QTL AND MICROARRAY MAPPING LEAD SENSITIVITY GENES

描述（由申请人提供）：人类活动导致许多有毒物质在环境中分布，其中重金属在我们的生物圈中传播。除了对接触铅（即铅涂料或受污染的饮用水）的人类产生急性毒性影响外，慢性接触对所有生物体的发育还有更阴险的影响。接触低水平铅的儿童会改变发育过程，这些儿童会出现多动、感觉功能改变和认知能力（“智商”）改变等症状。果蝇是研究发育过程中铅暴露影响的有希望的模式生物，因为(1)对其遗传学的复杂理解，以及易于操纵其基因组；(2)对极低剂量铅的微小影响敏感的行为学和形态学分析的可用性。人们认为铅暴露的敏感性存在很大差异，而且人类和果蝇细胞都被认为在接触铅时诱导“保护基因”的表达。这项提议的假设是，人们可以利用数量性状位点（QTL）定位技术，结合微阵列和复杂的遗传分析，识别出一些“保护基因”，这些基因使生物体抵抗铅毒性的行为和发育影响。为了验证这一假设，目的1是通过将90个重组近交系果蝇的幼虫暴露于环境相关剂量的铅中来绘制铅敏感QTL，并分析受铅影响的几个行为和突触连接方面，如幼虫运动、成虫学习、成虫运动、成虫运动与乙醇蒸汽暴露以及幼虫神经肌肉连接的结构。目的2是对来自重组自交系的整个幼虫和成虫头部的标记mRNA进行DNA微阵列分析，这些自交系显示出铅诱导的行为改变的最大差异，如目的1所确定的那样。目标3是通过使用现有的突变和RNA干扰技术，通过条件过表达或下调在目标1和2中进行的检测中重叠的基因，并确定铅对这些基因改变果蝇的认知、运动和突触功能的影响。这些研究的结果将确定在人类铅暴露期间发生改变的最重要基因的候选基因，并很可能导致达到或低于NOAEL和LOAEL值的重金属暴露的生物测定或治疗。

DESCRIPTION (provided by applicant): Human activity has resulted in the environmental distribution of many toxic substances, among them the heavy metals that are spread throughout our biosphere. In addition to the acute toxic effects to humans exposed to lead (i.e. in lead paint or in contaminated drinking water) there are more insidious effects of chronic exposure on the development of all organisms. Children exposed to low levels of lead have altered developmental processes, and these children develop symptoms such as hyperactivity, changes in sensory function, and changes in cognitive abilities ("IQ"). Drosophila is a promising model organism to study the effects of lead exposure during development because of (1) the sophisticated understanding of its genetics, and the ease of manipulating its genome; (2) the availability of behavioral and morphological assays sensitive to small effects of very low doses of lead. There is thought to be a great deal of variability in the sensitivity of lead exposure, and both human and Drosophila cells are thought to induce expression of "protective genes" upon exposure to lead. The hypothesis for this proposal is that one can identify some of the "protective genes" that make an organism resistant to the behavioral and developmental effects of lead toxicity using quantitative trait loci (QTL) mapping techniques combined with microarray and sophisticated genetic analyses. To test this hypothesis, Aim 1 is to map lead sensitive QTL by exposing larvae of 90 recombinant inbred Drosophila strains to environmentally relevant doses of lead, and assaying several behavioral and synaptic connection aspects that are affected by lead - such as larval locomotion, adult learning, adult locomotion, adult locomotion combined with ethanol vapor exposure, and the structure of the larval neuromuscular junction. Aim 2 is to perform DNA microarray analyses with labeled mRNA from whole larvae and adult heads from the recombinant inbred lines that show the greatest differences in lead-induced behavioral alterations, as determined in Aim 1. Aim 3 is to up-regulate, by conditional over-expression, or down-regulate, by using existing mutations and RNA interference techniques, the genes that overlap in the assays performed in Aims 1 and 2, and to determine the effects of lead on cognition, locomotion and synaptic function of these genetically altered flies. Results of these studies will identify candidates for the most important genes that are altered during lead exposure in humans, and could well lead to bioassays or treatments for heavy metal exposure at or below NOAEL and LOAEL values.