Transcription factor GATA-3 is involved in repression of promoter activity of the human interleukin-4 gene.

GATA-3 was shown to bind to two sites of the IL-4 gene promoter in human T-cell lines PER-117 and Jurkat. A motif located in the region of position -860 and responsible for GATA-3 binding was detected for the first time. Mutation or deletion of this site increased the promoter activity. The findings suggest a direct involvement of GATA-3 in regulation of the human IL-4 gene transcription as a repressor of the promoter activity.

Dexamethasone suppresses human interleukin-5 gene promoter.

Synthetic glucocorticoid dexamethasone suppressed interleukin-5 gene expression in PER-117 human T cells at the level of transcription. The conserved lymphokine element 0 in the interleukin-5 gene promoter context served as a target for dexamethasone.

GATA-3 has dual regulatory functions in human interleukin-5 transcription.

Interleukin-5 (IL-5) is a T-cell cytokine involved in Type 2 diseases and is commonly described as being coordinately regulated with other Type 2 cytokines, such as IL-4 and IL-13. Considering the unique control of eosinophilia by IL-5, such coordinate regulation would be surprising. In fact, the biological specificity of eosinophilia and its control by IL-5 suggests a unique and independent control of IL-5 regulation. In this report we show the binding of GATA-3 to three sites in the human IL-5 promoter in the human T-cell line PER117. The previously identified -70 site and another site at position -152 are shown to positively regulate IL-5 transcription. More importantly, the site located at -400 acts as a powerful repressor of IL-5 transcription with mutagenesis of this site allowing a high level expression of IL-5 without the activation of other factors normally required for IL-5 expression. Whereas GATA-3 has been proposed to be involved in the regulation of the IL-4/IL-5/IL-13 locus, we show here that it has another function in controlling IL-5 transcription that supports the observed unique biological function of this cytokine.

Binding of octamer factors to the murine IL-5 CLE0 in primary T-cells and a T-cell line.

Interleukin-5 (IL-5) is an inducible T-cell derived cytokine with remarkable specificity for the eosinophil lineage. It is controlled at the level of transcription and regulation of the gene is an obvious target for therapy of eosinophil-dependent allergic disorders such as asthma, eczema and rhinitis. Using a T-cell line and primary T-cells we have shown for the first time that the Oct1 and Oct2 transcription factors combine to form a complex with the functionally critical murine IL-5 cis-regulatory element, conserved lymphokine element 0 (CLE0), and contribute to positive regulation of the gene. These results show the increasingly important role of octamer factors in regulation of the IL-5 gene.

Nuclear factor of activated T cells and YY1 combine to repress IL-5 expression in a human T-cell line.

BACKGROUND: IL-5 is an inducible T-cell cytokine with the unique ability to induce eosinophilia without increases in other cell compartments. Regulation of IL-5 expression is controlled primarily at the level of transcription. The role of eosinophilia in allergic disorders indicates IL-5 as a target for therapy. OBJECTIVE: This report aims to increase our understanding of IL-5 gene regulation by identifying distal control elements in the human (h) IL-5 promoter, determining the transcription factors that bind these elements and elucidating their role in control of hIL-5 gene expression. METHODS: Methods used in this study include deoxyribonuclease I footprint analysis, electrophoretic mobility shift assay, and functional analysis by transfection of PER-117 cells with site-directed mutants of the hIL-5 promoter. RESULTS: We have identified a protected region in the distal hIL-5 promoter that has sequence homology to the previously identified negative regulatory element within BR3. This protected region has not been previously reported and is shown to contain overlapping binding sites for YY1 and nuclear factor of activated cells. The binding sites exist between positions -447 and -459, and this sequence was named hPRE2-IL5. Substitution mutations that abolish binding of these proteins to hPRE2-IL5 result in a 2- to 3-fold increase in hIL-5 promoter activity in activated human T cells. CONCLUSION: We report the novel combination of YY1 and nuclear factor of activated T cells transcription factors binding to a distal hIL-5 promoter element where both factors are involved in down-regulation of hIL-5 gene expression in human T cell.

Binding of YY1 and Oct1 to a novel element that downregulates expression of IL-5 in human T cells.

BACKGROUND: IL-5 controls development of eosinophilia and has been shown to be involved in the pathogenesis of allergic diseases. In both atopic and nonatopic asthma, elevated IL-5 has been detected in peripheral blood and the airways. IL-5 is produced mainly by activated T cells, and its expression is regulated at the transcriptional level. OBJECTIVE: This study focuses on the functional analysis of the human IL-5 (hIL-5) promoter and characterization of cis -regulatory elements and transcription factors involved in the suppression of IL-5 transcription in T cells. METHODS: Methods used in this study include DNase I footprint assays, electrophoretic mobility shift assays, and functional analysis by mammalian cell transfection involving deletion analysis and site-directed mutagenesis. RESULTS: We identified 5 protein binding regions (BRs) located within the proximal hIL-5 promoter. Functional analysis indicates that the BRs are involved in control of hIL-5 promoter activity. Two of these regions, BR3 and BR4 located at positions -102 to -73, have not previously been described as regulators of IL-5 expression in T cells. We show that the BR3 sequence contains a novel negative regulatory element located at positions -90 to -79 of the hIL-5 promoter, which binds Oct1, octamer-like, and YY1 nuclear factors. Substitution mutations, which abolished binding of these proteins to the BR3 sequence, significantly increased hIL-5 promoter activity in activated T cells. CONCLUSION: We suggest that Oct1, YY1, and octamer-like factors binding to the -90/-79 sequence within the proximal IL-5 promoter are involved in suppression of IL-5 transcription in T cells.

Identification of two novel palindromic regulatory elements in the murine interleukin-5 promoter.

Interleukin-5 has remarkable specificity for the eosinophil lineage. This fact, combined with the biological specificity of eosinophilia suggests tight and independent regulation of IL-5 expression. Here we report two novel palindromic regulatory elements (PRE) which contain positive regulatory motifs (PRM) that control transcription of the murine IL-5 gene. The first element, mPRE1-IL5 (-79 to -90) contains the mPRM1 at positions -87 to -89 which operates as a positive regulatory element with mutation of this motif resulting in a 64% decrease in gene activity. Gene expression was reduced by 67% when a similar mutation was introduced into the mPRM2 (-467 to -469) of mPRE2-IL5 (-459 to -470). Both elements specifically bind proteins from EL4-23 cell nuclear extracts forming constitutive DNA-protein complexes. EMSA experiments utilising mutated mPRE-IL5 oligonucleotides indicate that in both elements, the mPRMs are essential for protein binding.

New directions in understanding interleukin-5 gene expression.

Specific regulation of the interleukin-5 (IL-5) gene is implied by the unique control of eosinophilia which is regulated by IL-5. In studies of IL-5 gene expression, the only control elements identified for the IL-5 gene have been transcriptional elements in the 5' untranslated region (UTR). Significant differences exist in the arrangement of the murine and human IL-5 promoters, which is surprising considering the tight regulation of the gene. Novel palindromic regulatory elements involved in transcriptional regulation have been found in the 5' UTR and new results show the presence of transcriptional elements in the 3' UTR. Post-transcriptional control mechanisms in both the 5' and 3' UTRs have also been described.