Pseudomonas aeruginosa ExoS ADP-ribosyltransferase inhibits ERM phosphorylation.
Pseudomonas aeruginosa causes life-threatening infections in compromised and cystic fibrosis patients. Pathogenesis stems from a number of virulence factors, including four type III translocated cytotoxins: ExoS, ExoT, ExoY and ExoU. ExoS is a bifunctional toxin: the N terminus (amino acids 96-219) encodes a Rho GTPase Activating Protein (GAP) domain. The C terminus (amino acids 234-453) encodes a 14-3-3-dependent ADP-ribosyltransferase domain which transfers ADP-ribose from NAD onto substrates such as the Ras GTPases and vimentin. Ezrin/radixin/moesin (ERM) proteins have recently been identified as high-affinity substrates for ADP-ribosylation by ExoS. Expression of ExoS in HeLa cells led to a loss of phosphorylation of ERM proteins that was dependent upon the expression of ADP-ribosyltransferase activity. MALDI-MS and site-directed mutagenesis studies determined that ExoS ADP-ribosylated moesin at three C-terminal arginines (Arg553, Arg560 and Arg563), which cluster Thr558, the site of phosphorylation by protein kinase C and Rho kinase. ADP-ribosylated-moesin was a poor target for phosphorylation by protein kinase C and Rho kinase, which showed that ADP-ribosylation directly inhibited ERM phosphorylation. Expression of dominant active-moesin inhibited cell rounding elicited by ExoS, indicating that moesin is a physiological target in cultured cells. This is the first demonstration that a bacterial toxin inhibits the phosphorylation of a mammalian protein through ADP-ribosylation. These data explain how the expression of the ADP-ribosylation of ExoS modifies the actin cytoskeleton and indicate that ExoS possesses redundant enzymatic activities to depolymerize the actin cytoskeleton.
Maresso
Anthony W
AW
Department of Microbiology and Molecular Genetics, Medical College of Wisconsin, Milwaukee, WI 53226, USA.
Deng
Qing
Q
Pereckas
Michael S
MS
Wakim
Bassam T
BT
Barbieri
Joseph T
JT
D036002:ADP Ribose Transferases / Q000378:metabolism*; D000199:Actins / Q000378:metabolism; D001427:Bacterial Toxins / Q000378:metabolism*; D003598:Cytoskeletal Proteins / Q000378:metabolism*; D003599:Cytoskeleton / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008565:Membrane Proteins / Q000378:metabolism*; D008840:Microfilament Proteins / Q000737:chemistry / Q000378:metabolism*; D010766:Phosphorylation; D011493:Protein Kinase C; D011550:Pseudomonas aeruginosa / Q000201:enzymology* / Q000472:pathogenicity; D011994:Recombinant Proteins / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D037521:Virulence Factors / Q000378:metabolism; D000099149:Ezrin
D036002:ADP Ribose Transferases; D000199:Actins; D001427:Bacterial Toxins; D003598:Cytoskeletal Proteins; D008565:Membrane Proteins; D008840:Microfilament Proteins; D011493:Protein Kinase C; D011994:Recombinant Proteins; D037521:Virulence Factors; D000099149:Ezrin; C071058:moesin; C071776:radixin; C444820:exoenzyme S