SweC and SweD are essential co-factors of the FtsEX-CwlO cell wall hydrolase complex in Bacillus subtilis
Fig 6
Point mutations in ftsE and ftsX suppress the ΔlytE ΔsweDC synthetic lethality.
(A) Spot dilutions of the indicated strains on permissive and restrictive media. All strains were grown in LB in the presence of IPTG (500 μM) to an optical density of ∼2.0. The cultures were washed twice without inducer, resuspended at an OD600 of 1.5, and 10-fold serially diluted. Five microliters of each dilution was spotted onto the indicated agar plates without inducer. The permissive condition contains CH medium supplemented with 0.25 M sucrose and 20 mM MgCl2. The ΔsweDC (ΔDC) ΔlytE double mutant, like the parental strain used to select for suppressors, can only grow under permissive conditions. A walH deletion or point mutations in ftsE or ftsX support growth on CH medium lacking IPTG but not LB. Combining ΔwalH with either point mutation supports growth on LB. Representative plates from one of three biological replicates are shown. (B) Cytological analysis of the ftsEX suppressors strains under permissive and restrictive conditions. The indicated strains were grown to exponential phase in CH medium in presence of IPTG (500 μM). The cultures were washed twice in CH medium lacking inducer and back-diluted to an OD of 0.01 in permissive and restrictive culture media lacking IPTG. Cells were analyzed using differential interference contrast (DIC) microscopy after 4 generations. Mutations in ftsE or ftsX restore rod-shape morphology to the ΔsweDC ΔlytE double mutant when grown under permissive conditions and restored viability and partial rod-shape morphology when grown under restrictive conditions. Scale bar indicates 2 μm. Representative images from one of three independent experiments are shown.