School of Medicine, Jacqui Wood Cancer Centre, Ninewells Hospital and Medical School, University of Dundee, Dundee, United Kingdom (M.M., S.D., L.A.J., C.J.H., C.R.W.) and Preclinical Safety, Translational Medicine, Novartis Institutes for BioMedical Research, Basel, Switzerland (R.T., M.-A.G., A.V., J.M.).
School of Medicine, Jacqui Wood Cancer Centre, Ninewells Hospital and Medical School, University of Dundee, Dundee, United Kingdom (M.M., S.D., L.A.J., C.J.H., C.R.W.) and Preclinical Safety, Translational Medicine, Novartis Institutes for BioMedical Research, Basel, Switzerland (R.T., M.-A.G., A.V., J.M.)
Mol Pharmacol. 2019 Jan;95(1):97-105. doi: 10.1124/mol.118.113555. Epub 2018 Oct 25.
The constitutive androstane receptor (CAR) is a xenobiotic sensor expressed in hepatocytes that activates genes involved in drug metabolism, lipid homeostasis, and cell proliferation. Much progress has been made in understanding the mechanism of activation of human CAR by drugs and xenobiotics. However, many aspects of the activation pathway remain to be elucidated. In this report, we have used viral constructs to express human CAR, its splice variants, and mutant CAR forms in hepatocytes from mice in vitro and in vivo. We demonstrate CAR expression rescued the ability of hepatocytes to respond to a wide range of CAR activators including phenobarbital. Additionally, two major splice isoforms of human CAR, CAR2 and CAR3, were inactive with almost all the agents tested. In contrast to the current model of CAR activation, ectopic CAR1 is constitutively localized in the nucleus and is loaded onto gene in the absence of an inducing agent. In studies to elucidate the role of threonine T38 in CAR regulation, we found that the T38D mutant was inactive even in the presence of CAR activators. However, the T38A mutant was activated by CAR inducers, showing that T38 is not essential for CAR activation. Also, using the inhibitor erlotinib, we could not confirm a role for the epidermal growth factor receptor in CAR regulation. Our data suggest that CAR is constitutively bound to gene regulatory regions and is regulated by exogenous agents through a mechanism which involves protein phosphorylation in the nucleus.
组成型雄烷受体（CAR）是一种在肝细胞中表达的外源性传感器，可激活参与药物代谢、脂质稳态和细胞增殖的基因。在理解药物和外源性物质激活人 CAR 的机制方面已经取得了很大进展。然而，激活途径的许多方面仍有待阐明。在本报告中，我们使用病毒构建体在体外和体内表达了来自 小鼠的肝细胞中的人 CAR、其剪接变体和突变型 CAR 形式。我们证明 CAR 表达挽救了 肝细胞对包括苯巴比妥在内的广泛 CAR 激活剂的反应能力。此外，人 CAR 的两种主要剪接异构体 CAR2 和 CAR3，几乎对所有测试的药物均无活性。与当前的 CAR 激活模型相反，外源性 CAR1 持续定位于细胞核中，并且在没有诱导剂的情况下被加载到 基因上。在阐明 CAR 调节中苏氨酸 T38 作用的研究中，我们发现即使存在 CAR 激活剂，T38D 突变体也无活性。然而，T38A 突变体被 CAR 诱导剂激活，表明 T38 对于 CAR 激活不是必需的。此外，使用抑制剂厄洛替尼，我们无法确认表皮生长因子受体在 CAR 调节中的作用。我们的数据表明，CAR 与基因调节区域持续结合，并通过一种涉及核内蛋白磷酸化的机制被外源性试剂调节。
