Rewired m6A methylation of promoter antisense RNAs in Alzheimer’s disease regulates neuronal gene expression in the 3D nucleome

N6-methyladenosine (m6A) is an abundant internal RNA modification that can impact gene expression at both post-transcriptional and transcriptional levels. However, the landscapes and functions of m6A in human brains and neurodegenerative diseases, including Alzheimer's disease (AD), are under-explored. Here, we examined RNA m6A methylome using total RNA-seq and meRIP-seq in middle frontal cortex of post-mortem brains from individuals with or without AD, which revealed m6A alteration on both mRNAs and various noncoding RNAs. Notably, many promoter-antisense RNAs (paRNAs) displayed cell-type-specific expression and changes in AD, including one produced adjacent to MAPT that encodes the Tau protein. MAPT-paRNA is highly expressed in neurons, and m6A positively controls its expression. In iPSC-derived human excitatory neurons, MAPT-paRNA does not impact the nearby MAPT mRNA, but instead promotes expression of hundreds of neuronal and synaptic genes, and is protective against excitotoxicity. Analysis of single nuclei RNA-DNA interactome in human brains supports that brain paRNAs interact with both cis- and trans-chromosomal target genes to impact their transcription. These data reveal landscapes and functions of noncoding RNAs and m6A in brain gene regulation and AD pathogenesis. we first characterized RNA m6A methylome from individuals diagnosed with AD and from cognitively normal age-matched counterparts individuals. We uncovered a large number of m6A-modified RNAs in the human brain, including both mRNAs and multiple categories of ncRNAs, many of which are previously unannotated. RNA m6A methylome in AD displays alteration and correlation with expression levels of mRNA and ncRNAs

N6-甲基腺嘌呤(N6-methyladenosine, m6A)是一种广泛存在的内部RNA修饰，可在转录后与转录水平调控基因表达。然而，m6A在人类大脑及包括阿尔茨海默病(Alzheimer's disease, AD)在内的神经退行性疾病中的分布图谱与功能仍有待深入探索。本研究通过总RNA测序（total RNA-seq）与甲基化RNA免疫沉淀测序（meRIP-seq）技术，对伴或不伴阿尔茨海默病的死者大脑额中皮层组织开展RNA m6A甲基化组分析，结果揭示了信使RNA（mRNA）与多种非编码RNA（ncRNA）上的m6A修饰改变。值得关注的是，大量启动子反义RNA（promoter-antisense RNAs, paRNAs）呈现细胞类型特异性表达模式，并在阿尔茨海默病中发生表达变化，其中包括1种在编码Tau蛋白的MAPT基因邻近区域生成的paRNA。MAPT相关paRNA在神经元中高表达，且m6A对其表达具有正向调控作用。在诱导多能干细胞（induced pluripotent stem cell, iPSC）分化得到的人类兴奋性神经元中，MAPT-paRNA并未影响邻近的MAPT mRNA表达，反而可促进数百种神经元与突触相关基因的表达，并对兴奋性毒性起到保护作用。对人类大脑单细胞核RNA-DNA互作组的分析证实，大脑中的paRNAs可通过与顺式及反式染色体靶基因互作来调控其转录。本研究揭示了非编码RNA与m6A修饰在大脑基因调控及阿尔茨海默病发病机制中的分布图谱与功能。我们首先对确诊阿尔茨海默病的患者以及年龄匹配的认知正常对照个体的RNA m6A甲基化组进行了表征，在人类大脑中发现了大量携带m6A修饰的RNA，既包括mRNA，也涵盖多类ncRNA，其中许多为此前未被注释的RNA分子。阿尔茨海默病中的RNA m6A甲基化组存在修饰异常，并与mRNA及ncRNA的表达水平相关。