Department of Developmental Biology, Sloan-Kettering Institute, 1275 York Ave, Box 252, New York City, New York 10065, USA.
Department of Pharmacology, Weill Medical College, Cornell University, New York City, New York 10065, USA.
Nat Commun. 2017 Jul 4;8:15737. doi: 10.1038/ncomms15737.
The conserved modification N-methyladenosine (mA) modulates mRNA processing and activity. Here, we establish the Drosophila system to study the mA pathway. We first apply miCLIP to map mA across embryogenesis, characterize its mA 'writer' complex, validate its YTH 'readers' CG6422 and YT521-B, and generate mutants in five mA factors. While mA factors with additional roles in splicing are lethal, mA-specific mutants are viable but present certain developmental and behavioural defects. Notably, mA facilitates the master female determinant Sxl, since multiple mA components enhance female lethality in Sxl sensitized backgrounds. The mA pathway regulates Sxl processing directly, since miCLIP data reveal Sxl as a major intronic mA target, and female-specific Sxl splicing is compromised in multiple mA pathway mutants. YT521-B is a dominant mA effector for Sxl regulation, and YT521-B overexpression can induce female-specific Sxl splicing. Overall, our transcriptomic and genetic toolkit reveals in vivo biologic function for the Drosophila mA pathway.
保守修饰 N6-甲基腺苷（m6A）调节 mRNA 加工和活性。在这里，我们建立了果蝇系统来研究 m6A 途径。我们首先应用 miCLIP 来绘制胚胎发生过程中的 m6A 图谱，表征其 m6A“写入器”复合物，验证其 YTH“读取器”CG6422 和 YT521-B，并生成五个 m6A 因子的突变体。虽然具有额外剪接作用的 m6A 因子是致死的，但 m6A 特异性突变体是存活的，但存在某些发育和行为缺陷。值得注意的是，m6A 促进了主要的雌性决定因子 Sxl，因为多个 m6A 成分增强了 Sxl 敏感背景下的雌性致死率。m6A 途径直接调节 Sxl 的加工，因为 miCLIP 数据显示 Sxl 是主要的内含子 m6A 靶标，并且多个 m6A 途径突变体中的雌性特异性 Sxl 剪接受损。YT521-B 是 Sxl 调控的显性 m6A 效应因子，YT521-B 过表达可以诱导雌性特异性 Sxl 剪接。总体而言，我们的转录组学和遗传工具包揭示了果蝇 m6A 途径的体内生物学功能。
