Aeromonas salmonicida subsp. salmonicida contains a functional type III secretion system that is responsible for the secretion of the ADP-ribosylating toxin AexT. In this study, the authors identified AopP as a second effector protein secreted by this system. The aopP gene was detected in both typical and atypical A. salmonicida isolates and was found to be encoded on a small plasmid of approximately 6?4 kb. Sequence analysis indicates that AopP is a member of the YopJ family of effector proteins, a group of proteins that interfere with mitogen-activated protein kinase (MAPK) and/or nuclear factor kappa B (NF-kB) signalling pathways. AopP inhibits the NF-kB pathway downstream of IkB kinase (IKK) activation, while a catalytically inactivated mutant, AopPC177A, does not possess this inhibitory effect. Unlike other effectors of the YopJ family, such as YopJ and VopA, AopP does not inhibit the MAPK signalling pathway.

Type III protein secretion has been shown recently to be important in the virulence of the fish pathogen Aeromonas salmonicida. The ADP-ribosylating toxin Aeromonas exoenzyme T (AexT) is one effector protein targeted for secretion via this system. In this study, we identified muscular and nonmuscular actin as substrates of the ADP-ribosylating activity of AexT. Furthermore, we show that AexT also functions as a GTPaseactivating protein (GAP), displaying GAP activity against monomeric GTPases of the Rho family, specifically Rho, Rac, and Cdc42. Transfection of fish cells with wild type AexT resulted in depolymerization of the actin cytoskeleton and cell rounding. Point mutations within either the GAP or the ADP-ribosylating active sites of AexT (Arg-143 as well as Glu-398 and Glu-401, respectively) abolished enzymatic activity, yet did not prevent actin filament depolymerization. However, inactivation of the two catalytic sites simultaneously did. These results suggest that both the GAP and ADP-ribosylating domains of AexT contribute to its biological activity. This is the first bacterial virulence factor to be described that has a specific actin ADP-ribosylation activity and GAP activity toward Rho, Rac, and Cdc42, both enzymatic activities contributing to actin filament depolymerization.Aeromonas salmonicida subsp. salmonicida (A. salmonicida) is the causative agent of furunculosis, a systemic disease that affects salmonid fish (salmon, trout, and char). A. salmonicida expresses a variety of extracellular toxins, many of which have been implicated in virulence. Several of these factors, including the pore-forming toxin aerolysin, serine protease, and the phospholipase GCAT, are secreted into the environment via the well characterized type II or general secretory pathway (1, 2). However, like many Gram-negative pathogens, A. salmonicida also possesses a type III protein secretion system (3-5). Such secretion systems translocate toxins, or type III effector proteins, directly into the cytosol of eukaryotic cells. Here the effector proteins are able to modulate cell signaling pathways or, alternatively, disrupt the dynamics of the cytoskeleton (6, 7).To date, the following genes encoding four type III effector proteins have been identified in A. salmonicida: aexT, aopP, aopO, and aopH (8 -10). Although functional studies of AopO and AopH have not yet been carried out, sequence homology with type III effector proteins YopO and YopH of Yersinia sp. suggests that AopO and AopH modify the actin cytoskeleton of target eukaryotic cells. In contrast, AopP has been shown to interfere with the NF-B signaling pathway by inhibiting translocation of NF-B into the nucleus of target cells (10).AexT, the first type III effector of A. salmonicida to be characterized, has been shown to function as an ADP-ribosylating toxin (8). However, its substrate has never before been identified. AexT also displays significant sequence homology to the bifunctional type III effector proteins ExoS and ExoT, expressed by Pseudomonas aeruginosa. ...