Nucleotidyltransferase toxin MenT targets and extends the aminoacyl acceptor ends of serine tRNAs in vivo to control Mycobacterium tuberculosis growth

Toxins of toxin-antitoxin systems use diverse mechanisms to control bacterial growth and represent attractive therapeutic targets to fight pathogens. In this study, we characterized the translation inhibitor toxin MenT3 of Mycobacterium tuberculosis , the bacterium responsible for human tuberculosis in humans. We show that MenT3 is a robust cytidine specific tRNA nucleotidyltransferase in vitro , capable of modifying the aminoacyl acceptor ends of most tRNA but with a marked preference for tRNA ^Ser , to which long stretches of cytidines were added. Furthermore, transcriptomic-wide analysis of MenT3 targets in M. tuberculosis identified tRNA ^Ser as the sole target of MenT3 in vivo and revealed significant detoxification attempts by ribonuclease PH in response to MenT3 overexpression. Finally, under physiological conditions, only in the presence the native menAT3 operon, we found the unexpected presence of an active pool of endogenous MenT3 targeting tRNA ^Ser in M. tuberculosis , likely reflecting the importance of MenT3 during infection.