FIG 3 .
(A) The NvarAL mutant did not increase OmpU or ToxR levels in response to NRES. This phenotype could not be complemented with pVarA, suggesting a suppressor mutation. Strains N16961 (WT), NvarAL, NvarAL/pWKS30 (vector control), and NvarAL/pVarA (complemented strain) were grown overnight in LB broth and then subcultured 1:100 into T medium with or without 12.5 mM NRES mix. Cells were harvested in the mid-log phase, and whole-cell preparations were resolved by SDS-PAGE (10%) and stained by Coomassie blue (top panel) or immunoblotted using polyclonal anti-ToxR antisera (bottom panel). (B) The NvarAL strain did not have an inherent defect in OmpU production, since OmpU was made exclusively in response to the bile acid cholate. Strains N16961 (WT) and NvarAL were grown overnight in LB broth and then subcultured 1:100 into T medium with or without 0.1% cholate. Cells were harvested in the mid-log phase, and whole-cell preparations were resolved by SDS-PAGE (10%) and stained with Coomassie blue to visualize the OMPs.