secondary to the emptying of intracellular Ca2+ stores in Jurkat T-cells

1992 ◽  
Vol 288 (3) ◽  
pp. 785-789 ◽  
Author(s):  
C Pelassy ◽  
J P Breittmayer ◽  
C Aussel
Keyword(s):  
T Cells ◽  
Exchange System ◽  
Jurkat T Cells ◽  
Experimental Conditions ◽  
Intact Cells ◽  
Base Exchange ◽  
Permeabilized Cells ◽  
Atpase Inhibitors ◽  
Storage Organelle ◽  
Intracellular Compartments

The biosynthesis of phosphatidylserine (PtdSer) by the serine base-exchange enzyme system, in Jurkat T-lymphocytes, was inhibited in intact cells maintained in low-Ca(2+)-containing buffer (< 10 microM-Ca2+) by using Ca2+ ionophores (A23187 or ionomycin). The rise in cytosolic Ca2+ concentration under these experimental conditions was only due to the release of Ca2+ from intracellular compartments, suggesting that the inhibition of PtdSer synthesis was correlated with the emptying of intracellular Ca2+ pools. This was further studied in saponin-permeabilized cells, in which PtdSer synthesis was found to be inhibited by EGTA, Ca2+ ionophores (A23187 or ionomycin) and Ca(2+)-ATPase inhibitors [thapsigargin or 2,5-di-(t-butyl)-benzohydroquinone]. Since Ca(2+)-ATPase inhibitors impaired refilling of the Ca2+ stores with Ca2+, and since in CD3-activated Jurkat T-cells the Ca2+ stores remained empty after 1 h of treatment with anti-CD3 monoclonal antibodies, we suggest that PtdSer synthesis is mainly regulated by the level of Ca2+ in the intracellular compartments and that the Ca(2+)-dependent serine base-exchange system responsible for PtdSer synthesis is probably located within or close to a Ca(2+)-storage organelle.

scholarly journals T Cell Production of GM-CSF Protects the Host during Experimental Tuberculosis

mBio ◽  
2017 ◽  
Vol 8 (6) ◽  
Author(s):  
Richard T. Robinson
Keyword(s):  
T Cells ◽  
T Cell ◽  
Experimental Models ◽  
T Cell Subsets ◽  
Experimental Tuberculosis ◽  
Gm Csf ◽  
Macrophage Colony Stimulating Factor ◽  
Mechanistic Basis ◽  
Macrophage Colony ◽  
Stimulating Factor

ABSTRACT Although classically associated with myelopoiesis, granulocyte-macrophage colony-stimulating factor (GM-CSF) is increasingly recognized as being important for tuberculosis (TB) resistance. GM-CSF is expressed by nonhematopoietic and hematopoietic lineages following infection with Mycobacterium tuberculosis and is necessary to restrict M. tuberculosis growth in experimental models. Until the recent study by Rothchild et al. (mBio 8:e01514-17, 2017, https://doi.org/10.1128/mBio.01514-17 !), it was unknown whether GM-CSF-producing T cells contribute to TB resistance. Rothchild et al. identify which conventional and nonconventional T cell subsets produce GM-CSF during experimental TB, establish their protective nature using a variety of approaches, and provide a mechanistic basis for their ability to restrict M. tuberculosis growth. This commentary discusses the significance of these findings to basic and applied TB research. As translated to human disease, these findings suggest vaccine-mediated expansion of GM-CSF-producing T cells could be an effective prophylactic or therapeutic TB strategy.

scholarly journals Production of hematopoietic colony-stimulating factors by human natural killer cells.

1989 ◽  
Vol 169 (2) ◽  
pp. 569-583 ◽  
Author(s):  
M C Cuturi ◽  
I Anegón ◽  
F Sherman ◽  
R Loudon ◽  
S C Clark ◽  
...  
Keyword(s):  
T Cells ◽  
Biological Activity ◽  
Nk Cells ◽  
Negative Selection ◽  
Lymphoblastoid Cell Lines ◽  
Ionophore A23187 ◽  
Ifn Gamma ◽  
Gm Csf ◽  
Human Natural Killer Cells ◽  
Kinetics Of

We have analyzed the ability of highly purified preparations of human NK cells to produce CSF. NK cells, purified by negative selection from 10-d cultures of PBMC incubated with irradiated B-lymphoblastoid cell lines, were stimulated with rIL-2, FcR(CD16) ligands (particulate immune complexes or anti-CD16 antibodies bound to Sepharose), a combination of CD16 ligands and rIL-2, or the phorbol diester phorbol dibutyrate (PDBu) together with the Ca2+ ionophore A23187. Both rIL-2 and CD16 ligands induce accumulation of GM-CSF mRNA in NK cells and the combined effect of the two stimuli is synergistic. Maximal accumulation of GM-CSF mRNA is observed after PDBu/A23187 stimulation. The participation of contaminant T cells in the observed expression of the GM-CSF gene is excluded because CD16 ligands do not stimulate T cells and CD3 ligands, powerful stimulators of T cells, are inactive on NK cells. Accumulation of CSF-1 mRNA is observed only in NK cells stimulated with both CD16 ligands and rIL-2, whereas accumulation of IL-3 mRNA is observed only in NK cells stimulated with PDBu/A23187. Transcripts of the G-CSF, IL-1 alpha, and IL-1 beta genes were never detected in NK cells in these experiments. The kinetics of accumulation of GM-CSF and CSF-1 mRNA in NK cells stimulated with CD16 ligands and rIL-2 peaked at 2-4 h and was slower than that of TNF and IFN-gamma mRNA, which peak at 1 h. GM-CSF was precipitated from the supernatant fluids of NK cells stimulated with PDBu/A23187 and its biological activity was demonstrated by the ability of the supernatants to sustain proliferation of the TALL-101 cell line or CML blasts. Biological activity of IL-3 and CSF-1 was demonstrable in supernatant fluids of NK cells stimulated with PDBu/A23187 and CD16 ligands/rIL-2, respectively.

scholarly journals CD4+ effector cells default to the Th2 pathway in interferon gamma-deficient mice infected with Leishmania major.

