Equivalent results were found following Everolimus in vivo exposure of Mycobacterium bovis BCG to streptomycin. Exposure to antimicrobial agents with nonribosome targets did not affect tmRNA levels. The increased tmRNA levels were associated with increased output from the ssrA promoter, which controls tmRNA transcription, without evidence of a change in tmRNA

degradation. These results suggest that the upregulation of tmRNA expression was an important response of bacteria to exposure to ribosome-inhibiting antimicrobial agents. Prokaryotes and some eukaryotic mitochondria possess a specialized process, trans-translation, which rescues ribosomes that have stalled during translation of a transcript. This process is the subject of several recent reviews (Moore & Sauer, 2007; Keiler, 2008). The ribosome states that can lead to the triggering of trans-translation include encountering a rare codon when the ribosome has to wait for a low abundance tRNA (Roche & Sauer, 1999) and when the end

of a transcript is reached without an in-frame stop codon (Keiler et al., 1996). Rescue by trans-translation provides a stalled ribosome with an alternate coding region permitting normal termination of translation and dissociation of the translation complex. Central to trans-translation is Galunisertib solubility dmso a specialized RNA species, tmRNA, which has properties comparable to both tRNA and mRNA (Komine et al., 1994; Ushida et al., 1994; Tu et al., 1995). The tRNA-like domain is aminoacylated by alanyl-tRNA synthetase (Komine et al., 1994; Barends et al., 2000) and the mRNA-like domain provides a short coding region with a stop codon; the amino acid sequence of this coding region tags polypeptides for rapid degradation by the ClpXP and ClpAP proteases (Sauer et al., 2004). The tmRNA molecule is transcribed from the ssrA gene as a precursor tmRNA (pre-tmRNA), which becomes processed at the 5′ and 3′ ends by RNases including RNase P and possibly RNase E (Lin-Chao et al., 1999; Withey & Friedman, 2003). The tmRNA binds to the protein SmpB (Karzai et

al., 1999) and this complex is believed to be the unique functional unit of trans-translation. Previous studies demonstrated that disrupting trans-translation increased susceptibility to protein synthesis inhibitors in Escherichia coli, Salmonella typhimurium, and Synechocystis sp. (de la Cruz & out Vioque, 2001; Abo et al., 2002; Vioque & de la Cruz, 2003; Luidalepp et al., 2005). This suggested that trans-translation is important to bacteria for overcoming the effects of ribosome-targeting antimicrobial agents, although it was not clear whether ribosome inhibition by antimicrobial agents altered the rate of trans-translation. Evidence that such agents may affect trans-translation came from previous studies reporting that ribosome inhibitors caused an increase in the levels of tmRNA in Thermotoga maritima (Montero et al., 2006) and Streptomyces aureofaciens (Paleckova et al., 2006).