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有经验的研究者跨越差距，从他在基于微阵列的基因组学方面的专业知识到更具计算挑战性的下一代测序方法。该计划包括：（1）R编程培训，使研究人员能够利用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，（3）指导数据分析。试点研究利用了一种独特的资源，可以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）对于气味受体的严格调控表达至关重要。将通过ChIP-seq测量活性染色质修饰。气味受体表达的精确特异性控制可能是嗅觉感觉神经元功能的最大谜团。(2)气味剂激活气味剂受体的集合，这些集合可以重叠，但如果气味剂可以被区分，则必须具有至少一个不同的成员。通过从活性依赖性基因位点表达GFP标记的嗅觉感觉神经元的富集将允许RNA-seq方法来识别由特定气味激活的气味受体集。了解数百种气味受体如何允许检测和区分数千种气味化学物质的基本问题，应该有助于解释嗅觉减退和嗅觉丧失的一些原因;并应该允许未来开发可用于改善人类生活质量的气味激动剂和拮抗剂。

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.