Wang Jun, Shannon M Frances, Young Ian G
Division of Molecular Bioscience, John Curtin School of Medical Research, Australian National University, Mills Road, Acton, ACT 0200 Australia.
Int Immunol. 2006 Feb;18(2):313-23. doi: 10.1093/intimm/dxh370. Epub 2005 Dec 22.
IL-5 is a key regulator of eosinophilic inflammation and is selectively expressed by antigen-activated Th2 lymphocytes. An important role for the proximal AP-1 and GATA sites in regulating IL-5 transcription is generally accepted but the significance of an adjacent Ets/NFAT site has remained unclear. We have investigated its role using the mouse Th2 clone D10.G4.1. Transcription of IL-5 reporter gene plasmids could be induced in D10 cells by phorbol myristate acetate/cyclic adenosine monophosphate (PMA/cAMP) stimulation and significantly further enhanced by activation of the mitogen-activated protein (MAP) kinase pathways. Strong induction of IL-5 mRNA was also induced by PMA/cAMP. Mutagenesis showed that the Ets/NFAT site is of critical importance along with the AP-1 and GATA sites in regulating IL-5 transcription stimulated by PMA/cAMP and MAP kinase activation. Transactivation was used to investigate the transcription factors which could function at the three sites and possible synergistic interactions. AP-1 (c-Fos/c-Jun) strongly induced IL-5 transcription and dominant negative AP-1 constructs confirmed that AP-1 plays an important role in regulating IL-5 expression. Ets1, unlike other members of the Ets/NFAT family, synergized strongly with AP-1 suggesting that Ets1 is the family member which functions at the Ets/NFAT site. AP-1/Ets1 transactivation also stimulated IL-5 mRNA expression. Ets1 binding to the proximal promoter region, demonstrated by chromatin immunoprecipitation, was stimulated by PMA/cAMP. The absolute dependence on the binding sites for Ets1, AP-1 and GATA-3 together with the strong synergy between Ets1 and AP-1 suggest close cooperative interactions between the three transcription factors in the regulation of IL-5 expression in mouse T cells.
白细胞介素-5（IL-5）是嗜酸性粒细胞炎症的关键调节因子，由抗原激活的Th2淋巴细胞选择性表达。近端激活蛋白-1（AP-1）和GATA位点在调节IL-5转录中起重要作用，这一点已被广泛接受，但相邻的Ets/NFAT位点的意义仍不清楚。我们使用小鼠Th2克隆D10.G4.1研究了其作用。佛波酯肉豆蔻酸酯/环磷酸腺苷（PMA/cAMP）刺激可诱导D10细胞中IL-5报告基因质粒的转录，丝裂原活化蛋白（MAP）激酶途径的激活可进一步显著增强转录。PMA/cAMP也可强烈诱导IL-5 mRNA的表达。诱变表明，Ets/NFAT位点与AP-1和GATA位点一起，在调节PMA/cAMP和MAP激酶激活所刺激的IL-5转录中至关重要。采用反式激活研究了可在这三个位点发挥作用的转录因子以及可能的协同相互作用。AP-1（c-Fos/c-Jun）强烈诱导IL-5转录，显性负性AP-1构建体证实AP-1在调节IL-5表达中起重要作用。与Ets/NFAT家族的其他成员不同，Ets1与AP-1有强烈的协同作用，表明Ets1是在Ets/NFAT位点发挥作用的家族成员。AP-1/Ets1反式激活也刺激了IL-5 mRNA的表达。染色质免疫沉淀显示，PMA/cAMP可刺激Ets1与近端启动子区域的结合。对Ets1、AP-1和GATA-3结合位点的绝对依赖性以及Ets1与AP-1之间的强烈协同作用表明，这三种转录因子在调节小鼠T细胞中IL-5表达时存在密切的合作相互作用。
