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

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