FIG 3.

Anti-FASII treatment leads to transient acc repression and shifts in malonyl-CoA distribution. (A) Schematic diagram illustrating two known malonyl-CoA interactants, FapR and FabD. FapR is a repressor of FASII and phospholipid synthesis. Malonyl-CoA binds FapR (left), which reverses FapR repressor activity. FabD uses malonyl-CoA to produce malonyl-ACP, the FASII precursor (right). (B) Assessment of malonyl-CoA production by ELISA and P_accBC-lacZ. Sandwich ELISA was used to measure total malonyl-CoA (left) (see the Materials and Methods for detail) and P_accBC-lacZ reporter activity (right). (C) Assessment of FapR-bound malonyl-CoA. FASII-antibiotic-adapted S. aureus display altered malonyl-CoA distribution (see A). The Newman strain and derivatives were grown in nonselective (SerFA) and anti-FASII (SerFA-Tric) media. β-Gal activities were measured in SerFA at A₆₀₀ = ∼2.0 (exponential growth [Exp]) and in SerFA-Tric during latency after 3 h growth (A₆₀₀ = ∼0.3 [Latency]) and upon adaptive exponential outgrowth (A₆₀₀ = ∼2 [Outgrowth]). (D) Assessment of FapR-bound malonyl-CoA in a fabD mutant using FapR-Trap. β-Gal assays of wild-type Newman strain and fabD mutant (CondT^R-17, a point mutant [4]) carrying FapR-Trap were performed in SerFA after 3 h growth (A₆₀₀ = ∼1.0 [Exp]). Measurements in B to D represent means ± standard deviations from triplicate independent experiments. *, P ≤ 0.05, ns, not significant, using Mann-Whitney tests.