Microbial pathogens use a variety of mechanisms to disrupt the actin cytoskeleton during infection. Vibrio parahaemolyticus (V. para) is a Gram-negative bacterium that causes gastroenteritis, and new pandemic strains are emerging throughout the world. Analysis of the V. para genome revealed a type III secretion system effector, VopL, encoding three Wiskott-Aldrich homology 2 domains that are interspersed with three proline-rich motifs. Infection of HeLa cells with V. para induces the formation of long actin fibers in a VopL-dependent manner. Transfection of VopL promotes the assembly of actin stress fibers. In vitro, recombinant VopL potently induces assembly of actin filaments that grow at their barbed ends, independent of eukaryotic factors. Vibrio VopL is predicted to be a bacterial virulence factor that disrupts actin homeostasis during an enteric infection of the host.actin assembly ͉ microbial pathogenesis ͉ virulence ͉ stress fibers ͉ WH2 domains

During infection, bacterial pathogens utilize a type III secretion system to inject effectors into the cytoplasm of a target cell where they disrupt the defense system of the host cell. Vibrio parahaemolyticus, a causative agent of gastroenteritis endemic in Southeast Asia, has a type III secretion system that encodes a novel member of the YopJ-like protein effector family, VopA (Vibrio outer protein A). Our studies revealed that Vibrio VopA encodes an evolutionarily conserved activity that is extremely potent and requires an intact catalytic site to abrogate signaling pathways in a manner distinct from that of other YopJ-like effectors. We observed that VopA efficiently inhibits the MAPK signaling pathways but not the NFB pathway in mammalian cells. When expressed in yeast, VopA induces a growth arrest phenotype and also blocks yeast MAPK signaling pathways. Our observations provide insight into the immense diversity of targets utilized by YopJ-like effectors to manipulate eukaryotic signaling machineries that are important for the response and survival of the host cell during infection and/or symbiosis.

The bacterial pathogen Vibrio parahemeolyticus manipulates host signaling pathways during infections by injecting type III effectors. One of these effectors, Vibrio outer protein A (VopA), inhibits MAPK signaling via a novel mechanism, distinct from those described for other bacterial toxins, that disrupts this signaling pathway. VopA is an acetyltransferase that potently inhibits MAPK signaling pathways not only by preventing the activation of MAPK kinases (MKKs) but also by inhibiting the activity of activated MKKs. VopA acetylates a conserved lysine found in the catalytic loop of all kinases and blocks the binding of ATP, but not ADP, on the MKKs, resulting in an inactive phosphorylated kinase. Acetylation of this conserved lysine inhibits kinase activity by a new mechanism of regulation that has not been observed previously. Identifying the target of VopA reveals a way that the reversible post-translational modification of lysine acetylation can be used to regulate the activity of an enzyme.Vibrio parahemeolyticus is a marine bacterium and causative agent of gastroenteritis associated with the consumption of contaminated seafood. It is endemic to Southeast Asia and is the leading cause of gastroenteritis in Japan (1). Sequencing of the V. parahemeolyticus genome revealed the presence of two type III secretion systems each encoded in separate pathogenicity islands (2). Type III secretion systems are used by bacterial pathogens to inject effector proteins into the cytoplasm of the host cells (3). Upon delivery into the host cytosol, the effectors manipulate host signaling to gain an advantage for the infecting pathogen. The bacterial effectors are proposed to usurp or mimic eukaryotic activity that is then used to target and deregulate normal functions in the host cell by either up-or downregulating signaling pathways (4). However, the bacterial effector proteins must initially be kept in a quiescent state within the bacterium. This is accomplished by a variety of methods, including their association with a chaperone in the bacterial cytosol, their specificity for a substrate, and/or their requirement for an activator or modification that is only provided by a eukaryotic source (4, 5).VopA (Vibrio outer protein A) is an effector found within one of the V. parahemeolyticus pathogenicity islands, and is a member of a family of type III effectors called YopJ-like proteins that are found in a wide variety of pathogens including other animal pathogens, plant pathogens, and the plant symbiont Rhizobium (5-7). The founding member of the family, YopJ from Yersinia spp., inhibits the mitogen-activated protein kinase (MAPK) 3 and NFB signaling cascades within the host cell, thereby inhibiting the host's innate immune response (7). All YopJ-like proteins contain a catalytic triad, and it was recently shown that YopJ functions as an acetyltransferase (8 -10). It has been speculated that other members of the family of YopJ-like proteins might also possess this activity (8). YopJ was demonstrated to inhibit the MAPK and NFB...

SummaryThe pathogenic bacteria Yersinia spp. contain a virulence plasmid that encodes a type III secretion system and effectors. During infection, four of the effectors target the actin cytoskeleton, crippling the phagocytic machinery in the infected cell. The remaining two effectors dampen the innate immune response by targeting important signalling pathways. Although the biochemical activity for each of these effectors is known, the mechanisms involved in their ordered secretion and delivery remain elusive.