A role for cofilin in the activation of store-operated calcium entry by de novo conformational coupling in human platelets.
Store-operated Ca2+ entry (SOCE) is a major mechanism for Ca2+ influx in platelets and other cells. De novo conformational coupling between elements in the plasma membrane and Ca2+ stores, where the actin cytoskeleton plays an important regulatory role, has been proposed as the most likely mechanism to activate SOCE in platelets. Here we have examined for the first time changes in platelet F-actin levels on a subsecond time scale. Using stopped-flow fluorimetry and a quenched-flow approach, we provide evidence for the involvement of cofilin in actin filament reorganization and SOCE in platelets. Thrombin (0.1 U/mL) evoked an initial decrease in F-actin that commenced within 0.1 second and reached a minimum 0.9 second after stimulation, prior to the activation of SOCE. F-actin then increased, exceeding basal levels approximately 2.5 seconds after stimulation. Thrombin also induced cofilin dephosphorylation and activation, which paralleled the changes observed in F-actin, and rapid Btk activation. Inhibition of cofilin dephosphorylation by LFM-A13 resulted in the loss of net actin depolymerization and an increased delay in SOCE initiation. These results suggest that cofilin is important for the rapid actin remodeling necessary for the activation of SOCE in platelets through de novo conformational coupling.
Redondo
Pedro C
PC
Department of Physiology, University of Extremadura, Cáceres, Spain.
Harper
Matthew T
MT
Rosado
Juan A
JA
Sage
Stewart O
SO
D008841:Actin Cytoskeleton / Q000378:metabolism; D051339:Actin Depolymerizing Factors / Q000378:metabolism*; D000199:Actins / Q000378:metabolism; D000077329:Agammaglobulinaemia Tyrosine Kinase; D000577:Amides / Q000494:pharmacology; D001792:Blood Platelets / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects*; D002462:Cell Membrane / Q000378:metabolism; D005470:Fluorometry; D006490:Hemostatics / Q000378:metabolism / Q000494:pharmacology*; D006801:Humans; D009570:Nitriles / Q000494:pharmacology; D011487:Protein Conformation / Q000187:drug effects; D011499:Protein Processing, Post-Translational / Q000187:drug effects / Q000502:physiology; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D013917:Thrombin / Q000378:metabolism / Q000494:pharmacology*
D051339:Actin Depolymerizing Factors; D000199:Actins; D000577:Amides; D006490:Hemostatics; C118451:LFM A13; D009570:Nitriles; D011505:Protein-Tyrosine Kinases; D000077329:Agammaglobulinaemia Tyrosine Kinase; C000625949:BTK protein, human; D013917:Thrombin; D002118:Calcium