Project 4: Biotransformation Gene-Environment Interactions in Coho Salmon

美国西部的太平洋鲑鱼数量明显下降。特别令人关注的是，接触某些农药和微量金属的鲑鱼会出现亚致命性神经损伤。这些行为影响包括捕食者探测和猎物选择的丧失，繁殖时间的改变和家园的丧失。上述在个体中观察到的神经行为效应现在与群体影响联系在一起。鲑鱼的嗅觉系统是环境化学物质神经毒性的敏感目标，包括超级基金中常见的金属和杀虫剂。然而，人们对鱼类的化学嗅觉神经毒性机制知之甚少。我们第一个融资周期的研究产生了重要的发现，这些发现将在我们的竞争性更新中详细探讨。具体来说，我们知道：1)鲑鱼的嗅觉组织是化学生物转化的重要场所，特别是细胞色素P4503A和黄素单加氧酶（FMO）似乎介导了有机磷酸盐生物转化的组织特异性和化合物特异性差异，并可能对神经毒性产生影响；2)模型超级基金有机磷酸盐（氯吡虫啉）和金属（铜）的嗅觉损伤涉及嗅觉信号转导通路的破坏。然而，铜主要影响g蛋白偶联嗅觉受体信号，可能通过氧化应激，而毒死蜱激活参与嗅觉信号转导抑制的基因，并且3)转录特征可以帮助我们识别与混合物相关的独特基因靶标，以及区分金属和有机磷驱动的影响。根据我们的研究结果，竞争更新的目标是：1)利用cDNA克隆、重组蛋白表达、微阵列分析和酶学方法确定嗅觉CYP3A和黄素单加氧酶在鲑鱼从淡水向咸水迁移过程中有机磷神经毒性中的作用；2)利用原位杂交、免疫组织化学分析结合行为学研究了解氧化应激在铜和镉介导的嗅觉损伤中的作用。3)利用蛋白质组学方法鉴定和区分铜和毒死蜱的重要嗅觉蛋白靶点；4)利用上述研究产生的一套嗅觉生物标志物来评估通过超级基金站点迁移的鲑鱼的亚致死嗅觉神经毒性。

Pacific salmon populations have declined markedly in the Western United States. Of particular concern has been sublethal neurological injury occurring in salmon exposed to certain pesticides and trace metals. These behavioral impacts include loss of predator detection and prey selection, altered reproductive timing and loss of homing. These aforementioned neurobehavioral effects observed in individuals are now linked to population impacts. The salmon olfactory system is a sensitive target for the neurotoxicity of environmental chemicals, including metals and pesticides commonly found in Superfund sites. However, little is known about the mechanisms of chemical olfactory neurotoxicity in fish. Studies from our first funding cycle have produced important findings that will be explored in detail in our competitive renewal. Specifically, we know that: 1) the olfactory tissues of salmon are important site of chemical biotransformation, and in particular, cytochrome P4503A and flavin monooxygenases (FMO) appear to mediate tissue- and compound-specific differences in organophosphate biotransformation with potential impacts on neurotoxicity, 2) the olfactory injury by a model superfund organophosphate chemical (chlorpyrifos) and metal (copper) involves disruption of olfactory signal transduction pathways. However, copper primarily impacts G-protein coupled olfactory receptor signaling, likely through oxidative stress, whereas chlorpyrifos activates genes involved in the inhibition of olfactory signal transduction, and 3) transcriptional signatures can help us identify unique gene targets relevant to mixtures, as well differentiating metal- and organophosphate-driven affects. Based upon our findings, the objectives of the competing renewal are to: 1) use cDNA cloning, recombinant protein expression, microarray analysis and enzymatic approaches to determine the role of olfactory CYP3A and flavin monooxygenases in organophosphate neurotoxicity in salmon during movement from freshwater to saltwater, 2) use in situ hybridization and immunohistochemistry analyses coupled with behavioral studies to understand the role of oxidative stress in copper and cadmium-mediated olfactory injury, 3) use proteomics approaches to identify and discriminate important olfactory protein targets of copper and chlorpyrifos, 4) use a suite of olfactory biomarkers generated from the aforementioned studies to assess sublethal olfactory neurotoxicity in salmon migrating through Superfund sites.