Structure & Function of the Human PON1 Polymorphism

描述（由申请人提供）：拟议研究的主要目的是了解在人类对氧磷酶（PON1）遗传变异中观察到的功能差异的结构基础。我们已经部分表征了5个编码区多态性和5个上游调控多态性的影响。Q192R多态性影响PON1对多种底物水解的催化效率，而-108C/T位置多态性影响PON1的表达。三种新发现的多态性导致PON1等位基因失活。我们的目标是了解这些多态性的功能基因组学。我们将使用PON1小鼠模型来检查急性暴露于有机磷（OP）化合物的影响。小鼠模型包括PON1敲除小鼠以及表达每个人类PON1- q192r等位基因的一个或多个拷贝的敲除小鼠。该模型系统将允许我们验证这样的假设，即在给定水平上表达的多态性PON1- q192r等位基因将对由PON1直接水解或通过细胞色素P450 /PON1途径代谢的环境因子的毒性提供特定程度的抗性。该模型还将使我们能够研究PON1多态性在调节低水平OP暴露对大脑基因表达的影响中的作用，这是一个以前未涉足的研究领域。我们建议研究PON1遗传变异的五个方面，这将为人类PON1基因多态性的功能基因组学提供信息。特异性目的1研究了急性暴露于重氮唑（DZO）、毒死蜱（CPO）和毒死蜱（CPS）对PON1基因被人类PON1 Q192或R192的一个或两个拷贝所取代的小鼠的毒理学影响。特异性目的2研究了PON192变异对暴露于CPO和DZO的PON1-192转基因小鼠大脑受影响亚区/细胞中基因的毒理学表达谱的调节作用。特异性目标3旨在鉴定暴露于CPO修饰的新蛋白靶点。具体目标4包括确定人类PON1的晶体结构，并通过提高OP水解效率来表征重组PON1变体。具体目标5涉及使用表达不同水平的人类PON1-192亚型的人源化转基因PON1敲除小鼠，建立人类有机磷暴露的药代动力学/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.