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

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 e xhibits 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 prote ins. Exchange of arginine 144 with alanine, located in this arginine finger motif, results in an inactive form of YopE that can no longer stimulate GT P 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 c erevisiae inhibits growth, while in contrast, expression of the inactive fo rm 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 Ste2 0, suppressed the growth phenotype of YopE in yeast cells. These results pr ovide 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.