Dimeric RFX proteins contribute to the activity and lineage specificity ofthe interleukin-5 receptor alpha promoter through activation and repression domains

Interleukin-5 (IL-5) plays a central role in the differentiation, prolifera tion, and functional activation of eosinophils, The specific action of IL-5 on eosinophils and hematopoietically related basophils is regulated by the restricted expression of IL-5 receptor alpha (IL-5R alpha), a subunit of h igh-affinity IL-5R, on these cells. We have previously identified an enhanc er-like cis element in the IL-5R alpha promoter that is important for both full promoter function and lineage-specific activity, Here, we demonstrate by yeast one-hybrid screening that RFX2 protein specifically binds to this cis element. RFX2 belongs to the RFX DNA-binding protein family, the biolog ical role of which remains obscure. Using an electrophoretic mobility shift assay, we further show that RFX1, RFX2, and RFX3 homodimers and heterodime rs specifically bind to the cis element of the IL-5R alpha. promoter. The m RNA expression of RFX1, RFX2, and RFX3 was detected ubiquitously, but in tr ansient-transfection assays, multimerized RFX binding sites in front of a b asal promoter efficiently functioned in a tissue- and lineage-specific mann er. To further investigate RFX functions on transcription, full-length and deletion mutants of RFX1 were targeted to DNA through fusion to the GAL4 DN A binding domain. Tissue- and lineage-specific transcriptional activation w ith the full-length RFX I fusion plasmid on a reporter controlled by GAL4 b inding sites was observed. Distinct activation and repression domains withi n the RFX1 protein were further mapped. Our findings suggest that RFX prote ins are transcription factors that contribute to the activity and lineage s pecificity of the IL-5R alpha promoter by directly binding: to a target cis element and cooperating with other tissue- and lineage-specific cofactors.