Diacylglycerol elevations in control platelets are unaccompanied by pleckstrin phosphorylation. Implications for the role of diacylglycerol in platelet activation.
Several laboratories have reported that diacylglycerol levels in human platelets (approximately 100 pmol/10(9) platelets) increased severalfold in response to 0.5-1 U/ml thrombin. We report here fluctuations in diacylglycerol mass in control platelets, the magnitude of which were 60-90% of that measured in platelets treated with 0.2-0.5 U/ml of thrombin. These control platelets were not activated by such criteria as absence of aggregation, secretion, phosphatidic acid production and phosphorylation of the protein kinase C substrate, pleckstrin. Thrombin treatment evoked all of the above responses. Analysis of the diacylglycerol molecular species by reverse-phase HPLC of the dimethylated, phosphorylated derivatives showed that all of the molecular species that were present in control platelets were also present in thrombin-treated platelets. Most of the species appeared to fluctuate at random in control platelets with the exception of 1-stearoyl-2-arachidonoyl-sn-glycerol which was more or less stable and increased severalfold over control values only upon thrombin treatment. Furthermore, only this species accumulated as [32P]phosphorylated PtdOH in thrombin-treated platelets prelabelled with [32P]Pi. Our findings show that, in platelets, elevation of diacylglycerol molecular species other than the 1-stearoyl-2-arachidonoyl species occurs, but these changes are not necessarily linked to activation of protein kinase C as measured by pleckstrin phosphorylation which was observed only upon elevation of 1-stearoyl-2-arachidonoyl-sn-glycerol.
Fukami
M H
MH
Department of Biochemistry and Molecular Biology, University of Bergen, Norway.
Holmsen
H
H
D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism*; D001798:Blood Proteins / Q000378:metabolism*; D004075:Diglycerides / Q000097:blood*; D006358:Hot Temperature; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D010750:Phosphoproteins; D010766:Phosphorylation; D015539:Platelet Activation; D013917:Thrombin / Q000494:pharmacology
D001798:Blood Proteins; D004075:Diglycerides; D010750:Phosphoproteins; C039077:platelet protein P47; D013917:Thrombin