Regulation of the Human Interleukin-5 Promoter by Ets Transcription Factors (original) (raw)
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The Journal of Immunology, 2004
CD5 is a surface receptor constitutively expressed on thymocytes and mature T and B-1a cells. CD5 expression is tightly regulated during T and B cell development and activation processes. In this study we shown that the constitutive expression of CD5 on human T cells correlates with the presence of a DNase I-hypersensitive (DH) site at the 5-flanking region of CD5. Human CD5 is a TATA-less gene for which 5-RACE analysis shows multiple transcriptional start sites, the most frequent of which locates within an initiator sequence. Luciferase reporter assays indicate that a 282-bp region upstream of the initiation ATG displays full promoter activity in human T cells. Two conserved Ets-binding sites (at positions ؊239 and ؊185) were identified as functionally relevant to CD5 expression by site-directed mutagenesis, EMSAs, and cotransfection experiments. A possible contribution of Sp1 (؊115 and ؊95), c-Myb (؊177), and AP-1-like (؊151) motifs was also detected. Further DH site analyses revealed an inducible DH site 10 kb upstream of the human CD5 gene in both T and B CD5 ؉ cells. Interestingly, a 140-bp sequence showing high homology with a murine inducible enhancer is found within that site. The data presented indicate that the 5-flanking region of human CD5 is transcriptionally active in T cells, and that Ets transcription factors in conjunction with other regulatory elements are responsible for constitutive and tissue-specific CD5 expression.
Oncogene, 2000
Activation of Stat5 by many cytokines implies that it cannot alone insure the speci®city of the regulation of its target genes. We have evidenced a physical and functional interaction between members of two unrelated transcription factor families, Ets-1, Ets-2 and Stat5, which could contribute to the proliferative response to interleukin 2. Competition with GAS-and EBS-speci®c oligonucleotides and immunoassays with a set of anti-Stat and anti-Ets families revealed that the IL-2-induced Stat5-Ets complex recognizes several GAS motifs identi®ed as target sites for activated Stat5 dimers. Coimmunoprecipitation experiments evidenced that a Stat5/Ets-1/2 complex is formed in vivo in absence of DNA. GST-pull down experiments demonstrated that the C-terminal domain of Ets-1 is sucient for this interaction in vitro. Cotransfection experiments in Kit225 T cells resulted in cooperative transcriptional activity between both transcription factors in response to a combination of IL-2, PMA and ionomycin. A Stat5-Ets protein complex was the major inducible DNAbinding complex bound to the human IL-2rE GASd/ EBSd motif in long-term proliferating normal human T cells activated by CD2 and CD28. These results suggest that the inducible Stat5-Ets protein interaction plays a role in the regulation of gene expression in response to IL-2 in human T lymphocytes.
Lymphoid enhancer factor interacts with GATA-3 and controls its function in T helper type 2 cells
Immunology, 2008
GATA-3 is the master transcription factor for T helper 2 (Th2) cell differentiation and is critical for the expression of Th2 cytokines. Little is known, however, about the nature of the functional molecular complexes of GATA-3. We identified a high-mobility group (HMG)-box type transcription factor, lymphoid enhancer factor 1 (LEF-1), in the GATA-3 complex present in Th2 cells using a Flag-calmodulin-binding peptide (CBP)-tag based proteomics method. The interaction between GATA-3 and LEF-1 was confirmed by co-immunoprecipitation experiments using LEF-1-introduced T-cell lineage TG40 cells. The HMG-box domain of LEF-1 and two zinc finger domains of GATA-3 were found to be important for the physical association. The introduction of LEF-1 into developing Th2 cells resulted in the suppression of Th2 cytokine production. The suppression was significantly lower in the cells into which a HMG-boxdeleted LEF-1 mutant was introduced. Moreover, LEF-1 inhibited the binding activity of GATA-3 to the interleukin (IL)-5 promoter. These results suggest that LEF-1 is involved in the GATA-3 complex, while also regulating the GATA-3 function, such as the induction of Th2 cytokine expression via the inhibition of the DNA-binding activity of GATA-3.
The Journal of Immunology, 2007
Th2 cytokine expression is dependent on the transcription factor GATA-3. However, the molecular interactions of GATA-3 leading to Th2 cytokine gene activation have not been well characterized. Here, we reported a number of GATA-3 associated proteins in Th2 cells, and one of such proteins Pias1 functioned as a positive transcriptional coregulator for GATA-3. When overexpressed in Th2 cells, Pias1 enhanced the expression of IL-13, and to lesser degrees, IL-4 and-5. Conversely, Pias1 siRNA down-regulated the Th2 cytokine expression. In Leishmania major infection, manipulating Pias1 expression in parasite-reactive CD4 T cells altered severity of disease caused by Th2 responses. Mechanistically, Pias1 markedly potentiated GATA-3-mediated activation of the IL-13 promoter by facilitating the recruitment of GATA-3 to the promoter. In contrast, IL-5 promoter was modestly enhanced by Pias1 and no effect was observed on IL-4 promoter. Thus, both promoter activation and additional mechanisms are responsible for regulation by Pias1.
