Multiple pathways of thrombin-induced platelet activation differentiated by desensitization and a thrombin exosite inhibitor.
Recently a thrombin receptor with a unique mechanism of activation was cloned from a megakaryocyte-like cell line (Vu et al., Cell 64:1057-1068, 1991). Thrombin cleaves a portion of this receptor creating a new N-terminus that acts as a "tethered-ligand" to activate the receptor. A thrombin receptor activating peptide (SFLLRNPNDKYEPF) homologous to the new N-terminus was shown to activate platelets. We synthesized this peptide and demonstrated that it desensitized platelets to activation by low concentrations of alpha-thrombin but not gamma-thrombin. We also synthesized a thrombin exosite inhibitor (BMS 180742) that inhibited platelet aggregation induced by low, but not high, concentrations of alpha-thrombin. In contrast, a thrombin active site inhibitor, N alpha-(2-naphthylsulfonyl-glycyl)-D,L-amidinophenylalanylpiperi dide, competitively inhibited thrombin-induced platelet aggregation. We conclude that thrombin-induced platelet activation is mediated by at least two pathways: one activated by low concentrations of alpha-thrombin and blocked by a thrombin exosite inhibitor that appears to be coupled to the "tethered-ligand" thrombin receptor, and another that is stimulated by higher concentrations of alpha-thrombin and by gamma-thrombin and does not require the thrombin exosite for activation. Both pathways are blocked by a thrombin active site inhibitor.
Seiler
S M
SM
Department of Cardiovascular Biochemistry, Bristol-Myers Squibb Pharmaceutical Research Institute, Princeton, NJ 08543.
Goldenberg
H J
HJ
Michel
I M
IM
Hunt
J T
JT
Zavoico
G B
GB
D000595:Amino Acid Sequence; D004361:Drug Tolerance; D006801:Humans; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000494:pharmacology*; D010455:Peptides / Q000494:pharmacology*; D015539:Platelet Activation / Q000187:drug effects*; D015398:Signal Transduction; D013917:Thrombin / Q000037:antagonists & inhibitors / Q000494:pharmacology*
D010446:Peptide Fragments; D010455:Peptides; C072032:thrombin receptor peptide (42-55); C072028:BMS 180742; D013917:Thrombin