Genetic Analysis of Olfaction

描述(申请人提供)：昆虫化学传感系统与脊椎动物化学传感系统有许多相似之处，但在数值上更简单。果蝇强大的分子遗传学工具与简单的化学感觉行为分析和电生理分析相结合，使我们能够确定化学感觉系统各个组件的体内生物功能。最终，这些结果将提供对化学感觉信息处理的基本原理的见解，这些原理可能与脊椎动物共享。我们提出了以下具体目标： 具体目的1.阐明对雄性信息素11-顺式痘苗病毒醋酸酯敏感的周边受体事件和分子组成。在上一个资助周期中，我们发现气味结合蛋白LUSH是一小部分化学感觉神经元对信息素敏感所必需的。值得注意的是，Lush似乎是嗅觉神经元的直接激活器。我们将确定LUSH是否直接与11-顺式痘苗烯基乙酸酯结合，并使用靶向RNAi方法和遗传方法鉴定由LUSH激活的神经元受体。我们将确切地确定信息素、LUSH和嗅觉神经元受体是如何共同作用来调节信息素信号的。 具体目的2.将果蝇触角的功能外周嗅觉图与解剖图相关联。我们将使用反向遗传方法将特定气味受体基因产物的表达与嗅觉神经元的功能类别相关联。这将使我们能够将解剖外周嗅觉图与功能图合并，并建立在体内对天线中表达的每个气味感受器的气味特异性。这些信息最终将有助于理解大脑如何将混合气味转化为有意义的气味图像。我们将使用由我的实验室首次描述的转基因RNAi方法来靶向单个气味受体的表达，并使用单感受器电生理记录来识别转基因果蝇中的缺陷功能类别。 具体目的3.使用反向遗传方法阐明在触角中表达的其他OBP蛋白的作用。我们已经在果蝇基因组中发现了35个编码OBP蛋白的基因。16种气味结合蛋白在触角中表达。是所有这些蛋白质都参与了信息素反应，还是有些蛋白质参与了对一般气味的检测？我们将使用反向遗传学来消除这些蛋白质在天线中的表达，并确定嗅觉神经元的功能类别是否存在缺陷，或者交配或聚集行为是否存在缺陷。

DESCRIPTION (provided by applicant): The insect chemosensory system has many parallels with vertebrate chemosensory systems, but is numerically simpler. The potent molecular genetic tools available in Drosophila combined with simple chemosensory behavior assays and electrophysiological analysis allow us to determine the in vivo biological function of individual components of the chemosensory system. Ultimately these results will provide insights into underlying principles of chemosensory information processing that are likely to be shared with vertebrates. We propose the following specific aims: Specific Aim 1. Elucidate the perireceptor events and molecular components underlying sensitivity to the male pheromone, 11-cis vaccenvl acetate. In the last funding cycle we discovered the odorant binding protein LUSH is required for pheromone sensitivity in a small subset of chemosensory neurons. Remarkably, LUSH appears to function as a direct activator of olfactory neurons. We will determine whether LUSH binds directly to 11-cis vaccenyl acetate, and identify the neuronal receptor that is activated by LUSH using a targeted RNAi approach and a genetic approach. We will establish exactly how the pheromone, LUSH and the olfactory neuron receptor function together to mediate pheromone signaling. Specific Aim 2. Correlate the functional peripheral olfactory map to the anatomic map in the Drosophila antenna. We will correlate expression of specific odorant receptor gene products with functional classes of olfactory neurons using a reverse genetic approach. This will allow us to merge the anatomic peripheral olfactory map with the functional map and establish the in vivo odorant specificity for each odorant receptor expressed in the antenna. This information will ultimately help understand how the brain converts odorant blends into meaningful odor images. We will use a transgenic RNAi approach first described by my laboratory to target expression of individual odorant receptors and identify defective functional classes in the transgenic flies using single-sensillum electrophysiological recordings. Specific Aim 3. Use a Reverse Genetic Approach to elucidate the role of the other OBP proteins expressed in the antenna. We have identified 35 genes encoding OBP proteins in the Drosophila genome. 16 odorant binding proteins are expressed in the antenna. Are all of these proteins mediating pheromone responses or are some involved in detection of general odorants? We will use reverse genetics to eliminate expression of these proteins in the antenna and determine if there are defects in functional classes of olfactory neurons or in mating or aggregation behavior.