Efficient uptake of Yersinia pseudotuberculosis via integrin receptors involves a Rac1-Arp 2/3 pathway that bypasses N-WASP function

Efficient uptake of Yersinia pseudotuberculosis into cultured mammalian cel ls is the result of high-affinity binding of invasin to beta (1) chain inte grins. We demonstrate here that uptake requires Rac1 and Arp 2/3 function. Bacterial uptake was stimulated by GTP gammaS, but was inhibited in mammali an cells transfected with the interfering Rac1-N17 derivative. Rac1 was fou nd to be activated in response to integrin engagement by invasin, whereas R ac1 and Arp 2/3 were found to be intensely localized around phagosomes bear ing bacteria, indicating a specific role for Rac1 signalling from the nasce nt phagosome to downstream effectors. To determine whether the Arp 2/3 comp lex was a component of this proposed pathway, cells overproducing various d erivatives of Scar1/WAVE1, an Arp 2/3-binding protein, were analysed. Seque stration of Arp 2/3 away from the phagocytic cup as a result of Scar1/WAVE1 overproduction dramatically inhibited uptake. To determine whether signall ing from Rac1 to Arp 2/3 occurred via N-WASP, uptake was analysed in a cell line lacking expression of WASP and N-WASP. Uptake was unaffected by the a bsence of these proteins, indicating that beta (1) integrin signalling from Rac1 to Arp 2/3 can occur in the absence of N-WASP function.