Project 5: Phytorermediation of Pollutants Using Transgenic Plants

接触有毒挥发性有机化合物（VOC）继续对人类健康构成重大风险。 挥发性有机化合物，如氯仿、四氯化碳（CT）和三氯乙烯（TCE），沿着苯和 其它单环芳族化合物是水和空气中最常见的污染物。这些化合物 对哺乳动物肝细胞色素P450 2 E1的氧化具有敏感性。在过去的NIEHS资助 在UW工作，我们已经证明了TCE和CT被无菌白杨细胞培养物氧化， 研究表明，野生型白杨能够吸收和降解TCE和CT。 我们的工作使人们认识到，野生型植物对TCE和CT的吸收太弱， 降低其在根区水中的浓度，为植物的发育提供动力， 增加对VOC的降解活性。为此，我们表达了哺乳动物细胞色素 P450 2 E1（CYP 2 E1）在植物中，实现数量级更大的TCE氧化转基因白杨。 目前我们正在进行野生型杨树和转CYP 2 E1基因白杨对三氯乙烯的田间降解试验 能够进行质量平衡测量的试验台设施中的天平。在本申请中，我们提出 以下具体目的：比较野生型和现有的CYP 2 E1转基因白杨品系的能力， 在现场试验中降解TCE、CT和PCE;识别因污染物存在而上调的基因; 克隆和分析植物中降解三氯乙烯及其代谢产物的相关基因， 在CYP 2 E1转基因白杨中TCE和苯的代谢产物;通过 增加叶面开放气孔数，表达有机磷羟化酶， 对氧磷酶PON 1，用于降解植物中的有机磷类神经毒素;并测定其生态毒性 对蚯蚓暴露的CYP 2 E1转基因植物的根材料、叶凋落物和根际土壤 VOC。

Exposure to toxic volatile organic compounds (VOCs) continues to pose a significant risk to human health. VOCs such as chloroform, carbon tetrachloride (CT) and trichloroethylene (TCE), along with benzene and other single-ring aromatics are among the most common contaminants in water and air. These compounds share a susceptibility to oxidation by the mammalian hepatic cytochrome P450 2E1. In past NIEHS-funded work at UW, we have demonstrated that TCE and CT are oxidized by axenic poplar cell cultures using a pathway similar to that in mammals, and that wild-type poplar are able to take up and degrade TCE and CT. Our work has led to the realization that TCE and CT uptake by wild-type plants is too weak to significantly reduce their concentration in root zone water, providing motivation for the development of plants with increased degradative activity toward VOCs. Toward this end we have expressed mammalian cytochrome P450 2E1 (CYP2E1) in plants, achieving orders of magnitude greater oxidation of TCE in transgenic poplar. We are presently engaged in testing TCE degradation by wild-type and CYP2E1 transgenic poplar at field scale in a test bed facility capable of mass balance measurements. In this application we propose the following specific aims: compare the abilities of wild-type and existing CYP2E1 transgenic poplar lines to degrade TCE, CT, and PCE in field tests; identify genes that are up-regulated by the presence of pollutants; clone and analyze the plant genes involved in the degradation of TCE and its metabolites; determine the metabolites of TCE and benzene in CYP2E1 transgenic poplar; increase the uptake of VOCs from the air by increasing the number of open stomata on leaf surfaces; express the organophosphorus hydroxylase, paraoxonase PON1, in plants for the degradation of organophosphate neurotoxins; and determine the ecotoxicity to earthworms of root materials, leaf litter and rhizosphere soil of CYP2E1 transgenic plants exposed to VOCs.