FIG 6.

Nongrowing dormant intracellular S. Typhimurium is competent for translocating only defined SPI-2 effectors inside fibroblasts. (A) Expression of M45-tagged SPI-2 effectors in extracellular S. Typhimurium upon overnight growth at 37°C in LB medium at neutral pH under nonshaking conditions. Indicated are the respective SPI-2 effectors analyzed. (B) Selective translocation of tagged SseF-M45 and SseJ-M45 effectors occurs in nongrowing dormant intracellular fibroblasts. Note that there is no translocation of SifA-M45 or SifB-M45, even when there is prominent production of both effectors. (C) Control assays demonstrating that SseF-M45 and SseJ-M45 translocation depends on a functional SPI-2 TTSS. (D) Extraction of M45-tagged SPI-2 effectors in extracellular S. Typhimurium in buffer containing 1% (vol/vol) Triton X-100. Note the negligible amount of the bacterium-associated effector extracted by the detergent, which contrasts with the amount of effector detected outside the bacteria in infected fibroblasts (Fig. 4B and 5B). (E) Translocation rate of the effectors SifA, SseJ, and SifB in intracellular bacteria infecting NRK-49F fibroblasts pretreated for 30 h with a protease inhibitor cocktail or left untreated. (F) Efficient translocation of M45-tagged SseF, SseG, SifA, SifB, and SseJ by intracellular bacteria proliferating inside HeLa epithelial cells.