The role of linear ubiquitination in memory formation

线性泛素化在记忆形成中的作用

基本信息

批准号：
9977384
负责人：
TIMOTHY JOSEPH JAROME
金额：
$ 19.12万
依托单位：
VIRGINIA POLYTECHNIC INST AND ST UNIV
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-02-05 至 2021-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9977384
关键词：
Adult Amygdaloid structure Binding Biological Biological Process Brain C-terminal Cell Nucleus Cells Clustered Regularly Interspaced Short Palindromic Repeats Complex Cytoplasm Data Development Enzymes Fright Functional disorder Future Genes Genetic Transcription Goals Head Human In Vitro Knowledge Learning Literature Lysine Mass Spectrum Analysis Mediating Memory Memory Disorders Memory impairment Mental disorders Methods Modification Molecular N-terminal NF-Kappa B p65 Neurodegenerative Disorders Neurons Nuclear Pattern Physiological Polyubiquitin Polyubiquitination Post-Translational Protein Processing Process Proteins Research Role Small Interfering RNA Symptoms Synapses Synaptic plasticity System Tail Technology Testing Transcription Process Transcriptional Regulation Translational Research Ubiquitin Ubiquitination Work conditioned fear design experimental study fear memory genome analysis in vivo insight long term memory memory consolidation memory process memory retention multicatalytic endopeptidase complex nervous system disorder next generation novel therapeutics p65 protein degradation protein purification research study sharpin transcription factor transcriptome sequencing ubiquitin mediated proteasome degradation whole genome

项目摘要

Project Summary/Abstract The broad goal of this proposal is to understand how memories are formed and stored in the brain. Pavlovian fear conditioning has proven to be a useful paradigm in elucidating the molecular mechanisms underlying memory formation and storage in cells. Indeed, good evidence now exists suggesting that memories formed using this paradigm require dynamic post-translational modification of proteins in neurons. Recently, evidence has emerged demonstrating that protein ubiquitination is critically involved in memory formation in the brain, primarily through targeting proteins for degradation by the proteasome. However, much remains unknown about how other ubiquitin marks which are independent of the proteasome are involved in the memory storage process. In our preliminary studies, we found that in the amygdala fear learning increased the levels of linear ubiquitinated proteins, an atypical ubiquitin modification that is not targeted for degradation by the proteasome complex. Importantly, these increases occurred selectively in the nucleus, but not the cytoplasm or at synapses, and targeted the transcription factor p65, suggesting that linear ubiquitination may be involved in transcriptional control during memory formation. The work in this proposal is designed to answer important questions about whether linear ubiquitination is involved in gene transcription critical for fear memory formation in the amygdala. Using a combination of protein purification methods and mass spectrometry, Aim 1 will identify what proteins are targeted by linear ubiquitination following learning. Additionally, this aim will correlate these identified proteins with next generation RNA-seq data obtained following siRNA-mediated reductions in linear ubiquitination within the amygdala, which will allow an unbiased, whole genome analysis of how this ubiquitin mark is involved in transcriptional processes during the process of memory formation. Aim 2 directly tests the functional role of linear ubiquitination in memory formation by reducing or enhancing this ubiquitin mark in the amygdala via in vivo siRNA or CRISPR-dCas9 manipulations, respectively, and testing memory retention for a contextual fear conditioning task. Collectively, this research will provide critical information regarding how linear ubiquitination is involved in transcriptional regulation during memory formation and whether it is critical for learning-dependent synaptic plasticity. This could provide potentially useful information on how memories are stored in the brain which could have important implications for the treatment of memory impairments associated with a variety of psychiatric disorders.

项目摘要/摘要该提议的广泛目标是了解如何形成和存储在大脑中。事实证明，帕夫洛维亚恐惧调节是阐明分子机制的有用范式基础记忆形成和单元格中的存储。确实，现在存在充分的证据表明使用这种范式形成的记忆需要对神经元中蛋白质的动态翻译后修饰。最近，有证据表明蛋白质泛素化与记忆有关大脑形成，主要是通过靶向蛋白质降解蛋白酶体的。但是，很多对于独立于蛋白酶体的其他泛素标记如何与内存存储过程。在我们的初步研究中，我们发现在杏仁核中，恐惧学习增加了线性泛素化蛋白的水平，一种非典型的泛素修饰，不是降解的目标由蛋白酶体复合体。重要的是，这些增加发生在细胞核中，但没有细胞质或突触时，并针对转录因子p65，表明线性泛素化可能可能在记忆形成过程中参与转录控制。该提案的工作旨在回答关于线性泛素化是否参与基因转录至关重要的重要问题杏仁核中的形成。结合蛋白质纯化方法和质谱法，AIM 1 学习后，将确定哪些蛋白质是通过线性泛素化靶向的。此外，这个目标将将这些鉴定的蛋白与siRNA介导的下一代RNA-seq数据相关联杏仁核内线性泛素化的减少，这将允许对整个基因组分析在记忆形成过程中，该泛素标记如何参与转录过程。目标2 直接通过减少或增强它来直接测试线性泛素化在记忆形成中的功能作用杏仁核中的泛素标记分别通过体内siRNA或CRISPR-DCAS9操纵和测试上下文恐惧调节任务的记忆保留。总的来说，这项研究将提供关键关于在记忆形成期间，有关线性泛素化如何参与转录调节的信息以及这对于依赖学习的突触可塑性至关重要。这可能会提供潜在有用的有关记忆如何存储在大脑中的信息，这些记忆可能对治疗具有重要意义与各种精神疾病相关的记忆障碍。