Vacca A, Felli M P, Farina A R, Martinotti S, Maroder M, Screpanti I, Meco D, Petrangeli E, Frati L, Gulino A
Department of Experimental Medicine, University La Sapienza, Rome, Italy.
J Exp Med. 1992 Mar 1;175(3):637-46. doi: 10.1084/jem.175.3.637.
The immunosuppressant hormone dexamethasone (Dex) interferes with T cell-specific signals activating the enhancer sequences directing interleukin 2 (IL-2) transcription. We report that the Dex-dependent downregulation of 12-O-tetradecanoyl-phorbol-13-acetate (TPA) and calcium ionophore-induced activity of the IL-2 enhancer are mediated by glucocorticoid receptor (GR) via a process that requires intact NH2- and COOH-terminal and DNA-binding domains. Functional analysis of chloramphenicol acetyltransferase (CAT) vectors containing internal deletions of the -317 to +47 bp IL-2 enhancer showed that the GR-responsive elements mapped to regions containing nuclear factor of activated T cells protein (NFAT) (-279 to -263 bp) and AP-1 (-160 to -150 bp) motifs. The AP-1 motif binds TPA and calcium ionophore-induced nuclear factor(s) containing fos protein. TPA and calcium ionophore-induced transcriptional activation of homo-oligomers of the NFAT element were not inhibited by Dex, while AP-1 motif concatemers were not stimulated by TPA and calcium ionophore. When combined, NFAT and AP-1 motifs significantly synergized in directing CAT transcription. Such a synergism was impaired by specific mutations affecting the trans-acting factor binding to either NFAT or AP-1 motifs. In spite of the lack of hormone regulation of isolated cis elements, TPA/calcium ionophore-mediated activation of CAT vectors containing a combination of the NFAT and the AP-1 motifs became suppressible by Dex. Our results show that the IL-2-AP-1 motif confers GR sensitivity to a flanking region containing a NFAT element and suggest that synergistic cooperativity between the NFAT and AP-1 sites allows GR to mediate the Dex inhibition of IL-2 gene transcription. Therefore, a Dex-modulated second level of IL-2 enhancer regulation, based on a combinatorial modular interplay, appears to be present.
免疫抑制剂地塞米松（Dex）会干扰激活指导白细胞介素2（IL-2）转录的增强子序列的T细胞特异性信号。我们报告，12- O-十四烷酰佛波醇-13-乙酸酯（TPA）和钙离子载体诱导的IL-2增强子活性的Dex依赖性下调是由糖皮质激素受体（GR）通过一个需要完整的NH2和COOH末端以及DNA结合结构域的过程介导的。对含有-317至+47 bp IL-2增强子内部缺失的氯霉素乙酰转移酶（CAT）载体的功能分析表明，GR反应元件定位于含有活化T细胞核因子蛋白（NFAT）（-279至-263 bp）和AP-1（-160至-150 bp）基序的区域。AP-1基序结合TPA和钙离子载体诱导的含有fos蛋白的核因子。Dex不抑制TPA和钙离子载体诱导的NFAT元件同型寡聚体的转录激活，而AP-1基序串联体不受TPA和钙离子载体的刺激。当结合时，NFAT和AP-1基序在指导CAT转录方面有显著的协同作用。影响与NFAT或AP-1基序结合的反式作用因子的特定突变会损害这种协同作用。尽管分离的顺式元件缺乏激素调节，但含有NFAT和AP-1基序组合的CAT载体的TPA/钙离子载体介导的激活可被Dex抑制。我们的结果表明，IL-2-AP-1基序赋予GR对含有NFAT元件的侧翼区域的敏感性，并表明NFAT和AP-1位点之间的协同合作允许GR介导Dex对IL-2基因转录的抑制。因此，基于组合模块相互作用的Dex调节的IL-2增强子调控的第二水平似乎存在。
