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对氧磷酶（PON1）遗传变异体中观察到的功能差异的结构基础。我们已经部分表征了五个编码区多态性和五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位多态性影响PON1表达。三个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 等位基因一个或多个拷贝的敲除小鼠。该模型系统将使我们能够测试以下假设：在给定水平表达的多态性 PON1-Q192R 等位基因将对直接由 PON1 水解或通过细胞色素 P450 /PON1 途径代谢的环境物质的毒性提供特定程度的抵抗力。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方面的影响，这是一个以前无法实现的研究领域。我们建议研究 PON1 遗传变异性的五个方面，这将提供有关人类 PON1 基因多态性的功能基因组学信息。具体目标 1 检查急性暴露重氮磷 (DZO)、毒死蜱 (CPO) 和毒死蜱 (CPS) 对 PON1 基因已被一两个拷贝的人 PON1 Q192 或 R192 拷贝替换的小鼠的毒理学影响。具体目标 2 检查 PON192 变体对受影响亚区域/细胞中基因毒理学表达谱的调节作用暴露于 CPO 和 DZO 的 PON1-192 转基因小鼠的大脑。具体目标 3 旨在确定通过接触 CPO 修饰的新蛋白质靶标。具体目标 4 涉及确定人 PON1 的晶体结构以及具有更高 OP 水解效率的重组 PON1 变体的表征。具体目标 5 涉及使用表达不同水平的每种人类 PON1-192 同种型的人源化转基因 PON1 敲除小鼠开发有机磷暴露的人类药代动力学/药效 (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.