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镰刀菌感染会改变角膜中 mA 修饰的转录本图谱。

Fusarium infection alters the mA-modified transcript landscape in the cornea.

作者信息

Hu Jianzhang, Lin Yi

机构信息

Department of Ophthalmology, Fujian Medical University Union Hospital, Fu Zhou, China.

Department of Ophthalmology, Fujian Medical University Union Hospital, Fu Zhou, China.

出版信息

Exp Eye Res. 2020 Nov;200:108216. doi: 10.1016/j.exer.2020.108216. Epub 2020 Sep 3.

Abstract

N6-methyladenosine (mA) is the most common post-transcriptional modification of RNA in eukaryotes that regulates the post-transcriptional expression level of genes without changing the base sequence. The role of mA in fungal keratitis has not yet been elucidated. Here, we aimed to identify mA modification changes and their potential roles in fungal keratitis. The murine model of fungal keratitis was established by inoculating mice with Fusarium solani (F. solani). The overall mA level was detected via an mA RNA methylation assay kit. The expression levels of key mA modification-related genes were estimated by quantitative real-time polymerase chain reaction (PCR). The expression and localization of METTL (methyltransferase like)3, the key component of the mA methyltransferase complex, was determined by immunostaining and Western blotting (WB). Immunoprecipitation methylation microarray was used to describe the changes in mA modification in F. solani-infected corneal tissue. The overall mA level in corneal tissue on the 5th day in the F. solani-treated group was upregulated compared with that in the control group. The demethylase levels were unaltered, but the level of the methylase METTL3 was increased significantly after fungal infection. Additionally, differences were found in mA modifications in 1137 mRNAs, of which 780 were hypermethylated and 357 were hypomethylated. To the best of our knowledge, the present work is the first investigation on the mA modification profiles in experimental fungal keratitis, and it may provide a potential therapeutic target.

摘要

N6-甲基腺苷(mA)是真核生物中最常见的RNA转录后修饰,它在不改变碱基序列的情况下调节基因的转录后表达水平。mA在真菌性角膜炎中的作用尚未阐明。在此,我们旨在确定mA修饰变化及其在真菌性角膜炎中的潜在作用。通过用茄病镰刀菌(F. solani)接种小鼠建立真菌性角膜炎的小鼠模型。通过mA RNA甲基化检测试剂盒检测总体mA水平。通过定量实时聚合酶链反应(PCR)评估关键的mA修饰相关基因的表达水平。通过免疫染色和蛋白质印迹法(WB)确定mA甲基转移酶复合物的关键成分METTL(甲基转移酶样)3的表达和定位。免疫沉淀甲基化微阵列用于描述茄病镰刀菌感染的角膜组织中mA修饰的变化。与对照组相比,茄病镰刀菌处理组第5天角膜组织中的总体mA水平上调。去甲基化酶水平未改变,但真菌感染后甲基化酶METTL3的水平显著增加。此外,在1137个mRNA的mA修饰中发现了差异,其中780个发生了高甲基化,357个发生了低甲基化。据我们所知,目前的工作是对实验性真菌性角膜炎中mA修饰谱的首次研究,它可能提供一个潜在的治疗靶点。

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