One cannot rule them all: Are bacterial toxins-antitoxins druggable?

Chan, Wai Ting; Balsa, Dolors; Espinosa, Manuel

The first gene that was discovered to be involved in persistence was hipA from E. coli K-12, which is also the toxin gene of hipBA TA pair (Moyed and Bertrand 1983) . Toxin HipA inhibits aminoacylation and it is counteracted by its cognate antitoxin HipB which, in turn, is degraded by Lon protease (Gerdes, Christensen and Lobner-Olensen 2005; Schumacher et al., 2009; Hansen et al., 2012; Germain et al., 2013) . Upon exposure to ampicillin, the wild-type strain showed a persister cell ratio of 10 −5 to 10 −6 ; however, a double mutant of hipA, the hipA7 allele that abolished the toxicity of HipA, showed an increased frequency of persisters ratio up to 10 −2 under the same experimental conditions (Moyed and Bertrand 1983) . Overexpression of wild-type HipA also increased the frequency of persisters to the levels conferred by the HipA7 allele (Korch and Hill 2006) . HipA toxin was shown to phophorylate Ser 239 , which is near the active centre of tRNA Glu -bound glutamyl-tRNA synthetase, thus accumulating uncharged tRNA Glu (Germain et al., 2013) . This finding was taken as an indication that the entering of uncharged tRNA Glu into the A site of the ribosome would trigger activation and release of RelA, which is an alarmone pentaphosphate (pppGpp) synthetase. This in turn would lead to dramatic increases of pppGpp, whose levels are positively correlated to the persistence level (Germain et al., 2013; Maisonneuve, Castro-Camargo and Gerdes 2013) . In another study by the same group, formation of persisters was shown to be modulated by the signalling nucleotide pppGpp through a cascade involving inorganic polyphosphate (PolyP; a long linear polymer of orthophosphate residues), Lon protease and TAs. The concentration of PolyP is controlled by pppGpp, which competitively inhibits exopolyphosphatase from degrading PolyP. PolyP was demonstrated to stimulate Lon-mediated degradation of the antitoxins of TA pairs, stochastically, in small subpopulation of cells. This would lead to the release of toxins that eventually would promote inhibition of cellular biosynthesis, cessation of cell growth and persistence (Maisonneuve, Castro-Camargo and Gerdes 2013) . Previous studies showed that prolonged exposure to antibiotics activated transcription of TAs encoding mRNases in E. coli K-12; this activation led to inhibition of global cellular translation, rendering the cells to a state of dormancy and persistence. Single deletion of a toxin mRNase did not affect persister formation, and it required deletion of 10 TAs encoding mRNAses to cumulatively reduce the levels of persisters (Maisonneuve et al., 2011) . Redundancy of TAs can increase the frequency of persister cells and stochastic fluctuations can spontaneously switch on the TAs giving rise to the bistable state of normal cells and persisters (Fasani and Savageau 2013) .