FIG. 10.

Production of type III secretion effector proteins in the absence of Hfq. Y. pseudotuberculosis wild-type, Δhfq, and pYV⁻ strains were cultured in BHI broth for 3 h under Yop secretion-inducing conditions (37°C, low Ca²⁺). Cells were lysed, and culture supernatants were harvested, filtered, and precipitated with trichloroacetic acid. (a) The overall protein profile of the supernatant was determined by Coomassie brilliant blue staining. Molecular masses in kDa are indicated to the left. (b) Levels of YopE, YopH, YopJ, and YopT were assessed by immunoblotting in cells and culture supernatants. Blots are representative of 3 independent experiments. The relative density of each band compared to wild type is indicated. RpoA in whole-cell lysates was used as a loading control. (c) qRT-PCR of yopE, yopH, yopJ, and yopT transcripts. Bars represent the relative average fold change of each transcript compared to wild type for 3 independent experiments. There were no significant differences between wild-type and mutant transcript levels for all yop genes tested.