Dimeric RFX Proteins Contribute to the Activity and Lineage Specificity of the Interleukin-5 Receptor α Promoter through Activation and Repression Domains
Dimeric RFX Proteins Contribute to the Activity and Lineage Specificity of the Interleukin-5 Receptor α Promoter through Activation and Repression Domains
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DOI：
10.1128/mcb.19.6.3940
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发表时间：
1999-06
影响因子：
5.3
通讯作者：
Atsushi Iwama;Jing Pan;Pu Zhang;W. Reith;B. Mach;D. Tenen;Zijie Sun
中科院分区：
文献类型：
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作者：
Atsushi Iwama;Jing Pan;Pu Zhang;W. Reith;B. Mach;D. Tenen;Zijie Sun
白细胞介素-5（IL-5）在嗜酸性粒细胞的分化、增殖和功能活化中起着重要作用。IL-5对嗜酸性粒细胞和造血相关嗜碱性粒细胞的特异性作用受这些细胞上IL-5受体α（IL-5 R α）（高亲和力IL-5 R的亚基）的限制性表达调节。我们之前已经在IL-5 R α启动子中鉴定了一个增强子样顺式元件，该元件对完整的启动子功能和谱系特异性活性都很重要。在这里，我们证明酵母单杂交筛选RFX 2蛋白特异性结合到这个顺式元件。RFX 2属于RFX DNA结合蛋白家族，其生物学作用仍不清楚。使用电泳迁移率变动分析，我们进一步表明RFX 1、RFX 2和RFX 3同源二聚体和异源二聚体特异性结合IL-5 R α启动子的顺式元件。RFX 1，RFX 2和RFX 3的mRNA表达被普遍检测到，但在瞬时转染试验中，在基础启动子前的多聚化RFX结合位点以组织和谱系特异性的方式有效地发挥作用。为了进一步研究RFX对转录的功能，通过与GAL 4 DNA结合结构域融合将RFX 1的全长和缺失突变体靶向DNA。组织和谱系特异性转录激活与全长RFX 1融合质粒上的GAL 4结合位点控制的报告进行了观察。RFX 1蛋白内不同的激活和抑制结构域被进一步定位。我们的研究结果表明，RFX蛋白是转录因子，通过直接结合靶顺式元件并与其他组织和谱系特异性辅因子合作，促进IL-5 R α启动子的活性和谱系特异性。
ABSTRACT Interleukin-5 (IL-5) plays a central role in the differentiation, proliferation, 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 α (IL-5Rα), a subunit of high-affinity IL-5R, on these cells. We have previously identified an enhancer-like cis element in the IL-5Rα 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 ciselement. RFX2 belongs to the RFX DNA-binding protein family, the biological role of which remains obscure. Using an electrophoretic mobility shift assay, we further show that RFX1, RFX2, and RFX3 homodimers and heterodimers specifically bind to the ciselement of the IL-5Rα promoter. The mRNA expression of RFX1, RFX2, and RFX3 was detected ubiquitously, but in transient-transfection assays, multimerized RFX binding sites in front of a basal promoter efficiently functioned in a tissue- and lineage-specific manner. To further investigate RFX functions on transcription, full-length and deletion mutants of RFX1 were targeted to DNA through fusion to the GAL4 DNA binding domain. Tissue- and lineage-specific transcriptional activation with the full-length RFX1 fusion plasmid on a reporter controlled by GAL4 binding sites was observed. Distinct activation and repression domains within the RFX1 protein were further mapped. Our findings suggest that RFX proteins are transcription factors that contribute to the activity and lineage specificity of the IL-5Rα promoter by directly binding to a target cis element and cooperating with other tissue- and lineage-specific cofactors.