Abstract
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 tRNAPro 5ʹ half, a 35 residue long ncRNA associated with ribosomes and polysomes in several mammalian cell lines. Addition of tRNAPro 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 tRNAPro 5ʹ half-dependent translation product consists of both RNA and amino acids. Transfection of the tRNAPro 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 tRNAPro 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.
Highlights
In early high-throughput studies, tRNA-derived fragments were identified in a wide variety of model systems yet they were considered as degradation products devoid of physiological relevance [1]
It could be demonstrated that many tRNA-derived fragments (tdRs), which include tRNA half molecules as well as even shorter tRNA fragments, are processed by endonucleases to participate in and regulate diverse biological process [2]. tRNA halves and tRNA fragments are involved in controlling cell proliferation, cellular homoeostasis, priming of viral reverse transcriptases, regulating gene expression, trans-generational epigenetic inheritance, RNA processing, modulation of the DNA damage response, tumour suppression, regulation of transposition, neurodegeneration, ribosome biogenesis, and translation regulation [2,3,4,5]
Since there are characterized examples of tRNA fragments (tRFs) and tRNA halves directly binding to ribosomes and in this way regulating translation [13,15], northern blot screening was performed on total RNA and on RNA isolated from the crude ribosome pellets in parallel
Summary
In early high-throughput studies, tRNA-derived fragments (tdRs) were identified in a wide variety of model systems yet they were considered as degradation products devoid of physiological relevance [1]. With the improved deepsequencing protocols, tailor-made bioinformatics tools, and accumulating functional studies it became clear that tdRs represent a so far unknown class of regulatory ncRNAs. It could be demonstrated that many tdRs, which include tRNA half molecules as well as even shorter tRNA fragments (tRFs), are processed by endonucleases to participate in and regulate diverse biological process [2]. Ribosome-targeted tdRs have recently been linked to cell proliferation in mammalian cells and might promote cancer progression in a mouse model [16]. These findings reveal ribosome-bound tdRs as functionally versatile regulators capable of fine-tuning protein biosynthesis in a stress-dependent manner
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