FIG. 7.

YadA-mediated invasion requires tyrosine kinases, Ser/Thr kinases, and the PI3K of the host cells. About 5 × 10⁴ HEp-2 cells were preincubated for 30 min at 37°C in RPMI 1640 medium supplemented with 0.4% bovine serum albumin and 20 mM HEPES (pH 7.0) with increasing concentrations of the tyrosine kinase inhibitors genistein (A) and herbimycin (B), the Ser/Thr kinase inhibitor staurosporine (C), and the PI3K inhibitor wortmannin (D). Subsequently, about 5 × 10⁶ bacteria of strains CC118/pAM226 (inv⁺), grown in LB medium, and CC118/pPD284 (yadA⁺), grown in LB medium supplemented with 0.2% arabinose, were added to the cell monolayer, which was incubated for 1 h at 37°C to monitor invasion efficiency. The number of intracellular bacteria was determined by the gentamicin protection assay as described in reference 17. Each value represents the mean of three independent assays done in triplicate. E.c., E. coli.