Timed mRNA translation events in neocortical development and neurodevelopmental disorders

新皮质发育和神经发育障碍中的定时 mRNA 翻译事件

• 批准号: 8930467
• 负责人: Matthew Lee Kraushar
• 金额: $ 3.97万
• 依托单位: RBHS-ROBERT WOOD JOHNSON MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-08-28 至 2019-08-27
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8930467
• 关键词: Animal Model; Behavior; Brain; Cell Cycle; Cell Differentiation process; Cells; Cognitive deficits; Complex; Data; Density Gradient Centrifugation; Development; Disease; Elongation Factor; Embryo; Etiology; Event; FMRP; Fragile X Syndrome; Functional disorder; Gene Expression; Gene Expression Profile; Genes; Genetic Translation; Genomics; Health; Human; Immunohistochemistry; Lead; Measures; Messenger RNA; Meta-Analysis; Mitotic; Molecular; Molecular Target; Mus; Neocortex; Neurodevelopmental Disorder; Neuronal Differentiation; Neurons; Pathway interactions; Peptide Elongation Factor 2; Phosphorylation; Phosphotransferases; Polyribosomes; Predisposition; Protein Biosynthesis; Protein Dynamics; Proteins; Proteome; Regulation; Reverse Transcriptase Polymerase Chain Reaction; Ribosomal Proteins; Ribosomes; Role; Specificity; Staging; Stem cells; Sucrose; Susceptibility Gene; Syndrome; Techniques; Testing; Thymidine; Time; Transcript; Translating; Translations; Western Blotting; Work; autism spectrum disorder; base; cognitive function; disorder subtype; fetal; in vivo; microdeletion; neocortical; nerve stem cell; neurogenesis; neuronal circuitry; new therapeutic target; progenitor; small hairpin RNA; stem cell differentiation; stem cell fate; therapeutic target; transcriptomics; vector control

 DESCRIPTION (provided by applicant): Within the brain's neocortex originates our most evolutionarily advanced complex cognitive functions. The diverse subpopulations of neural stem cells and post-mitotic neurons underlying these intricate circuits are molecularly defined by their patterns of gene expression, where functional gene expression culminates in protein synthesis via mRNA translation at the level of the ribosome. Therefore, mRNA translation must be tightly regulated in development for specific mRNAs to generate molecularly defined subpopulations of neocortical neurons. Autism Spectrum Disorders (ASDs) are clinically heterogeneous disorders of complex cognitive functions. In the past decade, genomic and transcriptomic analyses have implicated genes and pathways that converge on protein synthesis in the cortex as a target in ASD etiology, reinforced by multiple syndromic ASD subtypes where translation abnormalities are central, such as Fragile-X Syndrome (FXS). While the regulation of core translation components has been implicated in ASDs, how abnormal translation of specific mRNAs leads to neocortical dysfunction remains unanswered. Furthermore, it is not known if ribosomal components are dynamic during neocortical development, nor whether this relates to how neocortical neurons are molecularly defined from the translation of specific mRNA transcripts. Our preliminary data suggest that components of the core translation machinery and the mRNAs that associate with them in the fetal neocortex are indeed dynamic, with a particular transition occurring at mid- neurogenesis - a susceptibility period highly implicated in ASD genomic meta-analyses. Our data show that this mid-neurogenic transition is marked by a dramatic increase in the phosphorylation of eukaryotic elongation factor 2 (eEF2), putatively modulating elongation activity via its kinase, eEF2k. Interestingly, eEF2k has been associated with ASDs as a variable genomic locus in multiple studies, and a unique microdeletion syndrome spanning this locus is characterized by severe cognitive deficits. eEF2k also partners with FMRP to regulate the translation of specific mRNAs in FXS. We found that dynamic eEF2 phosphorylation occurs in both cycling neural stem cells and differentiated neurons spanning neocorticogenesis, and loss of eEF2k results in abnormal neocortical neuron differentiation. We hypothesize that eEF2k regulates mRNA translation, neural stem cell cycling, and differentiation in fetal neocortical development and dysfunction. This will be tested by first analyzing the dynamic candidate protein components and mRNA cargo of actively translating ribosomes (polysomes) identified by our preliminary studies in eEF2k KO and WT neocortices throughout development. Second, neural stem cell cycle and differentiation will be analyzed in developing eEF2k KO vs. WT neocortices to extend our preliminary finding of abnormal neocortical circuits with eEF2k depletion. This project aims to advance our understanding of ASDs from previous studies of the genomic and transcriptomic levels towards an ASD proteome, employing advanced in vivo techniques to identify specific translational therapeutic targets.

 描述（由适用提供）：在大脑的新皮层内部，我们最先进的复杂认知功能。这些错综复杂的电路的神经元细胞和有序神经元的潜水员亚群是由它们的分子定义的 基因表达的模式，其中功能基因表达通过核糖体水平的mRNA翻译在蛋白质合成中达到顶点。因此，对于特定mRNA，必须在发育中严格调节mRNA翻译，以产生分子定义的新皮质神经元亚群。自闭症谱系障碍（ASD）是复杂认知功能的临床异质性疾病。在过去的十年中，基因组和转录组分析已经实施了基因和途径，这些基因和途径在皮质中融合了蛋白质合成作为ASD病因的靶标，并由多种综合综合症ASD亚型加强，其中翻译异常是中心的，例如脆弱的-X综合征（FXS）。尽管ASD中已经实施了核心翻译成分的调节，但特定mRNA的异常翻译如何导致新皮质功能障碍。此外，尚不清楚核糖体成分在新皮质发育过程中是否是动态的，也不知道这是否与新皮质神经元的分子如何从特定mRNA转录物的翻译中定义。我们的初步数据表明，在胎儿新皮层中与它们相关的核心翻译机制的组成部分确实是动态的，并且在神经中间发生时发生了特定的过渡 - 易感性周期在ASD基因组荟萃分析中高度涉及。我们的数据表明，这种神经发生的转变标志着真核伸长因子2（EEF2）的磷酸化急剧增加，通过其激酶EEF2K推测的伸长活性。有趣的是，在多项研究中，EEF2K与ASD相关联基因组基因座，而跨越该基因座的独特微骨骼综合征的特征是严重的认知缺陷。 EEF2K还与FMRP合作，以调节FXS中特定mRNA的翻译。我们发现，动态EEF2磷酸化发生在循环神经元细胞和跨越新皮质生成的分化神经元中，EEF2K的丧失导致新皮质神经元的异常分化。我们假设EEF2K调节mRNA翻译，神经干细胞循环以及胎儿新皮质发育和功能障碍的分化。这将通过首先分析我们在EEF2K KO和WT Neofortices的初步研究确定的积极翻译核糖体（多聚疾病）的动态候选蛋白成分和mRNA货物的测试。其次，将在开发EEF2K KO与WT Neotortices中分析神经干细胞周期和分化，以扩展我们对EEF2K耗竭异常的新皮质回路的初步发现。该项目旨在将我们对ASD的理解从先前对基因组和转录水平的研究转移到ASD蛋白质组，采用先进的体内技术来识别特定翻译的治疗靶标。