Fig. 7. Model connecting metabolic homeostasis, cytoplasmic acidification and two paths for persister cell formation.

A strong perturbation of metabolic homeostasis by nutrient shifts or entry in stationary phase acidifies the cytoplasm. Such acidification could occur due to an imbalance between ATP consumption and regeneration. In these conditions, complex I acts as a regulator of cytoplasmic pH: wild-type proton translocation activity counter-balances acidification; impaired proton translocation in the nuo* mutants enhances cytoplasmic acidification after ±30 min of perturbation of metabolic homeostasis. Mild cytoplasmic acidification leads to increased persistence by an increased RpoS response, possibly mediated by inhibition of SpoT-mediated hydrolysis of (p)ppGpp (dominant path in black arrows in the wild type). Stronger cytoplasmic acidification halts protein synthesis and renders antibiotic targets inactive leading to a second persistence mechanism (dominant path in red arrows in nuo* mutants with impaired complex I function.) At various points in this model, feedback loops are in place that affect persister formation. For instance, RpoS inhibits adaptation to maintain metabolic homeostasis, recovering from the initial pH drop. In turn, strong intracellular acidification via halted protein synthesis inhibits full RpoS activation.