Lysophosphatidic acid stimulation of platelets rapidly induces Ca2+-dependent dephosphorylation of cofilin that is independent of dense granule secretion and aggregation.
Cofilin is an actin dynamizing protein and inactivated after Ser3 phosphorylation by LIM-kinases (LIMKs). We studied whether in platelets stimulated by lysophosphatidic acid (LPA), Rho-kinase or p21-activated kinase (PAK) mediates LIMK-1 activation leading to subsequent phosphorylation and inactivation of cofilin and the increase of F-actin. During LPA (0.1 microM)-induced shape change, a rapid Rho-kinase activation and a slower activation of PAK were observed. Rho-kinase activation led to rapid LIMK-1 (Thr508) phosphorylation. Despite of LIMK-1 activation, cofilin net phosphorylation was not increased. Cofilin rapidly associated with F-actin and preceded the F-actin increase. Pretreatment with the Rho-kinase inhibitor Y-27632 inhibited LIMK-1 phosphorylation, unmasked cofilin dephosphorylation and inhibited the reversible F-actin increase during shape change. In the presence of fibrinogen, LPA (10 microM) induced ATP-secretion from dense granules and aggregation, and cofilin was rapidly dephosphorylated and then rephosphorylated in a Rho-kinase/LIMK-1-dependent manner. In the absence of fibrinogen, cofilin de- and rephosphorylation after LPA (10 microM) was unchanged, but secretion and aggregation were absent. Cofilin dephosphorylation was completely blocked by BAPTA-AM indicating that it was mediated by an increase of cytosolic Ca(2+). We conclude that in LPA-stimulated platelets, Rho-kinase-dependent LIMK-1 activation mediates the F-actin increase during shape change without enhancing cofilin net phosphorylation. However, a rapid dephosphorylation of cofilin occurs during secretion and aggregation, which is Ca(2+)-dependent, upstream of secretion and aggregation and might regulate these platelet responses.
Pandey
Dharmendra
D
Institute for Prevention of Cardiovascular Diseases, Klinikum Innenstadt, University of Munich, Pettenkoferstrasse 9, D-80336 Munich, Germany.
Goyal
Pankaj
P
Siess
Wolfgang
W
D051339:Actin Depolymerizing Factors / Q000378:metabolism*; D000577:Amides / Q000494:pharmacology; D001792:Blood Platelets / Q000187:drug effects / Q000502:physiology*; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects; D015550:Cell Degranulation / Q000187:drug effects; D048430:Cell Shape / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D008246:Lysophospholipids / Q000494:pharmacology*; D010766:Phosphorylation / Q000187:drug effects; D010974:Platelet Aggregation / Q000187:drug effects; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D011725:Pyridines / Q000494:pharmacology; D054460:rho-Associated Kinases
D051339:Actin Depolymerizing Factors; D000577:Amides; D004791:Enzyme Inhibitors; D047908:Intracellular Signaling Peptides and Proteins; D008246:Lysophospholipids; D011725:Pyridines; C108830:Y 27632; D017346:Protein Serine-Threonine Kinases; D054460:rho-Associated Kinases; D002118:Calcium