Identification of gene targets for morphine & other drugs of abuse

吗啡靶基因的鉴定

基本信息

批准号：
7452428
负责人：
RICHARD H. GOODMAN
金额：
$ 28.62万
依托单位：
OREGON HEALTH & SCIENCE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2005
资助国家：
美国
起止时间：
2005-09-28 至 2010-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7452428
关键词：
Actins Acute Address Adenylate Cyclase Affect Aftercare Amphetamines Antisense RNA Attention Behavior Behavioral Binding Binding Sites Bioinformatics Biological Assay CREB1 gene Cells Chromatin Chronic Class Cocaine Code Complement Custom Cyclic AMP Cytoskeleton DNA Dependence Drug Addiction Drug effect disorder Elements Enhancers Exposure to Forskolin Gene Expression Gene Expression Profile Gene Targeting Genes Genomics Goals Growth Human Laboratories Link Longevity Measures Mediating Mediator of activation protein Methodology Methods MicroRNAs Modeling Modification Morphine Morphology N-Methylaspartate Nature Nerve Growth Factor Receptors Neurites Neuroblastoma Neurons Opiates PC12 Cells Pathway interactions Personal Satisfaction Phenocopy Phosphorylation Population Process Proteins RNA Regulation Research Personnel Ribonucleotides Second Messenger Systems Sequence Analysis Signal Transduction Standards of Weights and Measures Testing Transcript Translations addiction chromatin immunoprecipitation concept design drug of abuse in vivo inhibitor/antagonist novel strategies programs promoter receptor coupling research study response rho second messenger serial analysis of gene expression transcription factor

项目摘要

DESCRIPTION (provided by applicant): Long term changes in gene expression are believed to contribute importantly to the mechanisms underlying drug addiction. Considerable attention has been devoted to the cAMP second messenger pathway because it is upregulated by opiates and other drugs of abuse. Both tolerance and dependence have been attributed to changes in function of the transcription factor CREB, which mediates cAMP-dependent gene expression. It is currently believed that CREB binds constitutively to a promoter element termed the CRE. This proposal challenges this model and tests a new hypothesis-that chronic exposure to morphine induces CREB binding to some genes but not others. To address this hypothesis the lab has developed a novel approach termed SACO for examining CREB binding to target genes in vivo. This method combines chromatin immunoprecipitation with a modification of Long SAGE (an approach designed for analysis of mixtures of RNA). SACO will be used to identify the entire complement of CREB targets and measure how the selection of these targets is affected by agents, such as morphine, that upregulate the cAMP pathway. Additional studies will address the mechanisms underlying the morphological changes in dendritic processes induced by CREB that occur after treatment with other drugs of abuse, namely cocaine and amphetamine. Specific goals of this project are to identify the entire set of CREB targets in human neuroblastoma cells and determine whether this set is altered by acute exposure to cAMP or chronic exposure to morphine. Previous studies have indicated that CREB regulates the expression of both protein-coding and noncoding transcripts. Many protein-coding transcripts and most noncoding transcripts are missing from conventional microarrays, however. Studies in this proposal will examine both classes of RNAs by developing a custom microarray representing the CREB transcriptome. One specific microRNA, designated miR132, was found to be induced by CREB in PC12 cells and to stimulate changes in dendritic morphology in neurons that are highly reminiscent of those caused by CREB activators. A candidate target for this miRNA has been identified and experiments are designed to elucidate the mechanism of miR132 action. The concept that CREB signaling induces mediators of translational arrest has profound implications for the understanding of drug action.

描述（由申请人提供）：基因表达的长期变化被认为对药物成瘾的机制有重要贡献。 cAMP 第二信使途径受到了相当多的关注，因为它会被阿片类药物和其他滥用药物上调。耐受性和依赖性均归因于转录因子 CREB 功能的变化，CREB 介导 cAMP 依赖性基因表达。目前认为 CREB 与称为 CRE 的启动子元件组成型结合。这一提议挑战了这一模型并测试了一个新的假设——长期接触吗啡会诱导 CREB 与某些基因结合，但不会与其他基因结合。为了解决这一假设，该实验室开发了一种称为 SACO 的新方法，用于检查 CREB 与体内靶基因的结合。该方法将染色质免疫沉淀与 Long SAGE 的改良（一种设计用于分析 RNA 混合物的方法）相结合。 SACO 将用于识别 CREB 靶标的全部补充，并测量上调 cAMP 通路的药物（例如吗啡）如何影响这些靶标的选择。其他研究将探讨用其他滥用药物（即可卡因和安非他明）治疗后 CREB 诱导的树突状过程形态变化的机制。该项目的具体目标是鉴定人神经母细胞瘤细胞中的整套 CREB 靶标，并确定该组靶标是否因急性暴露于 cAMP 或慢性暴露于吗啡而改变。先前的研究表明 CREB 调节蛋白质编码和非编码转录本的表达。然而，传统微阵列中缺少许多蛋白质编码转录本和大多数非编码转录本。该提案中的研究将通过开发代表 CREB 转录组的定制微阵列来检查这两类 RNA。一种名为 miR132 的特定 microRNA 被发现在 PC12 细胞中被 CREB 诱导，并刺激神经元树突形态的变化，这与 CREB 激活剂引起的变化高度相似。该 miRNA 的候选靶点已确定，并设计实验来阐明 miR132 的作用机制。 CREB 信号传导诱导翻译停滞介质的概念对于理解药物作用具有深远的影响。