Wei Lin Lee1,2, Jonathan M Grimes2,3,* & Robert C Robinson1,4
1 Institute of Molecular and Cell Biology, Agency for Science, Technology and Research (A*STAR), Singapore.
2 Division of Structural Biology, Wellcome Trust Centre for Human Genetics, University of Oxford, Oxford, UK.
3 Diamond Light Source, Oxfordshire, UK.
4 Department of Biochemistry, National University of Singapore, Singapore.
Published in Nature Structural & Molecular Biology 9 February 2015
Pathogenic Yersinia species evade host immune systems through the injection of Yersinia outer proteins (Yops) into phagocytic cells. One Yop, YopO, also known as YpkA, induces actin-filament disruption, impairing phagocytosis. Here we describe the X-ray structure of Yersinia enterocolitica YopO in complex with actin, which reveals that YopO binds to an actin monomer in a manner that blocks polymerization yet allows the bound actin to interact with host actin-regulating proteins. SILAC-MS and biochemical analyses confirm that actin-polymerization regulators such as VASP, EVL, WASP, gelsolin and the formin diaphanous 1 are directly sequestered and phosphorylated by YopO through formation of ternary complexes with actin. This leads to a model in which YopO at the membrane sequesters actin from polymerization while using the bound actin as bait to recruit, phosphorylate and misregulate host actin-regulating proteins to disrupt phagocytosis.
Structure of the YopO-actin complex. Two views of the structure (front and top view). Actin is shown as a cyan schematic within a gray semitransparent surface with the subdomains labeled. The YopO surface on actin is colored cyan. YopO GDI domain is in red, and the kinase domain is in blue. The catalytic residue Asp267 is shown as magenta spheres. Ordered N- and C-terminal residues are shown as green spheres.
Mechanism of YopO-inhibition of phagocytosis. YopO is produced in the bacteria as an inactive agent. On injection into the mammalian cell through the T3SS, YopO senses the environment of host cytoplasm through binding to actin, leading to kinase activation, and localizes via the membrane-association domain to the inner surface of the plasma membrane proximal to the bacterium site of attachment. YopO is further restricted in its localization to areas of actin polymerization through binding to Rac and RhoA, while inhibiting Rac/Rho-mediated actin assembly in the phagocytic cup. YopO’s sequestration of monomeric actin prevents its integration into a filament, yet the YopO–actin complex binds freely to VASP, EVL, formins, WASP and WIP. Thus, YopO counteracts host defenses by launching a three-pronged attack on disrupting actin dynamics within the macrophage: inhibiting signaling, blocking polymerization at the site of host pathogen contact and the misregulation of the actin remodeling machineries via phosphorylation.
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