GAP activity of the Yersinia YopE cytotoxin specifically targets the Rho pathway: a mechanism for disruption of actin microfilament structure

Abstract

The YopE cytotoxin of Yersinia pseudotuberculosis is an essential virulence determinant that is injected into the eukaryotic target cell via a plasmid‐encoded type III secretion system. Injection of YopE into eukaryotic cells induces depolymerization of actin stress fibres. Here, we show that YopE exhibits a GTPase‐activating protein (GAP) activity and that the presence of YopE stimulates downregulation of Rho, Rac and Cdc42 activity. YopE has an arginine finger motif showing homology with those found in other GAP proteins. Exchange of arginine 144 with alanine, located in this arginine finger motif, results in an inactive form of YopE that can no longer stimulate GTP hydrolysis by the GTPase. Furthermore, a yopE(R144A) mutant is unable to induce cytotoxicity on cultured HeLa cells in contrast to the corresponding wild‐type strain. Expression of wild‐type YopE in cells of Saccharomyces cerevisiae inhibits growth, while in contrast, expression of the inactive form of YopE, YopE(R144A), does not affect the yeast cells. Co‐expression of proteins belonging to the Rho1 pathway of yeast, Rho1, Rom2p, Bck1 and Ste20, suppressed the growth phenotype of YopE in yeast cells. These results provide evidence that YopE exhibits a GAP activity to inactivate RhoGTPases, leading to depolymerization of the actin stress fibres in eukaryotic cells and growth inhibition in yeast.

Proteasomal Degradation of Cytotoxic Necrotizing Factor 1-Activated Rac

“…In addition, it was shown recently that various bacterial effectors which are delivered into mammalian cells by type III secretion machinery affect Rho GTPases not by modifying but by modulating their functions. For example, several pathogens, including Pseudomonas aeruginosa and Salmonella and Yersinia species, produce GAP-like proteins, which terminate the active state of Rho GTPases (8,9,28). Moreover, Salmonella produces a noncovalently acting activator of Rho GTPases (SopE/ SopE2) (12), as well as an inactivator (SptP) (8), probably to switch on and off cytoskeletal changes in the host target cell in a space-and time-dependent manner according to the requirements of the pathogen.…”

Yersinia effectors target mammalian signalling pathways

“…Recently, YopE and ExoS were also demonstrated to possess GAP activity towards Rho family GTPases (RhoA, Rac and Cdc42) ( Fig. 1) (Goehring et al, 1999;Black and Bliska, 2000;Von Pawel-Rammingen et al, 2000). Activation of Rho GTP-binding proteins is a pivotal signalling event connecting receptor activation to cytoskeletal reorganization during phagocytosis in professional and non-professional phagocytic cells (Chimini and Chavrier, 2000;May and Machesky, 2001).…”

The Proteasome Pathway Destabilizes Yersinia Outer Protein E and Represses Its Antihost Cell Activities

“…This results from the blockade of Rho-GTPase activities (5,6). This effect of YopE, together with the action of YopH, YopT, and YopO/ YpkA, provides Yersinia with the ability to resist phagocytosis.…”

“…Interestingly, three of these Yops act on members of the Rho-GTPase family, which critically control rearrangements of the actin cytoskeleton (3,4). YopE is a GTPase-activating protein (GAP), which inactivates Rho-GTPases by increasing their intrinsic GTPase activity (5,6). This switches the GTPases from an active, GTP-loaded into an inactive, GDP-bound state.…”

“…This switches the GTPases from an active, GTP-loaded into an inactive, GDP-bound state. The preferred target of the YopE GAP activity in cells appears to be the Rac GTPase, although in vitro YopE can operate on Rho and Cdc42 as well (5)(6)(7). YopT acts as cysteine protease that inactivates Rho-GTPase members by cleaving their C-terminal geranylgeranyl isoprenoid moieties (8,9).…”