A bacterial regulatory uORF senses multiple classes of ribosome-targeting antibiotics

Expression of many bacterial genes is regulated by <i>cis</i>- and <i>trans</i>-acting elements in their 5' upstream regions (URs). <i>Cis</i>-acting regulatory elements in URs include upstream ORFs (uORFs), short ORFs that sense translation stress that manifests as ribosomes stalling at specific codons within the uORF. Here, we show that the transcript encoding the <i>Escherichia coli</i> TopAI-YjhQ toxin-antitoxin system is regulated by a uORF that we name '<i>toiL'</i>. We propose that in the absence of translation stress, a secondary structure in the UR represses translation of the <i>topAI</i> transcript by occluding the ribosome-binding site. Translation repression of <i>topAI</i> leads to premature Rho-dependent transcription termination within the <i>topAI</i> ORF. At least five different classes of ribosome-targeting antibiotics relieve repression of <i>topAI</i>. Our data suggest that these antibiotics function by stalling ribosomes at different positions within <i>toiL</i>, thereby altering the RNA secondary structure around the <i>topAI</i> ribosome-binding site. Thus, <i>toiL</i> is a multipurpose uORF that can respond to a wide variety of translation stresses.