Effect of pertussis toxin on the phosphodiesteratic cleavage of the polyphosphoinositides by guanosine 5'-O-thiotriphosphate and thrombin in permeabilized human platelets.
It has recently been shown in this laboratory that permeabilization of human platelets with 15-25 micrograms/ml saponin allows ADP-ribosylation by pertussis toxin of the alpha i-subunit of Gi (Ni), a guanine nucleotide-binding regulatory protein. The same assay conditions have been used to determine phospholipase C in permeabilized platelets. Guanosine 5'-O-thiotriphosphate- (GTP[gamma S]-) activated phospholipase C in permeabilized platelets whose inositol phospholipids were prelabeled with [3H]inositol. Phospholipase C activity was measured by [3H]polyphosphoinositide decreases and formation of [3H]inositol bisphosphate and [3H]inositol trisphosphate. Prostacyclin, cyclic AMP or pretreatment of permeabilized platelets with pertussis toxin did not alter this effect under conditions in which the alpha i-subunit was effectively ADP-ribosylated by pertussis toxin. This information indicated that ADP-ribosylation of Gi-protein was not directly related to activation or inhibition of platelet phospholipase C by GTP [gamma S]. Thrombin also activated phospholipase C in permeabilized platelets and, surprisingly, this action was enhanced by pertussis toxin pretreatment. This indicates that ADP-ribosylation of Gi-protein facilitates the action of thrombin on phospholipase C.
D000246:Adenosine Diphosphate Ribose / Q000097:blood; D001792:Blood Platelets / Q000201:enzymology*; D002118:Calcium / Q000494:pharmacology; D002463:Cell Membrane Permeability / Q000187:drug effects; D004789:Enzyme Activation / Q000187:drug effects; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006160:Guanosine Triphosphate / Q000031:analogs & derivatives* / Q000494:pharmacology; D006801:Humans; D006868:Hydrolysis; D037342:Pertussis Toxin; D018129:Phosphatidylinositol Phosphates; D010716:Phosphatidylinositols / Q000097:blood*; D012503:Saponins / Q000494:pharmacology; D013873:Thionucleotides / Q000494:pharmacology*; D013917:Thrombin / Q000494:pharmacology*; D010738:Type C Phospholipases / Q000097:blood*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology*
D018129:Phosphatidylinositol Phosphates; D010716:Phosphatidylinositols; D012503:Saponins; D013873:Thionucleotides; D010566:Virulence Factors, Bordetella; D000246:Adenosine Diphosphate Ribose; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006160:Guanosine Triphosphate; D037342:Pertussis Toxin; D010738:Type C Phospholipases; D013917:Thrombin; D002118:Calcium