RNA Control of Neural Function

RNA 控制神经功能

基本信息

批准号：
10622122
负责人：
Joel D Richter
金额：
$ 42.72万
依托单位：
UNIV OF MASSACHUSETTS MED SCH WORCESTER
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-05-01 至 2028-02-29
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10622122
关键词：
3&apos Untranslated Regions Ablation Affect Alternative Splicing Animal Behavior Area Behavioral Binding Brain Chromatin Codon Nucleotides Cognition Cytoplasm Electrophysiology (science)Epigenetic Process Event Excision FMR1 FMRP Fragile X Syndrome Functional disorder Genes Genetic Transcription Growth Health Human Knockout Mice Laboratories Lead Learning Length Link Mediating Memory Messenger RNA Minority Molecular Mus Neurodegenerative Disorders Neurodevelopmental Disorder Neurophysiology - biologic function Nucleotides Poly A Poly(A) Tail Polyadenylation Post-Transcriptional Regulation Protein Isoforms Proteins RNA RNA Degradation RNA Splicing RNA Stability RNA-Binding Proteins Regulation Research Resolution Ribosomes Synapses Synaptic plasticity Translation Initiation Translational Regulation Translations Work autism spectrum disorder experimental study mRNA Precursor mRNA Translation mouse model nanopore neuropathology neuroregulation postsynaptic ribosome profiling synaptic function transcriptome sequencing

项目摘要

Our work focuses on post-transcriptional control of neural function with emphasis on translational control of synaptic plasticity and learning and memory. We investigate the neurodevelopmental and neurodegenerative disorders that arise when this translation goes awry. We find that mis-regulated translation leads to changes in alternative splicing and RNA degradation, which in turn contribute to neuropathology. More specifically, our research is comprised of three distinct but complementary areas: (1) CPEB1 and cytoplasmic polyadenylation control of translation; (2) FMRP regulation of translation with emphasis on ribosome stalling and codon usage; (3) FMRP regulation of alternative splicing. CPEB1-regulated cytoplasmic polyadenylation governs translation in post-synaptic compartments, which in turn modifies synaptic strength, the underlying cellular basis of learning and memory. Molecular, electrophysiological, and behavioral experiments from our laboratory have demonstrated that CPEB1 regulates activity-dependent cytoplasmic polyadenylation-induced translation, which in turn modifies synaptic strength, and cognition. CPEB1 nucleates several proteins that promote poly(A) tail growth and removal and mediate translation initiation; they also regulate plasticity and animal behavior. Another RNA binding protein important for brain function is FMRP, the product of the Fragile X Syndrome gene FMR1. FMRP binds >1000 RNAs in the brain and regulates translation, primarily by stalling ribosome translocation on specific mRNAs. One such mRNA encodes the epigenetic factor SETD2, which catalyzes the chromatin mark H3K36me3. In FMRP-deficient mouse brain, SETD2 levels are elevated and the H3K36me3 chromatin landscape, which is principally located in gene bodies, is disrupted. H3K36me3 is linked to alternative pre-mRNA splicing and there is widespread mis-regulation of splicing in FMRP knockout (KO) mice. The main objectives our research going forward will address key unanswered questions regarding RNA regulation of neural function, primarily using mouse models: (a) CPEB1-deficiency rescues Fragile X pathophysiology in FMRP KO mice. Does this rescue involve ribosome stalling and/or polyadenylation? (b) CPEB1 also regulates 3’UTR length. What is the mechanism by which this occurs? (c) How does FMRP stall ribosomes on specific mRNAs? Does FMRP act as a molecular roadblock to ribosome transit and/or does FMRP interact with the ribosome? (d) How does FMRP regulate alternative splicing? Some of the mis-splicing events appear to involve H3K36me3, but these are in the minority. Does FMRP regulate the translation of mRNAs encoding splicing factors, and/or does FMRP, which is a shuttling protein, affect splicing directly? (e) How does FMRP employ codon optimality to regulate translation and RNA stability? We will address these issues by ribosome profiling, which we modified to distinguish between translocating and stalled ribosomes, and direct nanopore RNA sequencing, which yields poly(A) tail size at near-nucleotide resolution as well as RNA isoforms that arise by alternative splicing. We will deplete key regulatory factors from the mouse brain and assess synaptic function and animal behavior.

我们的工作重点是神经功能的转录后控制，重点是神经功能的翻译控制我们研究神经发育和神经退行性病变。当这种翻译出现问题时就会出现紊乱。我们发现，错误的翻译会导致变化。选择性剪接和 RNA 降解，这反过来又有助于神经病理学。研究由三个不同但互补的领域组成：(1) CPEB1 和细胞质多腺苷酸化翻译控制；(2) FMRP 翻译调控，重点是核糖体停滞和密码子使用； (3) CPEB1 调节的选择性剪接的 FMRP 调节控制翻译。在突触后区室中，这反过来又改变了突触强度，这是学习的细胞基础我们实验室的分子、电生理和行为实验。 CPEB1 调节活性依赖性的细胞质多聚腺苷酸化诱导的翻译，这反过来，CPEB1 会改变突触强度，并且使多种蛋白质成核，从而促进聚腺苷酸尾巴的形成。生长和去除以及介导翻译起始；它们还调节可塑性和动物行为。对大脑功能很重要的 RNA 结合蛋白是 FMRP，它是脆性 X 综合征基因 FMR1 的产物。 FMRP 在大脑中结合超过 1000 个 RNA 并调节翻译，主要是通过阻止核糖体易位一种这样的 mRNA 编码表观遗传因子 SETD2，它催化染色质标记。 H3K36me3 在 FMRP 缺陷的小鼠大脑中，SETD2 水平升高，并且 H3K36me3 染色质主要位于基因体中的景观被破坏，与替代的前 mRNA 相关。 FMRP 敲除 (KO) 小鼠中普遍存在剪接错误调节。我们未来的研究将解决有关 RNA 调节神经功能的关键未解答问题，主要使用小鼠模型：(a) CPEB1 缺陷可挽救 FMRP KO 小鼠中的脆性 X 病理生理学。这种拯救是否涉及核糖体停滞和/或多聚腺苷酸化？(b) CPEB1 也调节 3’UTR 长度。发生这种情况的机制是什么？(c) FMRP 如何阻止特定 mRNA 上的核糖体？ FMRP 充当核糖体转运的分子障碍和/或 FMRP 是否与核糖体相互作用？ FMRP 是否调控选择性剪接？一些错误剪接事件似乎涉及 H3K36me3，但是这些只是少数。FMRP 是否调节编码剪接因子的 mRNA 的翻译，和/或确实如此。 FMRP 是一种穿梭蛋白，直接影响剪接（e）FMRP 如何利用密码子最优性来实现剪接？调节翻译和 RNA 稳定性？我们将通过核糖体分析来解决这些问题，我们对其进行了修改区分易位核糖体和停滞核糖体，以及直接纳米孔 RNA 测序，从而产生我们将在近核苷酸分辨率下分析聚 (A) 尾部大小以及通过选择性剪接产生的 RNA 亚型。消除小鼠大脑中的关键调节因子并评估突触功能和动物行为。