(p)ppGpp and malonyl-CoA set the pace for Staphylococcus aureus adaptation to FASII antibiotics and provide a basis for bi-therapy inhibition

Fatty acid biosynthesis (FASII) enzymes have been considered as valid targets for antimicrobial drug development against the human pathogen . However, our findings showed that incorporation of compensatory host fatty acids confers FASII antibiotic adaptation that compromises prospective treatments. Adaptation to anti-FASII drugs is preceded by a non-replicative latency period. Here we investigated the factors that dictate the duration of latency prior to adaptation outgrowth, and identify stringent response as a regulator of both FASII and adaptation to FASII inhibition. We first show that (p)ppGpp induction inhibits malonyl-CoA synthesis, and constitutes a second regulator of FASII and phospholipid synthesis in , in addition to the FapR repressor. We then show that anti-FASII treatment triggers transient (p)ppGpp induction during the anti-FASII latency phase, with a concomitant reduction of FapR regulon expression. This effect is reversed upon adaptive outgrowth. Using three different sensors, we give evidence that anti-FASII treatment shifts the distribution of malonyl-CoA between its interactants, FapR and FabD, increasing expression of phospholipid synthesis genes and during outgrowth. We conclude that (p)ppGpp pools dictate malonyl-CoA availability in FASII regulation, and contribute to latency prior to anti-FASII adaptation. A combinatory approach, coupling a (p)ppGpp inducer and an anti-FASII, blocks growth, opening perspectives for bi-therapy treatment.