journal.ppat.1001056.pdf (5.4 MB)
Enterohemorrhagic E. coli Requires N-WASP for Efficient Type III Translocation but Not for EspFU-Mediated Actin Pedestal Formation
journal contributionposted on 2011-05-27, 00:00 authored by Didier Vingadassalom, Kenneth G. Campellone, Michael J. Brady, Brian Skehan, Scott E. Battle, Douglas Robbins, Archana Kapoor, Gail Hecht, Scott B. Snapper, John M. Leong
Upon infection of mammalian cells, enterohemorrhagic E. coli (EHEC) O157:H7 utilizes a type III secretion system to translocate the effectors Tir and EspFU (aka TccP) that trigger the formation of F-actin-rich ‘pedestals’ beneath bound bacteria. EspFU is localized to the plasma membrane by Tir and binds the nucleation-promoting factor N-WASP, which in turn activates the Arp2/3 actin assembly complex. Although N-WASP has been shown to be required for EHEC pedestal formation, the precise steps in the process that it influences have not been determined. We found that N-WASP and actin assembly promote EHEC-mediated translocation of Tir and EspFU into mammalian host cells. When we utilized the related pathogen enteropathogenic E. coli to enhance type III translocation of EHEC Tir and EspFU, we found surprisingly that actin pedestals were generated on N-WASP-deficient cells. Similar to pedestal formation on wild type cells, Tir and EspFU were the only bacterial effectors required for pedestal formation, and the EspFU sequences required to interact with N-WASP were found to also be essential to stimulate this alternate actin assembly pathway. In the absence of N-WASP, the Arp2/3 complex was both recruited to sites of bacterial attachment and required for actin assembly. Our results indicate that actin assembly facilitates type III translocation, and reveal that EspFU, presumably by recruiting an alternate host factor that can signal to the Arp2/3 complex, exhibits remarkable versatility in its strategies for stimulating actin polymerization.
This work was supported by National Institutes of Health (NIH) grants R01-AI46454 to J.M.L., R01-DK058964 and P01-DK050694 to G.H., and 5R01-AI052354 and 5P01-HL59561 to S.B.S.
Publisher Statement© 2010 Vingadassalom et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. The original publication is available through the Public Library of Science at DOI: 10.1371/journal.ppat.1001056.
PublisherPublic Library of Science