Mechanisms Controlling Sensitivity and Resistance to Dioxin-like Compounds: Role of AIP

项目总结/摘要 芳香烃受体（AHR）在许多植物的毒性机制中起着重要作用。 化学污染物，包括氯化二恶英，如2，3，7，8-四氯二苯并对二恶英（TCDD）， 一些多氯联苯（PCBs）和多环芳烃（PAH）。有一个inter- 个体对这些化合物的敏感性存在差异，但其机制尚不清楚。 人类AHR序列的变异不能完全解释对AHR配体应答的个体差异， 这表明AHR途径的其他成分参与控制敏感性。最近的基因组- 对进化出对多氯联苯、四氯二苯并对二恶英和多环芳烃抗性的鱼类种群进行的水平研究已经确定了AHR- 互作蛋白（AIP）作为候选抗性基因。AIP是一种AHR伴侣， 稳定性和核转位的AHR，但其确切的作用是不明确的。在人类中，AIP的突变 易患家族性孤立性垂体腺瘤（FIPA），有证据表明AIP序列变异 功能性后果。AIP变异是否改变了体内对AHR激动剂作用的敏感性， 不知道。这项基础研究的目的是阐明AIP及其序列变异体在 控制对不同AHR激动剂的敏感性，包括环境污染物以及天然AHR 配体。核心假设是AIP基因座的变异影响AIP和AHR之间的相互作用， 导致对引起毒性的化学物质的敏感性改变，并通过AHR改变基因表达。 这一假设将使用涉及斑马鱼（Danio rerio）在体内和人类的补充研究进行检验。 体外细胞在目标1中，使用CRISPR-Cas9基因组编辑产生的AIP无效斑马鱼将用于 确定AIP在控制对发育毒性和改变的基因表达的敏感性中的作用 由体内不同的AHR激动剂引起。在目标2中，将使用人肝细胞系HepaRG来研究 AIP及其变体（包括与FIPA相关的突变）影响AHR的分子机制 功能在目标3中，将AIP SNP靶向敲入斑马鱼中以确定在斑马鱼中AIP SNP的变异如何。 AIP蛋白影响体内对不同AHR配体的敏感性。这项研究代表了一种独特的 利用野生鱼类种群沿着的环境暴露的见解， 研究人类细胞和工程斑马鱼胚胎，以了解基本机制， 对通过AHR起作用的化学物质的敏感性的个体差异。这些研究涉及 NIEHS 2018-2023战略计划，包括参与介导的分子途径的基础研究 环境暴露的影响、对发育过程的研究和理解机制 潜在的个体易感性。

Project Summary/Abstract The aryl hydrocarbon receptor (AHR) plays an essential role in the mechanisms of toxicity of numerous chemical contaminants, including chlorinated dioxins such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), some polychlorinated biphenyls (PCBs), and polynuclear aromatic hydrocarbons (PAHs). There is inter- individual variation in sensitivity to effects of these compounds, but the mechanisms are poorly understood. Variation in the human AHR sequence does not fully explain individual differences in response to AHR ligands, suggesting that other components of the AHR pathway are involved in controlling sensitivity. Recent genome- level research in fish populations with evolved resistance to PCBs, TCDD, and PAHs has identified the AHR- interacting protein (AIP) as a candidate resistance gene. AIP is an AHR chaperone that influences the stability and nuclear translocation of AHR, but its exact role is poorly defined. In humans, mutations in AIP predispose patients to familial isolated pituitary adenomas (FIPA), evidence that AIP sequence variation has functional consequences. Whether AIP variation alters the susceptibility to effects of AHR agonists in vivo is not known. The goal of this basic research is to elucidate the role of AIP and its sequence variants in controlling sensitivity to diverse AHR agonists, including environmental contaminants as well as natural AHR ligands. The central hypothesis is that variation at the AIP locus affects the interaction between AIP and AHR, leading to altered sensitivity to chemicals that cause toxicity and altered gene expression through the AHR. This hypothesis will be tested using complementary studies involving zebrafish (Danio rerio) in vivo and human cells in vitro. In Aim 1, AIP-null zebrafish generated using CRISPR-Cas9 genome-editing will be used to determine the role of AIP in controlling the sensitivity to developmental toxicity and altered gene expression caused by diverse AHR agonists in vivo. In Aim 2, the human liver cell line HepaRG will be used to investigate the molecular mechanisms by which AIP and its variants, including mutations associated with FIPA, affect AHR function. In Aim 3, targeted knock-in of AIP SNPs into zebrafish will be used to determine how variation in the AIP protein affects the sensitivity to diverse AHR ligands in vivo. The proposed research represents a unique opportunity to use insights from environmental exposures in wild fish populations along with mechanistic studies in human cells and engineered zebrafish embryos to understand fundamental mechanisms underlying individual differences in susceptibility to chemicals that act through the AHR. The studies address goals of the NIEHS 2018-2023 Strategic Plan, including basic research on molecular pathways involved in mediating effects of environmental exposures, research on developmental processes, and understanding mechanisms underlying individual susceptibility.