Odorant Receptor Expression and Sensitivity to Odorants

描述（由申请人提供）：该申请建议允许在嗅上皮分子生物学方面经验丰富的研究者跨越他在基于微阵列的基因组学方面的专业知识与更具计算挑战性的下一代测序方法之间的差距。该计划包括：(1)R编程培训，使研究者能够利用ChIP-seq和RNA-seq技术的灵活分析方法，(2)指导这两种方法的试点实验，以及(3)指导数据分析。试点研究利用了一种独特的资源，允许纯化表达气味受体亚群的嗅觉神经元，以测试关于气味受体的两个基本假设。(1)活性染色质修饰（只能通过富集表达气味受体的神经元来检测）对气味受体的严格调控表达至关重要。活性染色质修饰将通过ChIP-seq测量。嗅觉感受器表达的精细特异性控制可能是嗅觉感觉神经元功能中最大的未解之谜。(2)气味剂激活了一组气味受体，这些受体可能重叠，但必须至少有一个不同的成员，才能区分气味剂。通过活性依赖基因位点表达GFP标记的嗅觉感觉神经元的富集，将允许RNA-seq方法识别由特定气味剂激活的气味受体集。了解数百种气味受体如何检测和区分数千种气味化学物质的基本问题，将有助于解释嗅觉减退和嗅觉缺失的一些原因；并应允许未来发展的气味激动剂和拮抗剂，可用于改善人类的生活质量。

DESCRIPTION (provided by applicant): The application proposes to allow an investigator experienced in the molecular biology of the olfactory epithelium to cross the gap from his expertise in microarray-based genomics to more computationally challenging next generation sequencing approaches. The plan includes: (1) training in R programming to allow the investigator take advantage of flexible analysis methods for ChIP-seq and RNA-seq techniques, (2) mentored pilot experiments in both of these approaches, and (3) mentored analyses of the data. The pilot studies take advantage of a unique resource that allows purification of olfactory neurons expressing subsets of odorant receptors to work toward testing two fundamental hypotheses about odorant receptors. (1) Active chromatin modifications (which can only be detected by enrichment for neurons expressing an odorant receptor) are critical for the tightly regulated expression of odorant receptors. Active chromatin modifications will be measured by ChIP-seq. The exquisitely specific control of odorant receptor expression is perhaps the greatest remaining mystery about the function of olfactory sensory neurons. (2) Odorants activate sets of odorant receptors that may overlap but must have at least one distinct member if the odorants can be discriminated. Enrichment for olfactory sensory neurons marked by expression of GFP from an activity-dependent gene locus will allow RNA-seq methods to identify sets of odorant receptors activated by specific odorants. Understanding the fundamental question of how hundreds of odorant receptors allow the detection and discrimination of thousands of odorant chemicals should help explain some causes of hyposmia and anosmia; and should allow the future development of odorant agonists and antagonists that could be used to improve the quality of human life.