Center for Medical Bioinformatics, Escola Paulista de Medicina, UNIFESP, São Paulo, Brazil.
Department of Microbiology, Immunology and Parasitology, Escola Paulista de Medicina, UNIFESP, São Paulo, Brazil.
Front Cell Infect Microbiol. 2022 Aug 16;12:906578. doi: 10.3389/fcimb.2022.906578. eCollection 2022.
The epitranscriptomics of the SARS-CoV-2 infected cell reveals its response to viral replication. Among various types of RNA nucleotide modifications, the m6A is the most common and is involved in several crucial processes of RNA intracellular location, maturation, half-life and translatability. This epitranscriptome contains a mixture of viral RNAs and cellular transcripts. In a previous study we presented the analysis of the SARS-CoV-2 RNA m6A methylation based on direct RNA sequencing and characterized DRACH motif mutations in different viral lineages. Here we present the analysis of the m6A transcript methylation of Vero cells (derived from African Green Monkeys) and Calu-3 cells (human) upon infection by SARS-CoV-2 using direct RNA sequencing data. Analysis of these data by nonparametric statistics and two computational methods (m6anet and EpiNano) show that m6A levels are higher in RNAs of infected cells. Functional enrichment analysis reveals increased m6A methylation of transcripts involved in translation, peptide and amine metabolism. This analysis allowed the identification of differentially methylated transcripts and m6A unique sites in the infected cell transcripts. Results here presented indicate that the cell response to viral infection not only changes the levels of mRNAs, as previously shown, but also its epitranscriptional pattern. Also, transcriptome-wide analysis shows strong nucleotide biases in DRACH motifs of cellular transcripts, both in Vero and Calu-3 cells, which use the signature GGACU whereas in viral RNAs the signature is GAACU. We hypothesize that the differences of DRACH motif biases, might force the convergent evolution of the viral genome resulting in better adaptation to target sequence preferences of writer, reader and eraser enzymes. To our knowledge, this is the first report on m6A epitranscriptome of the SARS-CoV-2 infected Vero cells by direct RNA sequencing, which is the RNA-seq.
SARS-CoV-2 感染细胞的表观转录组学揭示了其对病毒复制的反应。在各种类型的 RNA 核苷酸修饰中，m6A 是最常见的，涉及 RNA 细胞内定位、成熟、半衰期和翻译性等几个关键过程。这种表观转录组包含了混合的病毒 RNA 和细胞转录物。在之前的一项研究中，我们根据直接 RNA 测序分析了 SARS-CoV-2 RNA m6A 甲基化，并对不同病毒谱系中的 DRACH 基序突变进行了特征描述。在这里，我们利用直接 RNA 测序数据，展示了 SARS-CoV-2 感染 Vero 细胞（源自非洲绿猴）和 Calu-3 细胞（人）后的 m6A 转录物甲基化分析。通过非参数统计和两种计算方法（m6anet 和 EpiNano）对这些数据的分析表明，感染细胞的 RNA 中的 m6A 水平更高。功能富集分析显示，参与翻译、肽和胺代谢的转录物的 m6A 甲基化增加。这项分析确定了感染细胞转录物中差异甲基化的转录物和 m6A 独特位点。这里呈现的结果表明，细胞对病毒感染的反应不仅改变了 mRNAs 的水平，如前所述，还改变了其表观转录模式。此外，全转录组分析显示，在 Vero 和 Calu-3 细胞中，细胞转录物的 DRACH 基序存在强烈的核苷酸偏向性，这两个细胞使用的特征基序是 GGACU，而在病毒 RNA 中，特征基序是 GAACU。我们假设，DRACH 基序偏向性的差异可能迫使病毒基因组的趋同进化，从而更好地适应写入酶、读取酶和擦除酶的靶序列偏好。据我们所知，这是直接 RNA 测序首次报道 SARS-CoV-2 感染的 Vero 细胞的 m6A 表观转录组，即 RNA-seq。
