Transfer RNAs (tRNA) are best known for their role as adaptors during translation of the genetic code. Beyond their canonical role during protein biosynthesis, tRNAs also perform additional functions in both prokaryotes and eukaryotes for example in regulating gene expression. Aminoacylated tRNAs have also been implicated as substrates for non-ribosomal peptide bond formation, post-translational protein labeling, modification of phospholipids in the cell membrane, and antibiotic biosyntheses. Most recently tRNA fragments, or tRFs, have also been recognized to play regulatory roles. Here, we examine in more detail some of the new functions emerging for tRNA in a variety of cellular processes outside of protein synthesis.

Aminoacyl-tRNA synthetases use a variety of mechanisms to ensure fidelity of the genetic code and ultimately select the correct amino acids to be used in protein synthesis. The physiological necessity of these quality control mechanisms in different environments remains unclear, as the cost vs benefit of accurate protein synthesis is difficult to predict. We show that in Escherichia coli, a non-coded amino acid produced through oxidative damage is a significant threat to the accuracy of protein synthesis and must be cleared by phenylalanine-tRNA synthetase in order to prevent cellular toxicity caused by mis-synthesized proteins. These findings demonstrate how stress can lead to the accumulation of non-canonical amino acids that must be excluded from the proteome in order to maintain cellular viability.DOI: http://dx.doi.org/10.7554/eLife.02501.001

a b s t r a c tThe role of tRNA in translating the genetic code has received considerable attention over the last 50 years, and we now know in great detail how particular amino acids are specifically selected and brought to the ribosome in response to the corresponding mRNA codon. Over the same period, it has also become increasingly clear that the ribosome is not the only destination to which tRNAs deliver amino acids, with processes ranging from lipid modification to antibiotic biosynthesis all using aminoacyl-tRNAs as substrates. Here we review examples of alternative functions for tRNA beyond translation, which together suggest that the role of tRNA is to deliver amino acids for a variety of processes that includes, but is not limited to, protein synthesis. Ó 2009 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved. tRNA-dependent amino acid biosynthesisThe attachment of amino acids to the 3 0 -end of tRNAs is catalyzed by the aminoacyl-tRNA synthetase (aaRS) family of proteins [1]. aaRSs are ubiquitous and essential but only eukaryotes and a handful of bacteria have the full set of 20 enzymes, one for each canonical amino acid in the genetic code. Most bacteria and archaea lack asparaginyl-tRNA synthetase (AsnRS) and/or glutaminyl-tRNA synthetase (GlnRS) and some methanogenic archaea lack cysteinyl-tRNA synthetase (CysRS) [2]. Also, no aaRS for the rare amino acid selenocysteine has been found in any domain of life [3]. These organisms instead use indirect pathways to synthesize a number of amino acids (Asn, Cys, Gln and Sec) directly on their cognate tRNA: non-discriminating aaRSs first form misacylated aminoacyl-tRNA (aa-tRNA), which is not used by the ribosome but instead converted to cognate aa-tRNA by various RNA-dependent modifying enzymes [4].In organisms lacking GlnRS and AsnRS, Glu-tRNA Gln and AsptRNA Asn are synthesized by non-discriminating aaRSs and converted to cognate Gln-tRNA Gln and Asn-tRNA Asn by tRNA-dependent amidotransferases (AdT). Two types of AdT exist, the heterotrimeric GatCAB present in both bacteria and archaea and the homodimeric GatDE present in archaea [5,6]. The tRNA moiety is recognized by the B and E kinase subunits of GatCAB and GatDE, respectively, which phosphorylate the mischarged tRNAs to form activated intermediates [7][8][9]. The glutaminase subunit (GatA/D) liberates ammonia from an amide donor and amidates Glu or Asp on the tRNA to form Gln or Asn, respectively. In both types of AdT, a 40 Å-long hydrophilic channel connects the glutaminase and kinase subunits [9,10]. It has been proposed, but remains to be proven, that ammonia liberated in the glutaminase active site is transported through the channel via a series of protonations and deprotonations to the kinase active site, and that binding of mischarged tRNA is required for opening the channel. Another open question concerns the precise in vivo mechanism by which misacylated aa-tRNA species are stabilized and escape detection, and subsequent delivery to the ribosome, by e...

Significance Dual-function RNAs base pair with target messenger RNAs as small regulatory RNAs and encode small protein regulators. However, only a limited number of these dual-function regulators have been identified. In this study, we show that a well-characterized base-pairing small RNA surprisingly also encodes a 15–amino acid protein. The very small protein binds the cyclic adenosine monophosphate receptor protein transcription factor to block activation of some promoters, raising the question of how many other transcription factors are modulated by unidentified small proteins.

Bacteria are known to use RNA, either as mRNAs encoding proteins or as non-coding small RNAs (sRNAs), to regulate numerous biological processes. However, a few sRNAs have two functions: they act as base pairing RNAs and encode a small protein with additional regulatory functions. Thus, these so called “dual-function” sRNAs can serve as both a riboregulator and an mRNA. In some cases, these two functions can act independently within the same pathway while in other cases, the base-pairing function and protein function act in different pathways. Here, we discuss the five known dual-function sRNAs: SgrS from enteric species, RNAIII and Psm-mec from Staphylococcus aureus, Pel RNA from Streptococcus pyogenes, and SR1 from Bacillus subtilis, and review their mechanisms of action and roles in regulating diverse biological processes. We also discuss the prospect of finding additional dual-function sRNAs and future challenges in studying the overlap and competition between the functions.