Platelet shape change is mediated by both calcium-dependent and -independent signaling pathways. Role of p160 Rho-associated coiled-coil-containing protein kinase in platelet shape change

• Published in: The Journal of biological chemistry 1999-10, Vol.274 (40), p.28293-28300
• Main Authors: Paul, B Z, Daniel, J L, Kunapuli, S P
• Format: Article
• Language: English
• Subjects: 1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine - analogs & derivatives; 1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine - pharmacology; Amides - pharmacology; Blood Platelets - cytology; Calcium - metabolism; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Calcium-Calmodulin-Dependent Protein Kinases - metabolism; Egtazic Acid - analogs & derivatives; Enzyme Inhibitors - pharmacology; Humans; Intracellular Signaling Peptides and Proteins; Myosins - metabolism; Phosphorylation; Protein Kinase C - antagonists & inhibitors; Protein-Serine-Threonine Kinases - metabolism; Pyridines - pharmacology; Rho-associated coiled coil protein kinase; rho-Associated Kinases; Signal Transduction
• ISSN: 0021-9258
• DOI: 10.1074/jbc.274.40.28293

Platelets undergo shape change upon activation with agonists. During shape change, disc-shaped platelets turn into spiculated spheres with protruding filopodia. When agonist-induced cytosolic Ca(2+) increases were prevented using the cytosolic Ca(2+) chelator, 5, 5'-dimethyl-bis-(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (5, 5'-dimethyl-BAPTA), platelets still underwent shape change, although the onset was delayed and the initial rate was dramatically decreased. In the absence of cytosolic Ca(2+), agonist-stimulated myosin light chain phosphorylation was significantly inhibited. The myosin light chain was maximally phosphorylated at 2 s in control platelets compared with 30 s in 5,5'-dimethyl-BAPTA-treated platelets. ADP, thrombin, or U46619-induced Ca(2+)-independent platelet shape change was significantly reduced by staurosporine, a nonselective kinase inhibitor, by the selective p160 Rho-associated coiled-coil-containing protein kinase inhibitor Y-27632, or by HA 1077. Both Y-27632 and HA 1077 reduced peak levels of ADP-induced platelet shape change and myosin light chain phosphorylation in control platelets. In 5,5'-dimethyl-BAPTA-treated platelets, Y-27632 and HA 1077 completely abolished both ADP-induced platelet shape change and myosin light chain phosphorylation. Our results indicate that Ca(2+)/calmodulin-stimulated myosin light chain kinase and p160 Rho-associated coiled-coil-containing protein kinase independently contribute to myosin light chain phosphorylation and platelet shape change, through Ca(2+)-sensitive and Ca(2+)-insensitive pathways, respectively.