Many bacterial genes are regulated by RNA elements in their 5´ untranslated regions (UTRs). However, the full complement of these elements is not known even in the model bacterium Escherichia coli. Using complementary RNA-sequencing approaches, we detected large numbers of 3´ ends in 5´ UTRs and open reading frames (ORFs), suggesting extensive regulation by premature transcription termination. We documented regulation for multiple transcripts, including spermidine induction involving Rho and translation of an upstream ORF for an mRNA encoding a spermidine efflux pump. In addition to discovering novel sites of regulation, we detected short, stable RNA fragments derived from 5´ UTRs and sequences internal to ORFs. Characterization of three of these transcripts, including an RNA internal to an essential cell division gene, revealed that they have independent functions as sRNA sponges. Thus, these data uncover an abundance of cis- and trans-acting RNA regulators in bacterial 5´ UTRs and internal to ORFs.
Keywords: E. coli; RNA sponges; RNA-mediated regulation; RNA-seq; Rho-dependent termination; base pairing sRNAs; infectious disease; microbiology; uORF.
In most organisms, specific segments of a cell’s genetic information are copied to form single-stranded molecules of various sizes and purposes. Each of these RNA molecules, as they are known, is constructed as a chain that starts at the 5´ end and terminates at the 3´ end. Certain RNAs carry the information present in a gene, which provides the instructions that a cell needs to build proteins. Some, however, are ‘non-coding’ and instead act to fine-tune the activity of other RNAs. These regulatory RNAs can be separate from the RNAs they control, or they can be embedded in the very sequences they regulate; new evidence also shows that certain regulatory RNAs can act in both ways. Many regulatory RNAs are yet to be catalogued, even in simple, well-studied species such as the bacterium Escherichia coli. Here, Adams et al. aimed to better characterize the regulatory RNAs present in E. coli by mapping out the 3´ ends of every RNA molecule in the bacterium. This revealed many new regulatory RNAs and offered insights into where these sequences are located. For instance, the results show that several of these RNAs were embedded within RNA produced from larger genes. Some were nested in coding RNAs, and were parts of a longer RNA sequence that is adjacent to the protein coding segment. Others, however, were present within the instructions that code for a protein. The work by Adams et al. reveals that regulatory RNAs can be located in unexpected places, and provides a method for identifying them. This can be applied to other types of bacteria, in particular in species with few known RNA regulators.