Li Zhu, Liu Yi, Zhang Ling, Tian Jiahua, Wang Hongping, Ding Hongwei, Nie Jin, Pi Hang, Wang Bingyao, Liu Daishun
Medical College of Soochow University, 215000, Suzhou, China.
Department of Respiratory Medicine, The First People's Hospital of Zunyi, (The Third Affiliated Hospital of Zunyi Medical University), 563000, Guizhou, China.
Heliyon. 2023 Apr 6;9(4):e15307. doi: 10.1016/j.heliyon.2023.e15307. eCollection 2023 Apr.
Respiratory syncytial virus (RSV) is the second leading cause of death due to lower respiratory tract infections. Effective prevention and treatment measures are lacking, posing a huge socioeconomic burden to the world. -methyladenosine (mA) is the most common internal modification in messenger RNA and noncoding RNA. Numerous recent studies have shown that the dysregulation of mA modification is associated with diseases caused by pathogenic viruses.
The changes in mA modification were evaluated using mA RNA methylation assay. The differences in gene expression levels of various mA-modifying enzymes were observed using Quantitative Real-time PCR (qRT-PCR) during RSV infection. The autophagosomes were observed using transmission electron microscopy, and the expression of autophagy-associated protein Microtubule Associated Protein 1 Light Chain 3 Beta Ⅱ/Ⅰ (LC3B Ⅱ/Ⅰ) and Beclin1 in Human Normal Lung Epithelial Cells (BEAS-2B) cells using Western blot during RSV infection. The significantly differentially expressed genes were screened guided by bioinformatics. Their relationship with mA-modifying enzymes was analyzed through protein-protein interaction network and expression correlation analysis.
The mA abundance decreased and demethylase Fat Mass and Obesity- Associated Protein (FTO) significantly increased during RSV infection after 24 h. We also found that the DNA Damage-Inducible Transcript 3 Protein (DDIT3) level significantly increased during RSV infection after 24 h and observed autophagosomes in BEAS-2B cells. In addition, RSV infection could cause the upregulation of LC3B Ⅱ/Ⅰ and Beclin1. The expression correlation analysis showed that DDIT3 levels were positively correlated with the FTO level, and Methyltransferase Like 3 (METTL3), RNA Binding Motif Protein 15B (RBM15B), YTH Domain-Containing Family Protein 1 (YTHDF1), and levels were negatively correlated with the DDIT3 level.
We uncovered a significant role for mA modification during RSV infection. Also, a correlation was found between mA and autophagy, providing new ideas for therapeutic advancements in RSV treatment.
呼吸道合胞病毒（RSV）是下呼吸道感染致死的第二大病因。目前缺乏有效的预防和治疗措施，给全球带来了巨大的社会经济负担。N6-甲基腺苷（m⁶A）是信使核糖核酸和非编码核糖核酸中最常见的内部修饰。最近的大量研究表明，m⁶A修饰失调与致病性病毒引起的疾病有关。
采用m⁶A RNA甲基化检测法评估m⁶A修饰的变化。在RSV感染期间，使用定量实时聚合酶链反应（qRT-PCR）观察各种m⁶A修饰酶基因表达水平的差异。使用透射电子显微镜观察自噬体，并在RSV感染期间，通过蛋白质免疫印迹法观察人正常肺上皮细胞（BEAS-2B）中自噬相关蛋白微管相关蛋白1轻链3βⅡ/Ⅰ（LC3BⅡ/Ⅰ）和Beclin1的表达。在生物信息学的指导下筛选出显著差异表达的基因。通过蛋白质-蛋白质相互作用网络和表达相关性分析，分析它们与m⁶A修饰酶的关系。
RSV感染24小时后，m⁶A丰度降低，去甲基化酶脂肪量和肥胖相关蛋白（FTO）显著增加。我们还发现，RSV感染24小时后，DNA损伤诱导转录3蛋白（DDIT3）水平显著增加，并在BEAS-2B细胞中观察到自噬体。此外，RSV感染可导致LC3BⅡ/Ⅰ和Beclin1上调。表达相关性分析表明，DDIT3水平与FTO水平呈正相关，而甲基转移酶样3（METTL3）、RNA结合基序蛋白15B（RBM15B）、含YTH结构域家族蛋白1（YTHDF1）水平与DDIT3水平呈负相关。
我们揭示了m⁶A修饰在RSV感染过程中的重要作用。此外，还发现了m⁶A与自噬之间的相关性，为RSV治疗的治疗进展提供了新的思路。
