Rho-kinase is involved in the sustained phosphorylation of myosin and the irreversible platelet aggregation induced by PAR1 activating peptide.
We have addressed the role of Rho-kinase in the different steps of thrombin receptor agonist peptide (TRAP)-induced platelet activation. Interestingly, under physiological conditions, incubation of platelets with increasing concentrations of the specific Rho-kinase inhibitor Y-27632 resulted in a dose-dependent reversion of the aggregation induced by 10 microM TRAP, without affecting serotonin secretion. Addition of Y-27632 after three minutes of TRAP stimulation, when the maximal aggregation was reached, resulted in a rapid disaggregation of platelets. Accordingly, the early peak of myosin light chain (MLC) phosphorylation induced by TRAP was not affected by Y-27632 but its sustained phosphorylation, observed during the irreversible phase of aggregation, was dependent of Rho-kinase activity. The rapid decrease in MLC phosphorylation upon Y-27632 treatment correlated well with the specific disappearance of myosin heavy chain from the cytoskeleton and preceded platelet disaggregation. Finally, we provide evidence that secreted ADP, known to play a key role in TRAP-induced irreversible phase of aggregation, was involved in the sustained MLC phosphorylation through Rho-kinase and could be replaced by epinephrine.
Missy
K
K
INSERM, Unité 326, Institut Fédératif de Recherche 30, H pital Purpan, Toulouse, France.
Plantavid
M
M
Pacaud
P
P
Viala
C
C
Chap
H
H
Payrastre
B
B
D000244:Adenosine Diphosphate / Q000378:metabolism / Q000494:pharmacology; D000577:Amides / Q000494:pharmacology; D001792:Blood Platelets / Q000166:cytology / Q000187:drug effects; D003599:Cytoskeleton / Q000187:drug effects / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D007700:Kinetics; D009218:Myosins / Q000187:drug effects / Q000378:metabolism*; D010446:Peptide Fragments / Q000494:pharmacology*; D010766:Phosphorylation; D010974:Platelet Aggregation / Q000187:drug effects*; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000502:physiology*; D011725:Pyridines / Q000494:pharmacology; D054460:rho-Associated Kinases
D000577:Amides; D004791:Enzyme Inhibitors; D047908:Intracellular Signaling Peptides and Proteins; D010446:Peptide Fragments; D011725:Pyridines; C108830:Y 27632; C086309:thrombin receptor peptide SFLLRNP; D000244:Adenosine Diphosphate; D017346:Protein Serine-Threonine Kinases; D054460:rho-Associated Kinases; D009218:Myosins