Guidelines for Inferring and Characterizing a Family of Bacterial trans-Acting Small Noncoding RNAs

So far, every sequenced bacterial transcriptome encompasses hundreds of small regulatory noncoding RNAs (sRNAs). From those sRNAs that have been already characterized, we learned that their regulatory functions could span over almost every bacterial process, mostly acting at the posttranscriptional control of gene expression (Wagner and Romby, Adv Genet 90:133-208, 2015). Canonical molecular mechanisms of sRNA action have been described to rely on both sequence and/or structural traits of the RNA molecule. As for protein-coding genes, the conservation of sRNAs among species suggests conserved and adjusted functions across evolution. Knowing the phylogenetic distribution of an sRNA gene and how its functional traits have evolved may help to get a broad picture of its biological role in each single species. Here, we present a simple computational workflow to identify close and distant sRNA homologs present in sequenced bacterial genomes, which allows defining novel sRNA families. This strategy is based on the use of Covariance Models (CM) and assumes the conservation of sequence and structure of functional sRNA genes throughout evolution. Moreover, by carefully inspecting the conservation of the close genomic context of every member of the RNA family and how the patterns of microsynteny follow the path of species evolution, it is possible to define subgroups of sRNA orthologs, which in turn enables the definition of RNA subfamilies.