Takeda Yukimasa, Kang Hong Soon, Jetten Anton M
National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, NC, USA.
Methods Mol Biol. 2019;1966:193-202. doi: 10.1007/978-1-4939-9195-2_15.
Here, we describe several assays to analyze the transcriptional activity of retinoic acid-related orphan receptors (RORs) and the effect of inverse agonists on their activity. One assay measures the effect of an inverse agonist on the transcriptional activation of a luciferase reporter by RORs in a Tet-On cell system. A mammalian two-hybrid assay analyzes the interaction of the ROR ligand binding domain with a coactivator peptide. Two additional assays examine the effect of an inverse agonist on the activation of a luciferase reporter under control of the promoter of the ROR target gene, IL17, and on ROR-mediated activation using a mammalian monohybrid assay.
在此，我们描述了几种分析维甲酸相关孤儿受体（RORs）转录活性以及反向激动剂对其活性影响的检测方法。一种检测方法是在Tet-On细胞系统中测量反向激动剂对RORs介导的荧光素酶报告基因转录激活的影响。哺乳动物双杂交检测法分析ROR配体结合结构域与共激活肽之间的相互作用。另外两种检测方法分别是检测反向激动剂对ROR靶基因IL17启动子控制下的荧光素酶报告基因激活的影响，以及使用哺乳动物单杂交检测法检测反向激动剂对ROR介导的激活作用的影响。
