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Infection and Immunity, September 2008, p. 3854-3868, Vol. 76, No. 9
0019-9567/08/$08.00+0     doi:10.1128/IAI.00072-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

EspF Interacts with Nucleation-Promoting Factors To Recruit Junctional Proteins into Pedestals for Pedestal Maturation and Disruption of Paracellular Permeability

Departments of Cell Biology,¹ Molecular Biomedicine,² Physiology, Biophysics and Neurosciences, Centro de Investigación y de Estudios Avanzados (CINVESTAV-IPN), Ap. Postal 14-740, 07000 Mexico DF, Mexico,³ Morphology Unit, Universidad Nacional Autónoma de México (UNAM-FES Cuautitlan), Ap. Postal 54714, Cuautitlan, Mexico,⁴ Unité des Microbiologie Moléculaire (INRA-ENVT), Ecole Nationale Vétérinaire, 31000 Toulouse Cedex 3, France⁵

Received 18 January 2008/ Returned for modification 17 March 2008/ Accepted 9 June 2008

Many pathogenic bacteria subvert normal host cell processes by delivering effector proteins which mimic eukaryotic functions directly into target cells. EspF is a multifunctional protein injected into host cells by attaching and effacing pathogens, but its mechanism of action is not understood completely. In silico analyses of EspF revealed two key motifs: proline-rich domains and PDZ domain binding motifs. Such functional domains may allow EspF to act as an actin nucleation-promoting factor by mimicking host proteins. In agreement with these predictions, we found that EspF from rabbit enteropathogenic Escherichia coli (E22) participates in the regulation of actin polymerization by binding to a complex of proteins at the tight junctions (TJ). EspF bound to actin and profilin throughout the course of infection. However, after 2 h of infection, EspF also bound to the neural Wiskott-Aldrich syndrome protein and to the Arp2/3, zonula occludens-1 (ZO-1), and ZO-2 proteins. Moreover, EspF caused occludin, claudin, ZO-1, and ZO-2 redistribution and loss of transepithelial electrical resistance, suggesting that actin sequestration by EspF may cause local actin depolymerization leading to EspF-induced TJ disruption. Furthermore, EspF caused recruitment of these TJ proteins into the pedestals. An E22 strain lacking EspF did not cause TJ disruption and pedestals were smaller than those induced by the wild-type strain. Additionally, the pedestals were located mainly in the TJ. The overexpression of EspF caused bigger pedestals located along the length of the cells. Thus, actin sequestration by EspF allows the recruitment of junctional proteins into the pedestals, leading to the maturation of actin pedestals and the disruption of paracellular permeability.

Published ahead of print on 16 June 2008.

Editor: V. J. DiRita