1. Graduate Program in Chemistry and Chemical Biology and,
2. Department of Cellular and Molecular Pharmacology and the Cell Propulsion Laboratory (an NIH Nanomedicine Development Center), University of California, San Francisco, 600 16th Street, San Francisco, California 94158, USA
3. Raymond and Beverly Sackler Laboratory of Genetics and Molecular Medicine, Department of Genetics and Developmental Biology and Center for Cell Analysis and Modeling (CCAM), University of Connecticut Health Center, Farmington, Connecticut 06030, USA
4. Present address: Department of Synthetic Biology, California Institute of Quantitative Biomedical Research (QB3), University of California, Berkeley, 327 Stanley Hall, Berkeley, California 94720, USA.

Correspondence to: Wendell A. Lim² Correspondence and requests for materials should be addressed to W.A.L. (Email: lim@cmp.ucsf.edu).

Abstract

Enterohaemorrhagic Escherichia coli attaches to the intestine through actin pedestals that are formed when the bacterium injects its protein EspF_U (also known as TccP) into host cells¹. EspF_U potently activates the host WASP (Wiskott–Aldrich syndrome protein) family of actin-nucleating factors, which are normally activated by the GTPase CDC42, among other signalling molecules. Apart from its amino-terminal type III secretion signal, EspF_U consists of five-and-a-half 47-amino-acid repeats. Here we show that a 17-residue motif within this EspF_U repeat is sufficient for interaction with N-WASP (also known as WASL). Unlike most pathogen proteins that interface with the cytoskeletal machinery, this motif does not mimic natural upstream activators: instead of mimicking an activated state of CDC42, EspF_U mimics an autoinhibitory element found within N-WASP. Thus, EspF_U activates N-WASP by competitively disrupting the autoinhibited state. By mimicking an internal regulatory element and not the natural activator, EspF_U selectively activates only a precise subset of CDC42-activated processes. Although one repeat is able to stimulate actin polymerization, we show that multiple-repeat fragments have notably increased potency. The activities of these EspF_U fragments correlate with their ability to coordinate activation of at least two N-WASP proteins. Thus, this pathogen has used a simple autoinhibitory fragment as a component to build a highly effective actin polymerization machine.

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