P1: IwX
052182091Xc06.xml CB786/Lax 0 521 82091 X November 4, 2005 2:30
127
type iii–delivered toxins
on small GTP-binding proteins (Frank, 1997). In vivo,ExoS has been shown to
ADP-ribosylate Ras (H-Ras, Rap1B, and RalA) and Rab proteins (Figure 6.4)
(Barbieri et al., 2001; Fraylick et al., 2002; McGuffie et al., 1998; Riese et al.,
2001). ExoS ADP-ribosylation of H-Ras inhibits the interaction with its spe-
cific GEF, Cdc25, and consequently inhibits the nucleotide exchange reac-
tion catalysed by Cdc25 (Ganesan et al., 1999). Because only GTP-bound
Ras is capable of interacting with its downstream effectors, this disrupts
Ras-mediated signal transduction (Ganesan et al., 1998). Accordingly, in vivo
ExoS-mediated ADP-ribosylation of H-Ras inhibits the ability of the GTPase
to interact with its downstream effector Raf-1 (Vincent et al., 1999). The mod-
ification of H-Ras by ExoS correlates directly with its ability to inhibit DNA
synthesis (McGuffie et al., 1998).
ExoS ADP-ribosylation of Rap1B also inhibits the interaction between
Rap1B and its GEF, C3G, which could modulate the oxidative-burst in neu-
trophils or signal transduction through integrin-mediated pathways (Riese
et al., 2001). ExoS ADP-ribosylation of RalA alters its ability to bind its down-
stream effector RalBP1 (Fraylick et al., 2002). Similarly, the action of ExoS
on Rab5 inhibits the interaction of the GTPase with its effector EEA1 (early
endosome antigen 1) (Barbieri et al., 2001), which is predicted to affect phago-
some maturation. Therefore, ExoS may also inhibit phagocytosis through its
action on Rab.
ExoS displays GAP activity towards RhoA, Rac1, and Cdc42, both in vitro
and in vivo (Figure 6.4) (Goehring et al., 1999; Krall et al., 2002). Transfec-
tion of the amino terminal part of ExoS, or delivery of ExoS by the TTSS of
P. aeruginosa in eukaryotic cells, disrupts actin filaments (Krall et al., 2002;
Pederson et al., 1999). In addition, type III secretion delivered ExoS confers
phagocytosis resistance against macrophages (Frithz-Lindsten et al., 1997).
The primary structure of ExoT is 75% identical to that of ExoS. However,
the ADPRT activity of ExoT is only 0.2–1% of that of ExoS (Frank, 1997),
and ExoT does not interfere with cell viability or with Ras signalling (Sundin
et al., 2001). ExoT disrupts the cytoskeletal architecture of epithelial cells
(Vallis et al., 1999), and inhibits the uptake of P. aeruginosa by epithelial cells
and macrophages (Garrity-Ryan et al., 2000). These activities are partially
dependent on the GAP activity that ExoT exhibits towards RhoA, Rac1, and
Cdc42 both in vitro and in vivo (Figure 6.4) (Kazmierczak and Engel, 2002;
Krall et al., 2000). Furthermore, the GAP activity of ExoT is required for the
ability of the protein to inhibit lung epithelial wound repair in vitro (Geiser
et al., 2001). Since point mutations at the arginine finger of ExoT result
only in intermediate defects, it is predicted that its ADPRT activity may also
contribute to cell rounding and anti-internalisation activities (Sundin et al.,