Palvimo J J, Reinikainen P, Ikonen T, Kallio P J, Moilanen A, Jänne O A
Institute of Biomedicine, Department of Physiology, University of Helsinki, FIN-00014 Helsinki, Finland.
J Biol Chem. 1996 Sep 27;271(39):24151-6. doi: 10.1074/jbc.271.39.24151.
Cross-modulation between androgen receptor (AR) and NF-kappaB/Rel proteins was studied using various androgen- and NF-kappaB-regulated reporter genes under transient transfection conditions. In COS-1 cells, elevated expression of RelA (p65) repressed AR-mediated transactivation in a dose-dependent manner, whereas NFkappaB1 (p50), another major member of the NF-kappaB family, did not influence transactivation. The repression of AR appeared to involve the N-terminal region of the protein between residue 297 and the DNA-binding domain. RelA-mediated transrepression could not be overcome by increasing the amount of AR. Transcriptional interference between RelA and AR was mutual in that cotransfected AR was able to attenuate transactivation by RelA in a dose- and steroid-dependent fashion. An excess of RelA was able to rescue the repression to some extent. Immunological analyses of RelA and AR protein levels indicated that transrepression was not due to reciprocal decrease in their amounts. Neither did AR increase the concentration of IkappaBalpha, which can sequester and inactivate RelA. Electrophoretic mobility shift assays using extracts from cotransfected cells and purified recombinant proteins showed that AR and RelA did not significantly influence each other's DNA binding activity. Nevertheless, protein-protein interaction experiments demonstrated a weak association between AR and RelA. Collectively, these data suggest that the mutual repression in intact cells is due to formation of AR-RelA complexes that are held together by another partner or to competition for a coactivator required for transcription.
在瞬时转染条件下，利用各种雄激素和核因子κB（NF-κB）调控的报告基因，研究了雄激素受体（AR）与NF-κB/Rel蛋白之间的交叉调节作用。在COS-1细胞中，RelA（p65）表达升高以剂量依赖的方式抑制AR介导的反式激活，而NF-κB家族的另一个主要成员NF-κB1（p50）则不影响反式激活。AR的抑制似乎涉及该蛋白297位残基与DNA结合结构域之间的N端区域。增加AR的量并不能克服RelA介导的反式抑制作用。RelA和AR之间的转录干扰是相互的，因为共转染的AR能够以剂量和类固醇依赖的方式减弱RelA的反式激活。过量的RelA能够在一定程度上挽救这种抑制作用。对RelA和AR蛋白水平的免疫分析表明，反式抑制并非由于它们的量相互减少所致。AR也没有增加可隔离并使RelA失活的IκBα的浓度。使用共转染细胞提取物和纯化重组蛋白进行的电泳迁移率变动分析表明，AR和RelA不会显著影响彼此的DNA结合活性。然而，蛋白质-蛋白质相互作用实验证明AR和RelA之间存在弱关联。总体而言，这些数据表明完整细胞中的相互抑制是由于AR-RelA复合物的形成，该复合物由另一个伴侣结合在一起，或者是由于竞争转录所需的共激活因子所致。
