Structure & Function of the Human PON1 Polymorphism

描述(由申请人提供)：拟议研究的主要目标是了解在人类对氧磷酶(PON1)基因变体中观察到的功能差异的结构基础。我们已经部分地描述了五个编码区多态和五个上游调控多态的影响。Q192R多态影响PON1对多种底物的催化效率，而-108C/T多态影响PON1的表达。三个新发现的多态导致PON1等位基因无效。我们的目标是了解这些多态的功能基因组学。我们将使用PON1小鼠模型来检查急性暴露于有机磷化合物(OP)的影响。小鼠模型包括PON1基因敲除小鼠和表达每个人类PON1-Q192R等位基因的一个或多个副本的小鼠。这个模型系统将允许我们检验这样的假设，即在给定水平表达的多态PON1-Q192R等位基因将对由PON1直接水解或通过细胞色素P450/PON1途径代谢的环境毒物提供特定程度的抗性。该模型还将使我们能够调查PON1基因多态在调节低水平OP暴露对大脑基因表达影响方面的影响，这是一个以前从未接近过的研究领域。我们建议研究PON1基因变异的五个方面，这将为人类PON1基因多态的功能基因组学提供信息。特定目的1检测急性暴露二氮磷(DZO)、毒死蜱(CPO)和毒死蜱(CPS)对PON1基因已被一个或两个拷贝的人PON1 Q192或R192替换的小鼠的毒理学效应。特定目的2检测PON192变异对暴露于CPO和DZO的PON1-192转基因小鼠脑内受影响亚区/细胞中基因毒理基因组表达谱的调节作用。具体目标3旨在确定因接触CPO而修饰的新蛋白质靶标。具体目标4包括确定人PON1的晶体结构和鉴定具有更高OP水解率的重组PON1变异体。具体目标5涉及利用人源化的PON1基因敲除小鼠表达不同水平的每个人PON1-192亚型，建立有机磷暴露的人类药代动力学/药效学(PDPK/PD)模型。

DESCRIPTION (provided by applicant): The major objective of the proposed research is to understand the structural basis for the functional differences observed in human paraoxonase (PON1) genetic variants. We have partially characterized the effects of five coding region polymorphisms and five upstream regulatory polymorphisms. The Q192R polymorphism affects the catalytic efficiency of PON1 for hydrolysis of a number of substrates, while the position -108C/T polymorphism affects PON1 expression. Three newly discovered polymorphisms result in inactive PON1 alleles. Our goal is to understand the functional genomics of these polymorphisms. We will use a PON1 mouse model to examine the effects of acute exposure to organophosphorus (OP) compounds. The mouse model includes PON1 knockout mice as well as knockout mice expressing one or more copies of each human PON1-Q192R allele. This model system will allow us to test the hypothesis that a polymorphic PON1-Q192R allele expressed at a given level will provide a specific degree of resistance to the toxicity of environmental agents hydrolyzed directly by PON1 or metabolized through the cytochrome P450 /PON1 pathway. The model will also allow us to investigate the effects of PON1 polymorphisms in modulating the effects of low level OP exposure on gene expression in the brain, an area of research that has not been previously approachable. We propose to investigate five facets of PON1 genetic variability that will provide information on the functional genomics of the polymorphisms of the human PON1 gene. Specific aim 1 examines the toxicological effects of acute exposures of diazoxon (DZO), chlorpyrifos oxon (CPO) and chlorpyrifos (CPS) on mice whose PON1 genes have been replaced with one or two copies of human PON1 Q192 or R192.Specific aim 2 examines the modulating effects of the PON192 variants on the toxicogenomic expression profile of genes in the affected subregions/cells of brains of PON1-192 transgenic mice exposed to CPO and DZO. Specific aim 3 is aimed at identifying new protein targets modified by exposure to CPO. Specific aim 4 involves determination of the crystal structure of human PON1 and characterization of recombinant PON1 variants with increased efficiency of OP hydrolysis. Specific aim 5 involves development of a human pharmacokinetic/pharmacodynamic (PDPK/PD) model of organophosphate exposure using humanized transgenic PON1 knockout mice expressing varying levels of each human PON1-192 isoform.