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

项目摘要/摘要 芳香烃受体(AHR)在许多化合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)和多环芳香烃(PAHs)。有一件事-- 对这些化合物的影响的敏感度存在个体差异，但其机制尚不清楚。 人类AHR序列的变异不能完全解释个体对AHR配体的反应差异， 这表明AHR途径的其他组成部分参与了对敏感性的控制。最新的基因组- 对对多氯联苯、TCDD和多环芳烃具有进化抗性的鱼类种群进行的水平研究已经确定了AHR- 相互作用蛋白(AIP)作为候选抗性基因。AIP是AHR的监护人，它影响 AHR的稳定性和核转位，但其确切作用尚不清楚。在人类中，AIP的突变 易患家族性孤立性垂体腺瘤(FIPA)的证据表明，AIP序列变异 功能后果。AIP变异是否会改变体内AHR激动剂的敏感性 不知道。这项基础研究的目的是阐明AIP及其序列变体在 控制对各种AHR激动剂的敏感性，包括环境污染物和天然AHR 配基。中心假设是AIP基因座上的变异影响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。 这一假设将通过对斑马鱼(Danio Rerio)活体和人类的互补性研究来验证。 体外培养的细胞。在目标1中，使用CRISPR-Cas9基因组编辑产生的AIP-空斑马鱼将被用于 确定AIP在控制对发育毒性和基因表达改变的敏感性中的作用 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及其变异体，包括与FIPA相关的突变影响AHR的分子机制 功能。在目标3中，有针对性地将AIP SNP敲入斑马鱼将被用来确定如何在 AIP蛋白在体内影响对不同AHR配体的敏感性。这项拟议的研究代表了一项独特的 利用野生鱼类种群环境暴露的洞察力以及机械原理的机会 在人类细胞和工程斑马鱼胚胎中进行的研究，以了解 对通过AHR起作用的化学物质的敏感性存在个体差异。这些研究针对的是 NIEHS 2018-2023年战略计划，包括参与调解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.