1994 ◽  
Vol 179 (4) ◽  
pp. 1367-1371 ◽  
Author(s):  
Z E Wang ◽  
S L Reiner ◽  
S Zheng ◽  
D K Dalton ◽  
R M Locksley
Keyword(s):  
T Cells ◽  
Interferon Gamma ◽  
Knockout Mice ◽  
Leishmania Major ◽  
Interleukin 4 ◽  
Th2 Cells ◽  
Th1 Cells ◽  
Wild Type ◽  
Cd4 Cells ◽  
Ifn Gamma

Mice with homologous disruption of the interferon gamma (IFN-gamma) gene on the C57BL/6 background were infected with Leishmania major and the immune response assessed. In contrast to wild-type or heterozygous knockout mice, deficient animals were unable to restrict growth of the parasite and suffered lethal infection over 6-8 wk. Although wild-type and heterozygous littermates developed CD4+ cells that contained transcripts for IFN-gamma and lymphotoxin, typical of T helper type 1 (Th1) cells, the knockout mice developed CD4+ cells that contained transcripts for interleukin 4 (IL-4), IL-5, and IL-13, typical of Th2 cells. ELISPOT assays confirmed the reciprocal patterns of IFN-gamma or IL-4 production by T cells in similar frequencies in the respective groups of mice, and antibody analysis confirmed the presence of Th2-mediated isotype switching in the knockout mice. These data suggest that CD4+ T cells that normally respond to antigens by differentiation to Th1 cells default to the Th2 pathway in the absence of endogenous IFN-gamma.

scholarly journals Rapid activation of proteins that interact with the interferon gamma activation site in response to multiple cytokines

Blood ◽  
1994 ◽  
Vol 83 (8) ◽  
pp. 2063-2071 ◽  
Author(s):  
P Lamb ◽  
LV Kessler ◽  
C Suto ◽  
DE Levy ◽  
HM Seidel ◽  
...  
Keyword(s):  
Gene Expression ◽  
Dna Binding ◽  
Interferon Gamma ◽  
Regulation Of Gene Expression ◽  
Interleukin 2 ◽  
Sequence Element ◽  
Ifn Gamma ◽  
Gm Csf ◽  
Rapid Changes ◽  
Rapid Activation

Many cytokines and growth factors trigger rapid changes in gene expression upon binding to their receptors. In many cases, the mechanism by which these changes are affected is unknown. In this report, we show that interleukin-2 (IL-2), IL-3, IL-4, IL-6, leukemia inhibitory factor (LIF), erythropoietin (Epo), and granulocyte- macrophage colony-stimulating factor (GM-CSF) treatment of cells causes rapid activation of DNA-binding activities that recognize a DNA sequence element previously implicated in regulation of gene expression by interferon gamma (IFN gamma). The IL-4-, IL-6-, and GM-CSF-induced complexes can be distinguished from the recently characterized IFN gamma-activated protein p91 on the basis of mobility in polyacrylamide gels, sequence preferences, and lack of reactivity with an anti-p91 antiserum. The IL-4- and GM-CSF-induced complexes react with antiphosphotyrosine antibodies, demonstrating the presence of phosphotyrosine-containing proteins in these DNA-binding complexes. Transcriptional activation of a reporter gene linked to a synthetic IFN gamma-responsive promoter is observed in response to IFN gamma, IL-6, and LIF. These data suggest a pathway by which cytokines induce rapid changes in gene expression.

scholarly journals Idiotype-reactive T-cell subsets and tumor load in monoclonal gammopathies

Blood ◽  
1995 ◽  
Vol 86 (8) ◽  
pp. 3043-3049 ◽  
Author(s):  
Q Yi ◽  
A Osterborg ◽  
S Bergenbrant ◽  
H Mellstedt ◽  
G Holm ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Interleukin 2 ◽  
Human Tumor ◽  
Stage I ◽  
Stage Ii ◽  
T Cell Subsets ◽  
Ifn Gamma ◽  
Tumor Load ◽  
Th1 And Th2

The presence of idiotype-reactive T-cell subsets and their relation to the tumor load were analyzed in 9 patients with monoclonal gammopathy of undetermined significance (MGUS), in 12 patients with multiple myeloma (MM) clinical stage I, and in 9 patients with MM stage II/III. An enzyme-linked immunospot assay was used to identify interferon-gamma (IFN-gamma)-, interleukin-2 (IL-2)-, or IL-4-secreting T cells after stimulation by F(ab')2 fragments of monoclonal IgG. The response to autologous IgG was significantly higher than that induced by isotypic monoclonal IgG. Comparable results were obtained in a proliferation assay (3H-thymidine incorporation). A total of 8 of 9 patients with MGUS, 7 of 12 patients with MM stage I, and 3 of 9 with MM stage II/III had T cells secreting IFN-gamma and/or IL-2 (T helper [Th1] type-1 cells), whereas cells secreting both Th1 and Th2 or Th0 types of cytokines were more frequent in patients with MM, particularly in those with MM stage II/III. The number and frequency of Th1-type cells were significantly higher in MGUS patients as compared with those of MM stage II/III. The results indicate that idiotype-reactive T cells of the Th1 and Th2 or Th0 subsets were present in MGs and might provide indirect evidence that idiotype-reactive Th1-type cells may have a regulatory impact on the human tumor B cells.

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