Genome-Wide 5-Formylcytosine Redistribution in KCl-Stimulated Mouse Primary Cortical Neurons is Associated with Neuronal Activity
Genome-Wide 5-Formylcytosine Redistribution in KCl-Stimulated Mouse Primary Cortical Neurons is Associated with Neuronal Activity
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DOI：
10.1021/acschemneuro.3c00554
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发表时间：
2023-11-29
影响因子：
5
通讯作者：
Li，Xiang
中科院分区：
文献类型：
--
作者：
Wu，Du;Huang，Kaixin;Li，Xiang
在哺乳动物神经元中发现了大量的DNA去甲基化中间体。虽然5-甲基胞嘧啶（5mC）和5-羟甲基胞嘧啶（5hmC）在神经元功能中的作用已被广泛研究，但对5-甲酰胞嘧啶（5fC）在神经元中的作用知之甚少。因此，本研究旨在探讨5fC在神经元中的全基因组分布和潜在功能。在小鼠原代皮质神经元体外培养模型中，我们观察到氯化钾刺激后神经元基因组中总5fC水平的动态增加。随后，我们采用化学标记支持的c -t转换测序（cleverseq）在单碱基分辨率下检查5fC分布。生物信息学分析显示5fC在启动子区域富集，基因本体（GO）分析表明差异甲酰化位置（DFP）与神经元活动相关。此外，结合先前发表的新生RNA-seq数据显示，基因甲酰化与mRNA表达水平呈正相关。同时，证实了6个神经活动相关基因呈正相关。此外，通过多组学分析，我们在5fC位点附近观察到更高的染色质可及性和RNA pol II结合信号。基序分析确定了5fC的潜在解读蛋白。总之，我们的工作为研究5fC在激活的哺乳动物神经元中的动态变化和功能作用提供了宝贵的资源。
An abundant accumulation of DNA demethylation intermediates has been identified in mammalian neurons. While the roles of 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) in neuronal function have been extensively studied, little is known about 5-formylcytosine (5fC) in neurons. Therefore, this study was to investigate the genome-wide distribution and potential functions of 5fC in neurons. In an in vitro culture model of mouse primary cortical neurons, we observed a dynamic increase in the total 5fC level in the neuronal genome after potassium chloride (KCl) stimulation. Subsequently, we employed chemical-labeling-enabled C-to-T conversion sequencing (CLEVER-seq) to examine the 5fC distribution at a single-base resolution. Bioinformatic analysis revealed that 5fC was enriched in promoter regions, and gene ontology (GO) analysis indicated that the differential formylation positions (DFP) were correlated with neuronal activities. Additionally, integration with previously published nascent RNA-seq data revealed a positive correlation between gene formylation and mRNA expression levels. As well, 6 neuro-activity-related genes with a positive correlation were validated. Furthermore, we observed higher chromatin accessibility and RNA pol II binding signals near the 5fC sites through multiomics analysis. Motif analysis identified potential reader proteins for 5fC. In conclusion, our work provides a valuable resource for studying the dynamic changes and functional roles of 5fC in activated mammalian neurons.