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 n onatopic asthma, elevated IL-5 has been detected in peripheral blood and th e 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 eis-regulatory elements and transc ription factors involved in the suppression of IL-5 transcription in T cell s. Methods: Methods used in this study include DNase I footprint assays, elect rophoretic 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 p roximal hIL-5 promoter. Functional analysis indicates that the BRs are invo lved in control of hIL-5 promoter activity. Two of these regions, BR3 and B R4 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 con tains a novel negative regulatory element located at positions -90 to -79 o f the hIL-5 promoter, which binds Oct1, octamer-like, and YY1 nuclear facto rs. Substitution mutations, which abolished binding of these proteins to th e BR3 sequence, significantly increased hIL-5 promoter activity in activate d 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 supp ression of IL-5 transcription in T cells.