G-Protein Coupled Receptor Signaling: a Role for Small Interfering RNAs in Stimulus Specific Adaptation

G 蛋白偶联受体信号转导：小干扰 RNA 在刺激特异性适应中的作用

基本信息

批准号：
9764313
负责人：
Trang Huyen Duong
金额：
$ 5万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-09-01 至 2021-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9764313
关键词：
Affect Afferent Neurons Animals Behavioral Assay Benzaldehyde Biochemical Biological Models Butanones Caenorhabditis elegans Cells Chronic Clinical Complex Cyclic GMP-Dependent Protein Kinases Data Dental Drug Design Drug Targeting Excision Exonuclease Exoribonucleases Exposure to G Protein-Coupled Receptor Signaling G-Protein-Coupled Receptors GTP-Binding Proteins Genes Goals Hydrocodone Individual Intervention Lead Libraries Mechanics Mediating Messenger RNA Methadone Modeling Mutation Nematoda Neurobiology Neurons Odors Olfactory Pathways Oxycodone Pain management Pathway interactions Pharmaceutical Preparations Population Process Production RNA Interference Resolution Ribonucleases Role Second Messenger Systems Signal Pathway Signal Transduction Signal Transduction Pathway Signaling Protein Small Interfering RNA Specificity Stimulus System Testing Visual Work addiction cohort desensitization drug discovery drug market follow-up insight isopentyl alcohol member mutant olfactory sensory neurons public health relevance response side effect therapy development

项目摘要

DESCRIPTION (provided by applicant): A significant portion of currently approved drugs target G Protein Coupled Receptors (GPCRs) but most have off-target effects and many of these include dental sequelae. Such side effects can arise when distinct GPCRs in one cell signal through similar second messengers. Thus, the stimulation of one GPCR affects the down- stream signaling pathway and adaptation of the cell's other GPCRs. Indeed, one key question in the field is how different GPCRs, though they activate many of the same second messengers, signal independently without cross adaptation of their signals. Understanding how this signaling is segregated may lead to the ability to develop interventions that limit crosstalk and off-target effects. My overarching goal is to understand how adaptation of GPCR pathways can be independently regulated. This question will be studied in the C. elegans model system, where signaling from multiple GPCRs can be studied in individual olfactory neurons. Our lab recently discovered that an endogenous RNAi (endo-siRNA) pathway promotes adaptation to GPCR signaling by odorants. This observation provides a framework to explain how multiple odors sensed by a single sensory neuron are adapted to independently. This may thus constitute one mechanism by which highly similar signal transduction pathways are insulated from each other in the same cell. This leads me to the hypothesis that the specificity of adaptation to G protein signaling in olfactory neurons is directed by the engagement of different endogenous RNAi biosynthetic pathways acting downstream of olfactory GPCR stimulation. I will test this hypothesis in 3 aims: In aim 1, I will use behavioral assays to identify endogenous RNAi processing factors that act downstream of GPCRs to promote adaptation specifically to either isoamyl alcohol (IAA) or butanone (BU). Preliminary data suggests that the exonuclease ERI-1 is one such factor as it is required for adaptation only to IAA. In aim 2, I will use biochemical and visual means to determine whether ERI-1 is, in fact uniquely required for adaptation to IAA and if so, by what mechanism. In aim 3, I will use high-resolution endo-siRNA sequencing of either IAA or BU adapted populations to determine if specific siRNA species are produced in adaptation to each odor. I will then determine whether any of these sequences or removal of their targets are sufficient to promote IAA adaptation. Together, these results will provide insight into how GPCR signaling can remain separated within the same cell, and how endogenous siRNA mediates this process.

描述（由适用提供）：当前批准的药物中很大一部分靶向G蛋白偶联受体（GPCR），但大多数具有脱靶效应，其中许多包括牙齿后遗症。当通过类似的第二使者在一个单元信号中不同的GPCR中，可以产生这种副作用。这是一个GPCR的刺激会影响细胞其他GPCR的下游信号通路和适应。确实，该领域的一个关键问题是，尽管GPCR激活了许多相同的第二使者，但它们如何独立地发出信号的信号。了解该信号的分离方式可能会导致发展限制串扰和脱离目标效应的干预措施的能力。我的总体目标是了解如何独立调节GPCR途径的适应性。这个问题将在秀丽隐杆线虫模型系统中进行研究，其中来自多个GPCR的信号可以在单个嗅觉神经元中研究。我们的实验室最近发现，内源性RNAi（Endo-SiRNA）途径促进了气味剂对GPCR信号的适应。该观察结果提供了一个框架，可以解释单个感觉神经元如何独立调整单个感觉神经元的多种气味。因此，这可能构成一种机制，通过该机制，高度相似的信号转导途径在同一细胞中彼此隔离。这导致了以下假设：嗅觉神经元中对G蛋白信号的适应性的特异性是由嗅觉GPCR刺激下游作用的不同内源性RNAi生物合成途径的参与指导的。我将在3个目标中检验这一假设：在AIM 1中，我将使用行为测定来识别GPCR下游作用的内源性RNAI加工因子，以促进对异氧氨基醇（IAA）或丁烷（BU）的适应性。初步数据表明，外核酸酶ERI-1是仅适应IAA所需的一个因素。在AIM 2中，我将使用生化和视觉手段来确定ERI-1是否是通过哪种机制适应IAA的独特所需的。在AIM 3中，我将使用IAA或BU拟合种群的高分辨率内部 - 核NA测序来确定是否会适应每种气味的特定siRNA物种。然后，我将确定这些序列中的任何一个或删除其目标是否足以促进IAA适应。这些结果将共同洞悉GPCR信号如何在同一细胞内保持分离，以及内源性siRNA如何培养此过程。