A 3' external transcribed spacer in a tRNA transcript acts as a sponge for small RNAs to prevent transcriptional noise.

During ribosomal and transfer RNA maturation, external transcribed spacer (ETS) and internal transcribed spacer (ITS) sequences are excised and, as non-functional by-products, are rapidly degraded. However, we report that the 3'ETS of the glyW-cysT-leuZ polycistronic tRNA precursor is highly and specifically enriched by co-purification with at least two different small regulatory RNAs (sRNAs), RyhB and RybB. Both sRNAs are shown to base pair with the same region in the 3'ETS of leuZ (3'ETS(leuZ)). Disrupting the pairing by mutating 3'ETS(leuZ) strongly increased the activity of sRNAs, even under non-inducing conditions. Our results indicate that 3'ETS(leuZ) prevents sRNA-dependent remodeling of tricarboxylic acid (TCA) cycle fluxes and decreases antibiotic sensitivity when sRNAs are transcriptionally repressed. This suggests that 3'ETS(leuZ) functions as a sponge to absorb transcriptional noise from repressed sRNAs. Additional data showing RybB and MicF sRNAs are co-purified with ITS(metZ-metW) and ITS(metW-metV) strongly suggest a wide distribution of this phenomenon.