Smekalova Elena M, Gerashchenko Maxim V, O'Connor Patrick B F, Whittaker Charles A, Kauffman Kevin J, Fefilova Anna S, Zatsepin Timofei S, Bogorad Roman L, Baranov Pavel V, Langer Robert, Gladyshev Vadim N, Anderson Daniel G, Koteliansky Victor
David H. Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, MA 02142, USA.
Division of Genetics, Department of Medicine, Brigham and Women's Hospital and Harvard Medical School, Boston, MA 02115, USA.
Mol Ther Nucleic Acids. 2020 Mar 6;19:252-266. doi: 10.1016/j.omtn.2019.11.009. Epub 2019 Nov 18.
Translation is an essential biological process, and dysregulation is associated with a range of diseases including ribosomopathies, diabetes, and cancer. Here, we examine translation dysregulation in vivo using RNAi to knock down the m-subunit of the translation initiation factor eIF3 in the mouse liver. Transcriptome sequencing, ribosome profiling, whole proteome, and phosphoproteome analyses show that eIF3m deficiency leads to the transcriptional response and changes in cellular translation that yield few detectable differences in the translation of particular mRNAs. The transcriptional response fell into two main categories: ribosome biogenesis (increased transcription of ribosomal proteins) and cell metabolism (alterations in lipid, amino acid, nucleic acid, and drug metabolism). Analysis of ribosome biogenesis reveals inhibition of rRNA processing, highlighting decoupling of rRNA synthesis and ribosomal protein gene transcription in response to eIF3m knockdown. Interestingly, a similar reduction in eIF3m protein levels is associated with induction of the mTOR pathway in vitro but not in vivo. Overall, this work highlights the utility of a RNAi-based in vivo approach for studying the regulation of mammalian translation in vivo.
翻译是一个重要的生物学过程，其失调与一系列疾病相关，包括核糖体病、糖尿病和癌症。在此，我们利用RNA干扰在小鼠肝脏中敲低翻译起始因子eIF3的m亚基，以研究体内的翻译失调。转录组测序、核糖体谱分析、全蛋白质组和磷酸蛋白质组分析表明，eIF3m缺陷导致转录反应以及细胞翻译的变化，而这些变化在特定mRNA的翻译中几乎检测不到差异。转录反应主要分为两类：核糖体生物合成（核糖体蛋白转录增加）和细胞代谢（脂质、氨基酸、核酸和药物代谢改变）。对核糖体生物合成的分析揭示了rRNA加工受到抑制，这突出了rRNA合成与核糖体蛋白基因转录在响应eIF3m敲低时的解偶联。有趣的是，eIF3m蛋白水平的类似降低在体外与mTOR途径的诱导相关，但在体内并非如此。总体而言，这项工作突出了基于RNA干扰的体内方法在研究哺乳动物体内翻译调控方面的实用性。
