Ezrin/radixin/moesin proteins are high affinity targets for ADP-ribosylation by Pseudomonas aeruginosa ExoS.
Pseudomonas aeruginosa ExoS is a bifunctional type III-secreted cytotoxin. The N terminus (amino acids 96-233) encodes a GTPase-activating protein activity, whereas the C terminus (amino acids 234-453) encodes a factor-activating ExoS-dependent ADP-ribosyltransferase activity. The GTPase-activating protein activity inactivates the Rho GTPases Rho, Rac, and Cdc42 in cultured cells and in vitro, whereas the ADP-ribosylation by ExoS is poly-substrate-specific and includes Ras as an early target for ADP-ribosylation. Infection of HeLa cells with P. aeruginosa producing a GTPase-activating protein-deficient form of ExoS rounded cells, indicating the ADP-ribosyltransferase domain alone is sufficient to elicit cytoskeletal changes. Examination of substrates modified by type III-delivered ExoS identified a 70-kDa protein as an early and predominant target for ADP-ribosylation. Matrix-assisted laser desorption ionization mass spectroscopy identified this protein as moesin, a member of the ezrin/radixin/moesin (ERM) family of proteins. ExoS ADP-ribosylated recombinant moesin at a linear velocity that was 5-fold faster and with a K(m) that was 2 orders of magnitude lower than Ras. Moesin homologs ezrin and radixin were also ADP-ribosylated, indicating the ERMs collectively represent high affinity targets of ExoS. Type III delivered ExoS ADP-ribosylated moesin and ezrin (and/or radixin) in cultured HeLa cells. The ERM proteins contribute to cytoskeleton dynamics, and the ability of ExoS to ADP-ribosylate the ERM proteins links ADP-ribosylation with the cytoskeletal changes associated with ExoS intoxication.
Maresso
Anthony W
AW
Microbiology and Molecular Genetics, Medical College of Wisconsin, Milwaukee, Wisconsin 53226, USA.
Baldwin
Michael R
MR
Barbieri
Joseph T
JT
D036002:ADP Ribose Transferases / Q000378:metabolism*; D000199:Actins / Q000378:metabolism; D000244:Adenosine Diphosphate / Q000378:metabolism*; D001427:Bacterial Toxins / Q000378:metabolism*; D001798:Blood Proteins / Q000737:chemistry*; D003598:Cytoskeletal Proteins / Q000737:chemistry*; D003599:Cytoskeleton / Q000378:metabolism; D003603:Cytotoxins / Q000737:chemistry; D004926:Escherichia coli / Q000378:metabolism; D020558:GTP Phosphohydrolases / Q000737:chemistry / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D008565:Membrane Proteins / Q000737:chemistry*; D008840:Microfilament Proteins / Q000737:chemistry*; D010750:Phosphoproteins / Q000737:chemistry*; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011550:Pseudomonas aeruginosa / Q000378:metabolism*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013744:Tetanus Toxin / Q000737:chemistry; D013997:Time Factors; D014162:Transfection; D000099149:Ezrin
D036002:ADP Ribose Transferases; D000199:Actins; D000244:Adenosine Diphosphate; D001427:Bacterial Toxins; D001798:Blood Proteins; D003598:Cytoskeletal Proteins; D003603:Cytotoxins; D020558:GTP Phosphohydrolases; D008565:Membrane Proteins; D008840:Microfilament Proteins; D010750:Phosphoproteins; D013744:Tetanus Toxin; D000099149:Ezrin; C071058:moesin; C071776:radixin; C026167:tetanolysin; C444820:exoenzyme S