Transcriptome-wide m6A Methylation Profiling of Wild Type and ALKBH5-/- Peritoneal Macrophages by m6A-seq

By performing m6A-seq analysis on the peritoneal macrophages that derived from ALKBH5-/- mice and littermate mice infected with or without vesicular stomatitis virus (VSV), we want to investigate whether ALKBH5 deficiency-mediated m6A RNA methylation contributes to the regulation of its target genes expression. m6A-seq analysis revealed enriched and specific m6A peaks on the transcript of ALKBH5-targeted gene, which were substantially increased in ALKBH5-deficient peritoneal macrophages than that in wild-type cells whatever infected with or without VSV. Meanwhile we didn’t observe up-regulation of m6A signal on VSV in ALKBH5-deficient cells; and also didn't find significant difference of m6A signals on IFN-β mRNA between ALKBH5-deficient and wild type cells whatever infected with or without VSV. These demonstrated that deficiency of ALKBH5 controls viral replication by increasing the m6A modification of ALKBH5 target gene to regulate its expression. Examination of m6A RNA modifications in wild type and ALKBH5-/- peritoneal macrophages infected with or without VSV, two biological replicates.

本研究通过对感染或未感染水泡性口炎病毒（vesicular stomatitis virus, VSV）的ALKBH5基因敲除（ALKBH5-/-）小鼠及其同窝野生型小鼠来源的腹腔巨噬细胞开展m6A测序（m6A-seq）分析，旨在探究ALKBH5缺失介导的m6A RNA甲基化是否参与调控其靶基因的表达。m6A测序分析结果显示，ALKBH5靶基因的转录本上存在富集且特异性的m6A修饰峰；无论是否感染VSV，ALKBH5缺失型腹腔巨噬细胞中的该峰信号均显著高于野生型细胞。与此同时，我们未在ALKBH5缺失细胞中观察到VSV转录本上的m6A信号上调；且无论是否感染VSV，ALKBH5缺失型与野生型细胞的干扰素-β（IFN-β）mRNA上的m6A信号均无显著差异。上述结果表明，ALKBH5缺失可通过增强ALKBH5靶基因的m6A修饰以调控其表达，进而控制病毒复制。本数据集涵盖感染或未感染VSV的野生型与ALKBH5-/-腹腔巨噬细胞的m6A RNA修饰检测数据，共设置2次生物学重复。