Institute of Biochemistry, Department of Biology, ETH Zurich, 8093 Zurich, Switzerland.
Molecular Life Sciences Ph.D. Program, 8057 Zurich, Switzerland.
Nucleic Acids Res. 2022 Mar 21;50(5):2872-2888. doi: 10.1093/nar/gkac072.
Ribosome assembly is an essential process that is linked to human congenital diseases and tumorigenesis. While great progress has been made in deciphering mechanisms governing ribosome biogenesis in eukaryotes, an inventory of factors that support ribosome synthesis in human cells is still missing, in particular regarding the maturation of the large 60S subunit. Here, we performed a genome-wide RNAi screen using an imaging-based, single cell assay to unravel the cellular machinery promoting 60S subunit assembly in human cells. Our screen identified a group of 310 high confidence factors. These highlight the conservation of the process across eukaryotes and reveal the intricate connectivity of 60S subunit maturation with other key cellular processes, including splicing, translation, protein degradation, chromatin organization and transcription. Intriguingly, we also identified a cluster of hits comprising metabolic enzymes of the polyamine synthesis pathway. We demonstrate that polyamines, which have long been used as buffer additives to support ribosome assembly in vitro, are required for 60S maturation in living cells. Perturbation of polyamine metabolism results in early defects in 60S but not 40S subunit maturation. Collectively, our data reveal a novel function for polyamines in living cells and provide a rich source for future studies on ribosome synthesis.
核糖体组装是一个与人类先天性疾病和肿瘤发生有关的重要过程。尽管在破译真核生物核糖体生物发生的机制方面已经取得了很大的进展，但仍缺乏支持人类细胞核糖体合成的因素清单，特别是关于大 60S 亚基的成熟。在这里，我们使用基于成像的单细胞测定法进行了全基因组 RNAi 筛选，以揭示促进人类细胞 60S 亚基组装的细胞机制。我们的筛选鉴定出了一组 310 个高可信度的因素。这些因素突出了该过程在真核生物中的保守性，并揭示了 60S 亚基成熟与其他关键细胞过程（包括剪接、翻译、蛋白质降解、染色质组织和转录）之间的复杂连接。有趣的是，我们还鉴定出了一组包含多胺合成途径的代谢酶的命中。我们证明，多胺长期以来一直被用作缓冲添加剂以支持体外核糖体组装，在活细胞中 60S 的成熟是必需的。多胺代谢的扰乱会导致 60S 但不是 40S 亚基成熟的早期缺陷。总的来说，我们的数据揭示了多胺在活细胞中的新功能，并为核糖体合成的未来研究提供了丰富的资源。
