Enteric virulence associated protein VapC inhibits translation by cleavage of initiator tRNA

Abstract

Eukaryotic PIN (PilT N-terminal) domain proteins are ribonucleases involved in quality control, metabolism and maturation of mRNA and rRNA. The majority of prokaryotic PIN-domain proteins are encoded by the abundant vapBC toxin—antitoxin loci and inhibit translation by an unknown mechanism. Here we show that enteric VapCs are site-specific endonucleases that cleave tRNA^fMet in the anticodon stem-loop between nucleotides +38 and +39 in vivo and in vitro. Consistently, VapC inhibited translation in vivo and in vitro. Translation-reactions could be reactivated by the addition of VapB and extra charged tRNA^fMet. Similarly, ectopic production of tRNA^fMet counteracted VapC in vivo. Thus, tRNA^fMet is the only cellular target of VapC. Depletion of tRNA^fMet by vapC induction was bacteriostatic and stimulated ectopic translation initiation at elongator codons. Moreover, addition of chloramphenicol to cells carrying vapBC induced VapC activity. Thus, by cleavage of tRNA^fMet, VapC simultaneously may regulate global cellular translation and reprogram translation initiation.