biostudies-literatureBarth VCHHS | National Institutes of HealthNIAID NIH HHSCoordenação de Aperfeiçoamento de Pessoal de Nível Superiornew jersey commission on cancer researchNIH HHSe2022136118https://www.ebi.ac.uk/biostudies/studies/S-EPMC8364209biostudies-literatureUnknown118(32)The <i>Mycobacterium tuberculosis</i> (Mtb) VapBC4 toxin-antitoxin system is essential for the establishment of Mtb infection. Using a multitier, systems-level approach, we uncovered the sequential molecular events triggered by the VapC4 toxin that activate a circumscribed set of critical stress survival pathways which undoubtedly underlie Mtb virulence. VapC4 exclusively inactivated the sole transfer RNA^Cys (tRNA^Cys) through cleavage at a single site within the anticodon sequence. Depletion of the pool of tRNA^Cys led to ribosome stalling at Cys codons within actively translating messenger RNAs. Genome mapping of these Cys-stalled ribosomes unexpectedly uncovered several unannotated Cys-containing open reading frames (ORFs). Four of these are small ORFs (sOProceedings of the National Academy of Sciences of the United States of America<i>Mycobacterium tuberculosis</i> VapC4 toxin engages small ORFs to initiate an integrated oxidative and copper stress response.PMC836420999999.013433/2013-00DFHS18PPC045R21 AI135461S10 OD025140AI135461R01 AI154464AI154464Husson RNBarth VCWoychik NAZheng HZeng JSu XChauhan Ufalse<i>Mycobacterium tuberculosis</i> VapC4 toxin engages small ORFs to initiate an integrated oxidative and copper stress response.The <i>Mycobacterium tuberculosis</i> (Mtb) VapBC4 toxin-antitoxin system is essential for the establishment of Mtb infection. Using a multitier, systems-level approach, we uncovered the sequential molecular events triggered by the VapC4 toxin that activate a circumscribed set of critical stress survival pathways which undoubtedly underlie Mtb virulence. VapC4 exclusively inactivated the sole transfer RNA^Cys (tRNA^Cys) through cleavage at a single site within the anticodon sequence. Depletion of the pool of tRNA^Cys led to ribosome stalling at Cys codons within actively translating messenger RNAs. Genome mapping of these Cys-stalled ribosomes unexpectedly uncovered several unannotated Cys-containing open reading frames (ORFs). Four of these are small ORFs (sO2021-01-01T00:00:00Z2021 Aug2026-05-08T10:35:22.292Z2025-04-07T00:26:05.46ZS-EPMC83642093436284110.1073/pnas.2022136118