Friend of GATA-1 Represses GATA-3-dependent Activity in CD4+ T Cells
Journal of Experimental Medicine, 2001
The development of naive CD4 ϩ T cells into a T helper (Th) 2 subset capable of producing interleukin (IL)-4, IL-5, and IL-13 involves a signal transducer and activator of transcription (Stat)6-dependent induction of GATA-3 expression, followed by Stat6-independent GATA-3 autoactivation. The friend of GATA (FOG)-1 protein regulates GATA transcription factor activity in several stages of hematopoietic development including erythrocyte and megakaryocyte differentiation, but whether FOG-1 regulates GATA-3 in T cells is uncertain. We show that FOG-1 can repress GATA-3-dependent activation of the IL-5 promoter in T cells. Also, FOG-1 overexpression during primary activation of naive T cells inhibited Th2 development in CD4 ϩ T cells. FOG-1 fully repressed GATA-3-dependent Th2 development and GATA-3 autoactivation, but not Stat6-dependent induction of GATA-3. FOG-1 overexpression repressed development of Th2 cells from naive T cells, but did not reverse the phenotype of fully committed Th2 cells. Thus, FOG-1 may be one factor capable of regulating the Th2 development.
Journal of Biological Chemistry, 2016
IL-2 is the first cytokine produced when naive T helper (Th) cells are activated and differentiate into dividing pre-Th0 proliferating precursors. IL-2 expression is blocked in naive, but not activated or memory, Th cells by the transcription factor Ets-2 that binds to the antigen receptor response element (ARRE)-2 of the proximal IL-2 promoter. Ets-2 acts as an independent preinduction repressor in naive Th cells and does not interact physically with the transcription factor NFAT (nuclear factor of activated T-cells) that binds to the ARRE-2 in activated Th cells. In naive Th cells, Ets-2 mRNA expression, Ets-2 protein levels, and Ets-2 binding to ARRE-2 decrease upon cell activation followed by the concomitant expression of IL-2. Cyclosporine A stabilizes Ets-2 mRNA and protein when the cells are activated. Ets-2 silences directly constitutive or induced IL-2 expression through the ARRE-2. Conversely, Ets-2 silencing allows for constitutive IL-2 expression in unstimulated cells. Ets-2 binding to ARRE-2 in chromatin is stronger in naive compared with activated or memory Th cells; in the latter, Ets-2 participates in a change of the IL-2 promoter architecture, possibly to facilitate a quick response when the cells re-encounter antigen. We propose that Ets-2 expression and protein binding to the ARRE-2 of the IL-2 promoter are part of a strictly regulated process that results in a physiological transition of naive Th cells to Th0 cells upon antigenic stimulation. Malfunction of such a repression mechanism at the molecular level could lead to a disturbance of later events in Th cell plasticity, leading to autoimmune diseases or other pathological conditions.
Stat6Independent GATA3 Autoactivation Directs IL4Independent Th2 Development and Commitment
Immunity, 2000
we presented evidence that Th2 cells Washington University School of Medicine can develop from naive T cells independently of IL-4 St. Louis, Missouri 63110 produced by non-T cells (Schmitz et al., 1994), sug- † Deutsches Rheumaforschungszentrum gesting either that naive T cells are capable of autono-Hannoversche Strasse 27 mous IL-4 production or that non-IL-4 signals can direct 10115 Berlin Th2 development. However, a mechanistic explanation Germany for factor-independent IL-4 production by naive T cells ‡ Miltenyi Biotec was not available at that time. Friedrich-Ebert-Strasse 68 The transcription factors GATA-3 and c-Maf are selec-51429 Bergisch Gladbach tively expressed in type 2 Th cells (Ho et al., 1996; Zhang Germany et al., 1997; Zheng and Flavell, 1997). Transgenic § Department of Experimental Rheumatology GATA-3 overexpression increases Th2 cytokine produc-Charite tion, and antisense-GATA-3 blockade reduces Th2 cyto-Humboldt University kines in Th2 clones (Zheng and Flavell, 1997). GATA-3 10115 Berlin activates the IL-5 promoter (Zhang et al., 1997), and Germany GATA-3-dependent enhancer activity has been found within several regions surrounding the IL-4 gene (Ranganath et al., 1998). GATA-3 expression by retrovirus in Summary developing Th1 cells blocks IL-12 receptor expression and prevents Th1 development independently of the The initial source of IL-4-inducing Th2 development induction of IL-4 (Ouyang et al., 1998). c-Maf directly and the mechanism of stable Th2 commitment remain augments IL-4 promoter activity through cooperative obscure. We found the reduced level of IL-4 production interactions with Nip-45, NF-AT (Ho et al., 1996, 1998), in Stat6-deficient T cells to be significantly higher than and JunB (Rincon et al., 1997b). Transgenic c-Maf overin Th1 controls. Using a novel cell surface affinity maexpression increases IL-4 and alters the isotype patterns trix technique, we found that IL-4-secreting Stat6-defiof antibody with increased IgE (Ho et al., 1998), and cient T cells stably expressed GATA-3 and Th2 phenoc-Maf-deficient T cells show a selective reduction in type. Introducing GATA-3 into Stat6-deficient T cells IL-4 but not other Th2-specific cytokines (Kim et al., completely restored Th2 development, inducing c-Maf, 1999a, 1999b).