Depletion of the DarG antitoxin in Mycobacterium tuberculosis triggers the DNA‐damage response and leads to cell death

Of the ~80 putative toxin‐antitoxin (TA) modules encoded by the bacterial pathogen Mycobacterium tuberculosis (Mtb), three contain antitoxins essential for bacterial viability. One of these, Rv0060 (DNA ADP‐ribosyl glycohydrolase, DarG(Mtb)), functions along with its cognate toxin Rv0059 (DNA ADP‐ribosyl transferase, DarT(Mtb)), to mediate reversible DNA ADP‐ribosylation (Jankevicius et al., 2016). We demonstrate that DarT(Mtb)‐DarG(Mtb) form a functional TA pair and essentiality of darG(Mtb) is dependent on the presence of darT(Mtb), but simultaneous deletion of both darT(Mtb)‐darG(Mtb) does not alter viability of Mtb in vitro or in mice. The antitoxin, DarG(Mtb), forms a cytosolic complex with DNA‐repair proteins that assembles independently of either DarT(Mtb) or interaction with DNA. Depletion of DarG(Mtb) alone is bactericidal, a phenotype that is rescued by expression of an orthologous antitoxin, DarG(Taq), from Thermus aquaticus. Partial depletion of DarG(Mtb) triggers a DNA‐damage response and sensitizes Mtb to drugs targeting DNA metabolism and respiration. Induction of the DNA‐damage response is essential for Mtb to survive partial DarG(Mtb)‐depletion and leads to a hypermutable phenotype.