Molecular mechanisms of m6A mRNA modification in Drosophila neurogenesis

果蝇神经发生中 m6A mRNA 修饰的分子机制

• 批准号: 404953711
• 负责人: Professor Dr. Christoph Dieterich
• 金额: --
• 依托单位: Institut für Pharmazeutische und Biomedizinische Wissenschaften (IPBW)
• 依托单位国家: 德国
• 项目类别: Priority Programmes
• 财政年份: 2018
• 资助国家: 德国
• 起止时间: 2017-12-31 至 2021-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/404953711?language=en
• 关键词: Molecular; mechanisms; m6A; mRNA; modification

The field of RNA modifications, now termed Epitranscriptomics, has made major strides to the central stage of gene expression control in the recent years. Many modified nucleotides were first discovered in transfer RNAs and ribosomal RNAs in all studied organisms. Few of the same nucleotide modifications were later identified in messenger RNAs (mRNA) of eukaryotes, where they were recently revealed as an entire new layer of regulation of gene expression. The most prevalent and best-studied internal modification on mRNA is N6-methyladenosine (m6A). This modification was shown to regulate several physiological processes in various organisms via posttranscriptional control of gene expression. We recently characterized the m6A pathway in Drosophila melanogaster by identifying its main components and demonstrating its critical role during brain development. Accumulating evidences suggest that this function in the nervous system is conserved in vertebrates. However, due to the difficulty of studying m6A (and RNA modifications in general) in vivo, the molecular mechanisms by which m6A controls neurogenesis remains elusive. In this collaborative proposal, we want i) to develop new tools to map m6A RNA modification in vivo by a non-invasive RNA labeling approach (TRIBE) and by low input immunoprecipitation protocols and in quantitative manner ii) to explore the effect of m6A on gene expression in sub-populations of neuronal cells and iii) to identify and molecularly characterize m6A reader proteins, which are relevant to neuronal functions. We have generated promising results in regards to the identification of methylated transcripts in vivo using a genetic approach combined with high throughput sequencing and sophisticated computational analysis. Furthermore, we have delineated the cell types that are sensitive to the loss of m6A in the nervous system. Lastly our unpublished work suggests that additional m6A readers exist. We plan to characterize their physiological functions as well as their cross talks within the m6A pathway. While our work will be initially focused on m6A, we plan to later adapt the developed tools to additional modifications. The suggested cooperative study has the potential to foster several collaborations within the SPP, and to lead to a better understanding of the nascent field of mRNA modifications.

RNA修饰领域，现在被称为表观转录组学，近年来在基因表达控制的中心阶段取得了重大进展。在所有被研究的生物体中，许多修饰的核苷酸是在转移rna和核糖体rna中首次发现的。很少有相同的核苷酸修饰后来在真核生物的信使rna （mRNA）中被发现，它们最近被揭示为基因表达调控的一个全新层面。最普遍和研究最多的mRNA内部修饰是n6 -甲基腺苷（m6A）。这种修饰被证明通过基因表达的转录后控制来调节多种生物的几种生理过程。我们最近通过鉴定其主要成分并证明其在大脑发育过程中的关键作用，对黑腹果蝇的m6A通路进行了表征。越来越多的证据表明，神经系统的这种功能在脊椎动物中是保守的。然而，由于在体内研究m6A（以及一般的RNA修饰）的困难，m6A控制神经发生的分子机制仍然是难以捉摸的。在这项合作提案中，我们希望i)开发新的工具，通过非侵入性RNA标记方法（TRIBE）和低输入免疫沉淀协议，以定量的方式绘制体内m6A RNA修饰；ii)探索m6A对神经细胞亚群基因表达的影响；iii)鉴定和分子表征与神经元功能相关的m6A读取器蛋白。我们利用结合高通量测序和复杂计算分析的遗传方法在体内鉴定甲基化转录本方面取得了令人鼓舞的结果。此外，我们还描述了神经系统中对m6A缺失敏感的细胞类型。最后，我们未发表的工作表明，存在额外的m6A阅读器。我们计划表征它们的生理功能以及它们在m6A通路中的相互作用。虽然我们的工作最初将集中在m6A上，但我们计划稍后对开发的工具进行额外的修改。建议的合作研究有可能促进SPP内部的几次合作，并有助于更好地理解新兴的mRNA修饰领域。