FIGURE 4.

Induction of actin cytoskeletal rearrangements by OMVs harboring T3SS1 effectors in host cell. (A) Comparison of F-actin polymerization elicited by treatment with OMVs isolated from wild-type and ΔSPI-1, sopB, sopE2 strains. Aliquots of cell lysates from equivalent HeLa cells were ultracentrifuged to separate F-actin as described in the Section “Materials and Methods.” F-actin fractions and remnants of total cell lysates were analyzed by immunoblot assay using anti-actin antibody. Cytochalasin D (CytoD) was used to inhibit host cell actin polymerization. The levels of F-actin formation were normalized using the levels of total actin and the ratio of F-actin/total actin in the mock treatment was set to 1.0. Asterisks indicate significant differences with a P-value < 0.05. (B) Microscopic analysis for cytoskeletal reorganization of HeLa cells induced by OMVs treatments. HeLa cells were treated with OMVs isolated from wild-type and ΔSPI-1, sopB, sopE2 strains as described above. OMVs, actin, and nucleus are located using anti-Salmonella antibody (red), anti-actin antibody (green), and DAPI (blue), respectively, in confocal immunofluorescence microscopy.