Figure 5.

A benzamide‐dependent strain of B. subtilis.

A. Strain DWA454 is benzamide dependent. Strains 168 (WT) and DWA454 (ftsZ sup9[FtsZ E251K]) were streaked on NA in the absence and presence of either 3‐MBA (10 mM) or 8J (2 μg ml⁻¹), as indicated.

B. The re‐engineered ftsZ sup9 mutation is sufficient for benzamide dependence. Strains DWA458 (ftsZ cat[FtsZ WT]) and DWA455 (ftsZ cat[FtsZ E251K]) (Site‐directed‐mutant; SDM) were streaked on NA + Cm in the absence and presence of either 3‐MBA (10 mM) or 8J (2 μg ml⁻¹), as indicated. Plates were photographed after incubation at 37°C for 18 h.

C and D. Cell division is benzamide dependent. Exponentially growing cells of strain DWA454 were examined following growth in the absence (C) and presence (D) of 8J (8 μg ml⁻¹). Arrowheads highlight unusual cell division events. Cell membranes were stained with FM5‐95.

E–H. Localisation of early‐assembling division proteins in the benzamide‐dependent strain background. Exponentially growing cells of strains DWA638 (FtsZ E251K, YFP‐FtsA) and DWA637 (FtsZ E251K, YFP‐ZapA) were observed following growth in the absence (E and G) and presence (F and H) of 8J (8 μg ml⁻¹).

I and J. Cell division defects associated with mutations at R168 and E251. Strains DWA410 (ftsZ cat[FtsZ R168A]) and DWA412 (ftsZ cat[FtsZ E251A]) were observed after growth overnight at 37°C on NA + Cm. Scale bars = 5 μm.

K. sep F is synthetic lethal with ftsZ sup9. Strains DWA460 (ΔsepF, P_xyl‐sep F, FtsZ G196S) and DWA461 (ΔsepF, P_xyl‐sepF, FtsZ E251K) were streaked on plates containing 0.5% w/v xylose, 8J (2 μg ml⁻¹) and both, as indicated. Plates were photographed after incubation at 37°C for 18 h.