Neuronal Activity-dependent Pomoter Usage

摘要 依赖经验的神经回路重塑使动物能够使它们的行为适应永远的 不断变化的环境。由感觉输入引起的神经元活性依赖的分子改变 从转录到翻译后修饰的水平是神经重新连接的基础。不足为奇的是， 神经元活动的分子反应受损是一系列认知障碍的共同特征 精神障碍，如抑郁症、精神分裂症和自闭症。因此，理解活性依赖的分子 改变对于理解这些精神障碍的机制至关重要。在过去几年中， 广泛的研究在描绘转录组活性依赖的变化方面取得了很好的进展， 选择性剪接和蛋白质组。这些研究为我们提供了关于大脑如何 整合感觉输入并重组特定的神经网络。 然而，RNA分子的一部分逃过了全基因组的审查--5‘端。广泛使用 RNA-Seq方法不能很好地代表RNA的5‘端。分析转录起始点的现有方法 然而，(TS)主要代表5‘端；因此，大多数转录组在这种情况下都不存在 文库，这使得将TSS的使用与动态mRNA表达联系起来具有挑战性。 我们最近开发了一种全长新生转录组图谱方法，Bru-seq-DLAF，它 结合敏感的TSS检测和新生RNA测序。通过Bru-Seq-DLAF，我们已经揭开了 在减少或增加网络活跃度时动态改变TS的数十个基因。这些TS中的大多数 导致其氨基末端的多肽发生变化。此外，蛋白质异构体是保守的。 表明依赖活动的TSS的使用可能代表了一种保守的新的 突触可塑性背后的基因调节层。提出的研究目标是：1)彻底 在小鼠模型中描述这一新现象，并2)确定活动依赖的TSS在 突触可塑性。我们假设依赖神经元活性的TSS通过以下方式控制突触可塑性 改变蛋白质的亚细胞定位。我们的团队独一无二地准备用它的 在基因组学和突触可塑性方面的统一专业知识。 拟议研究的一个积极结果是阐明了依赖活动的TSS选择是一种 突触可塑性的复杂分子机制。据我们所知，我们的数据代表了第一个基因组- 动物细胞中TSS变化对细胞外刺激的广泛表征。此外，许多基因 接受依赖活动的TSS选择与精神障碍有关，如精神分裂症 以及自闭症谱系障碍。因此，这项拟议的研究将改善精神障碍的遗传学和 指导未来疾病相关基因的功能研究。

ABSTRACT Experience-dependent remodeling of neural circuits enables animals to adapt their behavior to ever- changing environments. Neuronal activity-dependent molecular alterations, elicited by sensory inputs, at all levels from transcription to post-translational modifications are the basis of neural rewiring. Not surprisingly, impaired molecular responses to neuronal activity are a common hallmark of a broad range of cognitive disorders, such as depression, schizophrenia, and autism. Thus, comprehending activity-dependent molecular changes is crucial to understanding the mechanisms of these mental disorders. Over the past several years, extensive studies have made excellent strides in profiling activity-dependent changes in the transcriptome, alternative splicing, and proteome. These studies have provided significant new insights into how the brain integrates sensory inputs and reorganizes specific neural networks. However, a portion of the RNA molecule has escaped genome-wide scrutiny―the 5'-ends. Widely-used RNA-seq methods underrepresent the 5'-ends of RNAs. Existing methods to profile transcription start sites (TSS), however, predominantly represent the 5' ends; therefore, most of the transcriptome is absent in such libraries, making it challenging to connect TSS usage to dynamic mRNA expression. We recently developed a full-length nascent-transcriptome profiling method, Bru-seq-DLAF, which combines sensitive detection of TSS and nascent RNA sequencing. With Bru-seq-DLAF, we have unveiled tens of genes that dynamically change TSS upon reducing or increasing network activity. Most of these TSS led to alterations in polypeptides at their amino termini. Furthermore, the protein isoforms were conserved between mice and humans, indicating that activity-dependent TSS usage may represent a conserved new layer of gene regulation underlying synaptic plasticity. The proposed research goals are to 1) thoroughly characterize this novel phenomenon in mouse models and 2) determine the roles of activity-dependent TSS in synaptic plasticity. We hypothesize that neuronal activity-dependent TSS controls synaptic plasticity by changing the subcellular localization of proteins. Our team is uniquely poised to test this hypothesis with its unified expertise in genomics and synaptic plasticity. A positive outcome of the proposed study is to illuminate activity-dependent TSS selection as an intricate molecular mechanism for synaptic plasticity. To our knowledge, our data represent the first genome- wide characterization of TSS alterations upon extracellular stimuli in animal cells. Furthermore, many genes that undergo activity-dependent TSS selection have been implicated in mental disorders such as schizophrenia and autism spectrum disorders. Thus, the proposed study will improve the genetics of mental disorders and guide future functional studies on disease-associated genes.