Figure 3: Zinc-responsive expression of the zitB gene in comparison with the zinc uptake (znuA) gene.

(a) S1 mapping analysis of znuA and zitB transcripts under TPEN or zinc-treated conditions. The wild-type cells grown in YEME medium to exponential growth (OD₆₀₀ of 0.4 to ∼0.5) were treated for 30 min with TPEN (5.0–6.5 μM as indicated), ZnSO₄ (25–150 μM as indicated) or none, before cell harvest. Quantifications of S1 mapping results were done from 11 independent experiments for znuA, and 3–6 experiments for zitB transcript analysis. The relative expression values with s.d.’s were presented, taking the untreated sample values as 1.0. The P values for all the measurements were <0.001 by Student’s t-test, except the zitB value for 5 μM TPEN treatment (P>0.8). (b) The relative expression levels of znuA and zitB mRNAs were plotted against the concentrations of zinc in the medium. The maximally expressed levels were drawn as equal heights in the y axis for both znuA (solid circle; left axis) and zitB (open circle; right axis). The concentration of zinc in the TPEN-treated samples was calculated from the concentrations of treated TEPN (μM; in blue number). The added amount of Zn (μM) was indicated in purple. The amount of zinc in non-treated YEME medium (−) estimated by ICP-MS was 1.33 μM. The biphasic induction of the zitB gene was labeled as phase I (at sub-femtomolar zinc) and phase II (at >micromolar zinc).