Figure 1

Response of the σ^E-dependent and the CpxA/R-regulated envelope stress pathways to inactivation and overexpression of ppiD. Eσ^E (A) and Cpx (B) activities in the indicated strains carrying SurA (light gray bars), PpiD (dark gray bars), and Skp (black bars) encoding plasmids or an empty vector (pASK75; white bars) were assayed by monitoring the accumulation of β-galactosidase resulting from σ^E-dependent rpoH P3::lacZ and from Cpx-meditated cpxP-lacZ reporter expression, respectively. Cells were grown in LB (σ^E) or in LB buffered at pH 7.0 (Cpx) at 37°C and β-galactosidase activities were determined as described in Methods and compared to that of wild-type cells. Results represent the average of at least two independent experiments (*P ≤ 0.05; **P ≤ 0.01 Student's t-test). Qualitatively similar results were obtained from cells grown at 30°C (data not shown). (C) Western blot analysis of crude extracts derived from cells with (+) and without (-) pPpiD. A volume of sample equivalent to 4 × 10⁷ cells was loaded onto each lane. The anti-PpiD antiserum showed a weak unspecific cross-reaction with a similar sized unknown protein. The intensity of the PpiD signal relative to that in the wild-type strain (rel. Int.) was calculated using MalE as the internal standard for each lane.