Posttranscriptional 3'-Terminal Modifications of Escherichia coli RNA Fragments Evolved for Diversity Boosting.

An emerging area of microbial biology focuses on oligonucleotides excised from functional RNAs and subsequently fulfilling an independent cellular role. Some of these products are subjected to modifications that may expand their functional inventory. Here, we applied a differential analysis of intra- and extracellular RNA fragments produced by wild-type Escherichia coli and its dps-null mutant and discovered leucine tRNA fragments with random 3'-terminal extensions among oligonucleotides with Dps-dependent secretion. We observed an exclusive intracellular enrichment of modified LeuT(VPQ) tRNA fragments compared to secretomes, with abundance level dependent on growth medium and the presence of competing bacteria. To assess the pervasiveness of this phenomenon, we developed a custom computational pipeline for detecting variable RNA termini in RNA-seq data. Beyond LeuT(VPQ) tRNA fragments, several other genomic loci yielded oligos with highly heterogeneous ends, indicating that terminal elongation, most prevalent in LeuT(VPQ), is not exclusive to these fragments. Ex vivo testing using synthetic LeuT(VPQ) analogs revealed their stimulatory effect on the persistence of multiple taxa in an artificial microbiome, which was attenuated by 3'-end elongation. We propose that non-template extensions may serve to broaden the spectrum of target molecules for elimination of unused mRNAs by an interference-like mechanism or to generate sequences absent from the E. coli genome as part of a primitive defense system.