In this attempt, we run into a previously described phenomenon that may become a source of erroneous results. If toxins are expressed from the arabinose-inducible P BAD promoter and antitoxins from an IPTG-inducible promoter, MDV3100 it is important to consider that

IPTG inhibits P BAD directly [71]. When we used an expression vector that encoded for the IPTG-insensitive C280* version of AraC transcriptional activator, we could not see any cross-inhibition. Based on that, a recent report on functional non-cognate TA interactions in Mycobacterium tuberculosis[67] may require retesting. Selective targeting of mRNA by toxins as a mechanism of gene regulation In the current study, we found that the cleavage products produced by TA toxins differ in stability. Selective targeting of mRNAs by endoribonucleolytic toxins and different ZD1839 stabilities of the resulting cleavage products may constitute another layer of gene regulation in the bacterial stress response. Differences in half-life and translational efficiency of mRNA cleavage products, along with generation of a pool of ribosomes

lacking the anti-Shine-Dalgarno sequence (as shown for MazF [22]), could profoundly affect the proteome composition. An example of such an effect is the occurrence of a MazF-resistant protein pool in E. coli[72]. The accumulation of toxin-encoding mRNA fragments may have potential use as a marker of toxin activation in studies of stressed and non-growing bacteria. Increase Cell press of the JNK assay T/A ratio may possibly trigger a positive feedback loop consisting of transcriptional activation of the TA operon, successive cleavage of the TA transcript, buildup of the toxin-encoding mRNA fragments, and translation of them, shifting the T/A balance (Figure 7). Thus, it can be related to TA-linked growth heterogeneity in bacterial populations (Additional file 1: Figure S6) [38, 39,

54]. Conclusions The main finding of this study is that bacterial toxin-antitoxin systems affect mutually each others’ expression and activity (Figure 7). We show that overexpression of one toxin can activate transcription of the other TA operons. Toxins with endoribonuclease activity add another layer of complexity to these interactions. They cleave TA mRNA, which is followed by degradation of the antitoxin-encoding RNA fragments and accumulation of the toxin-encoding fragments. We show that these accumulating mRNA fragments can be translated to produce more toxin. Most of bacteria have many different TA systems. Although their function is debatable, many TA toxins have similar activity and the inhibitory effect on bacterial cells is common to all of them. Therefore, an important question is whether TA systems are redundant or not. Another intriguing issue is whether different TA systems are functionally connected and do cross-talk [44, 70]. Here we over-expressed toxins to show that TA systems have a potential to form a network of cross-reacting regulators in E. coli.