Comparative Biology Elucidation of Environmental Pathways and Susceptibility

描述（由申请人提供）： 许多人类疾病，如亨廷顿氏病、帕金森氏病和阿尔茨海默氏病，以及正常的生理状态，如衰老，都受到暴露于环境压力的强烈影响。然而，很少有人知道的机制，环境压力的检测和区分彼此在动物。本工作的长期目标是利用功能基因组学方法在线虫C。elegans来定义基本的环境压力传感和信号机制。这些研究将更准确地确定环境在人类健康和疾病中的作用，并有助于确定许多有毒物质的作用方式。在这个提议中要检验的具体假设是，隔室特异性蛋白质损伤构成了区分不同形式的环境应激的反馈抑制机制。该假说基于以下观察结果：1）通常防止某些形式的蛋白质损伤的积累的蛋白质稳态（PH）基因的敲低激活对蛋白质敏感的基因表达; 2）激活对蛋白质敏感的基因表达的PH基因的敲低加速蛋白质聚集; 3）由热和氧化应激诱导的蛋白质损伤不激活对蛋白质敏感的基因表达。本研究的具体目标是：1）确定受环境应激因子转录调控的PH基因。我们将研究PH基因在响应环境应激中的空间和时间以及功能参与，使用i）野生型和转录因子突变动物中热、氧化和免疫诱导的基因表达的比较全基因组微阵列分析，ii）暴露于环境应激期间GFP标记的PH基因的体内定位，iii）过表达PH靶标的转基因动物的应激存活测定。2)测试蛋白质错误折叠作为应激诱导基因表达的激活剂的作用。蛋白质损伤将使用特定PH复合物的RNAi敲低来遗传诱导。每个基因敲低对热、氧化和渗透应激信号传导途径的激活的影响将通过以下方式定量：i）表达应激特异性GFP报告基因的动物的体内高通量荧光测量，ii）使用应激感应转录因子突变体的上位性分析，iii）PH敲低动物中的整个动物存活测定。

DESCRIPTION (provided by applicant): Many human diseases, such as Huntington's, Parkinson's, and Alzheimer's, and normal physiological states, such as aging, are strongly influenced by exposure to environmental stress. However, little is known about the mechanisms by which environmental stressors are detected and discriminated from one another in animals. The long term objective of this work is to utilize functional genomic approaches in the nematode C. elegans to define fundamental environmental stress sensing and signaling mechanisms. Such studies will more precisely define the role of the environment in human health and disease and help to define the modes of action of numerous toxic substances. The specific hypothesis to be tested in this proposal is that compartment specific protein damage constitutes a feedback inhibition mechanism for discrimination between different forms of environmental stress. This hypothesis is based on the observations that 1) knockdown of protein homeostasis (PH) genes that normally prevent the accumulation of some forms of protein damage activates osmosensitive gene expression 2) Knockdown of PH genes that activate osmosensitive gene expression accelerates protein aggregation 3) Protein damage induced by heat and oxidative stress does not activate osmosensitive gene expression. The specific aims of this proposal are: 1) Define the PH genes that are transcriptionally regulated by environmental stressors. We will examine the spatial and temporal, and functional involvement of PH genes in response to environmental stress using i) comparative whole genome microarray analysis of heat, oxidative, and osmotically induced gene expression in wild type and transcription factor mutant animals ii) in vivo localization of GFP tagged PH genes during exposure to environmental stress iii) stress survival assays in transgenic animals overexpressing PH targets. 2) Test the role of protein misfolding as an activator of stress induced gene expression. Protein damage will be genetically induced using RNAi knockdown of specific PH complexes. The effects of each gene knockdown on the activation of heat, oxidative, and osmotic stress signaling pathways will be quantified by i) in vivo high throughput fluorescence measurements of animals expressing stress-specific GFP reporters ii) epistasis analysis using stress sensing transcription factor mutants iii) whole animal survival assays in PH knockdown animals.