ABSTRACT

Originally considered futile degradation products, tRNA-derived RNA fragments (tdRs) have been shown over the recent past to be crucial players in orchestrating various cellular functions. Unlike other small non-coding RNA (ncRNA) classes, tdRs possess a multifaceted functional repertoire ranging from regulating transcription, apoptosis, RNA interference, ribosome biogenesis to controlling translation efficiency. A subset of the latter tdRs has been shown to directly target the ribosome, the central molecular machine of protein biosynthesis. Here we describe the function of the mammalian tRNA^Pro 5ʹ half, a 35 residue long ncRNA associated with ribosomes and polysomes in several mammalian cell lines. Addition of tRNA^Pro halves to mammalian in vitro translation systems results in global translation inhibition and concomitantly causes the upregulation of a specific low molecular weight translational product. This tRNA^Pro 5ʹ half-dependent translation product consists of both RNA and amino acids. Transfection of the tRNA^Pro half into HeLa cells leads to the formation of the same product in vivo. The migration of this product in acidic gels, the insensitivity to copper sulphate treatment, the resistance to 3ʹ polyadenylation, and the association with 80S monosomes indicate that the accumulated product is peptidyl-tRNA. Our data thus suggest that binding of the tRNA^Pro 5ʹ half to the ribosome leads to ribosome stalling and to the formation of peptidyl-tRNA. Our findings revealed a so far unknown functional role of a tdR thus further enlarging the functional heterogeneity of this emerging class of ribo-regulators.

摘要

tRNA衍生的RNA片段（tdRs）原本被认为是徒劳的降解产物，但最近证明在协调各种细胞功能方面起着至关重要的作用。与其他小型非编码RNA（ncRNA）类不同，tdR具有多方面的功能库，范围从调节转录，凋亡，RNA干扰，核糖体生物发生到控制翻译效率。后者的tdRs的一个子集已显示直接靶向核糖体，这是蛋白质生物合成的中心分子机器。这里，我们介绍了哺乳动物的tRNA的功能^Pro的5'半，一个35残长非编码RNA与核糖体和多核糖体在几种哺乳动物细胞系相关。将tRNA ^Pro一半加入体外哺乳动物翻译系统导致整体翻译抑制，并随之引起特定的低分子量翻译产物的上调。这tRNA的^临5'半依赖性翻译产品由RNA和氨基酸。将tRNA ^Pro的一半转染到HeLa细胞中会导致在体内形成相同的产物。该产物在酸性凝胶中的迁移，对硫酸铜处理的不敏感性，对3′聚腺苷酸的抗性以及与80S单核糖体的结合，表明所积累的产物是肽基-tRNA。因此，我们的数据表明，tRNA ^Pro的结合核糖体的5/5一半导致核糖体停滞并形成肽基-tRNA。我们的发现揭示了迄今未知的tdR的功能作用，从而进一步扩大了这种新兴的核糖调节剂类别的功能异质性。