PKCdelta regulates collagen-induced platelet aggregation through inhibition of VASP-mediated filopodia formation.
Protein kinase Cdelta (PKCdelta) has been shown by pharmacologic approaches to negatively regulate collagen-induced platelet aggregation. Here we addressed the molecular and cellular mechanisms underlying this negative regulation. Using PKCdelta-/- platelets, we show that the mechanism did not involve altered inside-out signaling to integrin alpha(IIb)beta3 and did not affect early signaling events downstream of GPVI, because there was no difference in tyrosine phosphorylation of PLCgamma2 between wild-type and PKCdelta-/- platelets. There was also no increase in secretion of dense granule content, in contrast to studies using rottlerin where secretion was enhanced. Importantly, however, there was marked enhancement of filopodia generation in PKCdelta-/- platelets upon adhesion to collagen compared with wild-type platelets. Filopodia play an essential role regulating adhesive events leading to platelet aggregation by increasing platelet-platelet contact. We show that the critical effector for PKCdelta is vasodilator-stimulated phosphoprotein (VASP), a major regulator of actin cytoskeleton dynamics. PKCdelta physically interacts with VASP constitutively and regulates its phosphorylation on Ser157. In VASP-/- platelets, the enhancement of filopodia generation, actin polymerization, and platelet aggregation by rottlerin is ablated. PKCdelta is therefore a critical negative regulator of filopodia, and hence platelet aggregation, through a functional interaction with the actin organizer VASP.
Pula
Giordano
G
Department of Pharmacology, School of Medical Sciences, University of Bristol, United Kingdom.
Schuh
Kai
K
Nakayama
Keiko
K
Nakayama
Keiichi I
KI
Walter
Ulrich
U
Poole
Alastair W
AW
D000098:Acetophenones / Q000494:pharmacology; D000199:Actins / Q000378:metabolism; D000818:Animals; D001578:Benzopyrans / Q000494:pharmacology; D001792:Blood Platelets / Q000201:enzymology*; D015815:Cell Adhesion Molecules / Q000172:deficiency / Q000378:metabolism*; D003094:Collagen / Q000378:metabolism / Q000494:pharmacology*; D003599:Cytoskeleton / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D051379:Mice; D018345:Mice, Knockout; D008840:Microfilament Proteins / Q000172:deficiency / Q000378:metabolism*; D051966:Phospholipase C gamma / Q000378:metabolism; D010750:Phosphoproteins / Q000172:deficiency / Q000378:metabolism*; D010974:Platelet Aggregation / Q000187:drug effects; D010980:Platelet Membrane Glycoproteins / Q000378:metabolism; D051745:Protein Kinase C-delta / Q000172:deficiency / Q000378:metabolism*; D011554:Pseudopodia / Q000378:metabolism; D022142:Secretory Vesicles / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D000098662:Vasodilator-Stimulated Phosphoprotein
D000098:Acetophenones; D000199:Actins; D001578:Benzopyrans; D015815:Cell Adhesion Molecules; D004791:Enzyme Inhibitors; D008840:Microfilament Proteins; D010750:Phosphoproteins; D010980:Platelet Membrane Glycoproteins; C098341:platelet membrane glycoprotein VI; D000098662:Vasodilator-Stimulated Phosphoprotein; D003094:Collagen; C085746:rottlerin; D051745:Protein Kinase C-delta; D051966:Phospholipase C gamma