Pflügers Archiv - European Journal of Physiology

, Volume 457, Issue 5, pp 1173–1185

Rac1 is essential for phospholipase C-γ2 activation in platelets

Authors

  • Irina Pleines
    • Rudolf Virchow Center for Experimental BiomedicineUniversity of Würzburg
  • Margitta Elvers
    • Rudolf Virchow Center for Experimental BiomedicineUniversity of Würzburg
  • Amrei Strehl
    • Rudolf Virchow Center for Experimental BiomedicineUniversity of Würzburg
  • Miroslava Pozgajova
    • Rudolf Virchow Center for Experimental BiomedicineUniversity of Würzburg
  • David Varga-Szabo
    • Rudolf Virchow Center for Experimental BiomedicineUniversity of Würzburg
  • Frauke May
    • Rudolf Virchow Center for Experimental BiomedicineUniversity of Würzburg
  • Anna Chrostek-Grashoff
    • Cardiovascular Research CenterUniversity of Virginia Health System
  • Cord Brakebusch
    • BRIC, Biomedical InstituteUniversity of Copenhagen
    • Rudolf Virchow Center for Experimental BiomedicineUniversity of Würzburg
    • Institute of Clinical Biochemistry and PathobiochemistryUniversity of Würzburg
Signaling and Cell Physiology

DOI: 10.1007/s00424-008-0573-7

Cite this article as:
Pleines, I., Elvers, M., Strehl, A. et al. