Lee Hai-Chon, Shibata Hirofumi, Ogawa Shinya, Maki Kazushige, Ikuta Koichi
Laboratory of Biological Protection, Department of Biological Responses, Institute for Virus Research, Kyoto University, Kyoto, Japan.
J Immunol. 2005 Jun 15;174(12):7800-6. doi: 10.4049/jimmunol.174.12.7800.
Expression of the IL-7R alpha-chain (IL-7Ralpha) is strictly regulated during the development and maturation of lymphocytes. Glucocorticoids (GC) have pleiotypic effects on the growth and function of lymphocytes. Although GC have been reported to induce the transcription of IL-7Ralpha gene in human T cells, its molecular mechanism is largely unknown. In this study, we show that GC up-regulate the levels of IL-7Ralpha mRNA and protein in mouse T cells. This effect does not require protein synthesis de novo, because protein synthesis inhibitors do not block the process. Mouse IL-7Ralpha promoter has striking homology with human and rat, containing consensus motifs of Ikaros, PU.1, and Runx1 transcription factors. In addition, a conserved noncoding sequence (CNS) of approximately 270 bp was found 3.6-kb upstream of the promoter, which was designated as CNS-1. A GC receptor (GR) motif is present in the CNS-1 region. Importantly, we show by reporter assay that the IL-7Ralpha promoter has specific transcription activity in T cells. This activity highly depends on the PU.1 motif. Furthermore, GC treatment augments the transcriptional activity through the GR motif in the CNS-1 region. We also demonstrate that GR binds to the GR motif by EMSA. In addition, by chromatin immunoprecipitation assay, we show that GR is rapidly recruited to endogenous CNS-1 chromatin after GC stimulation. These results demonstrate that GR binds to the GR motif in the CNS-1 region after GC stimulation and then activates the transcription of the IL-7Ralpha promoter. Thus, this study identifies the IL-7Ralpha CNS-1 region as a GC-responsive element.
白细胞介素-7受体α链（IL-7Rα）的表达在淋巴细胞的发育和成熟过程中受到严格调控。糖皮质激素（GC）对淋巴细胞的生长和功能具有多效性作用。尽管已有报道称GC可诱导人T细胞中IL-7Rα基因的转录，但其分子机制仍 largely未知。在本研究中，我们发现GC可上调小鼠T细胞中IL-7Rα mRNA和蛋白的水平。此效应不需要从头合成蛋白质，因为蛋白质合成抑制剂不会阻断该过程。小鼠IL-7Rα启动子与人和大鼠的启动子具有显著同源性，包含Ikaros、PU.1和Runx1转录因子的共有基序。此外，在启动子上游3.6 kb处发现了一个约270 bp的保守非编码序列（CNS），被命名为CNS-1。在CNS-1区域存在一个糖皮质激素受体（GR）基序。重要的是，我们通过报告基因测定表明IL-7Rα启动子在T细胞中具有特异性转录活性。这种活性高度依赖于PU.1基序。此外，GC处理通过CNS-1区域中的GR基序增强转录活性。我们还通过电泳迁移率变动分析（EMSA）证明GR与GR基序结合。此外，通过染色质免疫沉淀测定，我们表明GC刺激后GR迅速被募集到内源性CNS-1染色质上。这些结果表明，GC刺激后GR与CNS-1区域中的GR基序结合，然后激活IL-7Rα启动子的转录。因此，本研究确定IL-7Rα CNS-1区域为GC反应元件。
