Rho protein inhibition blocks protein kinase C translocation and activation.
Small GTP-binding proteins of the Ras and Rho family participate in various important signalling pathways. Large clostridial cytotoxins inactivate GTPases by UDP-glucosylation. Using Clostridium difficile toxin B-10463 (TcdB) for inactivation of Rho proteins (RhoA/Rac/Cdc42) and Clostridium sordellii lethal toxin-1522 (TcsL) for inactivation of Ras-proteins (Ras/Rac/Ral, Rap) the role of these GTPases in protein kinase C (PKC) stimulation was studied. Phorbol-myristate-acetate (PMA) induced a rapid PKC translocation to and activation in the particulate cell fraction as determined by PKC-activity measurements and Western blots for PKC alpha. These effects were blocked by TcdB inhibiting Rho proteins in endothelial cells, but not in TcsL-treated cells (i.e., cells without Ras activity), suggesting that Rho GTPases (RhoA and/or Cdc42) are the most likely GTP-binding proteins responsible for PKC activation. The Rho requirement for PKC activation/translocation was also verified for human epithelial cells and for lipopolysaccharide-stimulated endothelial cells. In summary, the data presented indicate that Rho protein inhibition blocked PKC translocation/activation in endothelial and epithelial cells.
Hippenstiel
S
S
Department of Internal Medicine, Justus-Liebig-University, Giessen, Germany.
Kratz
T
T
Krüll
M
M
Seybold
J
J
von Eichel-Streiber
C
C
Suttorp
N
N
D000818:Animals; D001980:Bronchi / Q000187:drug effects / Q000378:metabolism; D018797:Cell Cycle Proteins / Q000037:antagonists & inhibitors*; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000187:drug effects / Q000378:metabolism; D004789:Enzyme Activation; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000037:antagonists & inhibitors*; D006801:Humans; D008070:Lipopolysaccharides / Q000494:pharmacology; D011493:Protein Kinase C / Q000378:metabolism*; D013552:Swine; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D020764:cdc42 GTP-Binding Protein; D018631:ras Proteins / Q000037:antagonists & inhibitors*; D020742:rhoA GTP-Binding Protein
D018797:Cell Cycle Proteins; D008070:Lipopolysaccharides; D011493:Protein Kinase C; D019204:GTP-Binding Proteins; D020764:cdc42 GTP-Binding Protein; D018631:ras Proteins; D020742:rhoA GTP-Binding Protein; D013755:Tetradecanoylphorbol Acetate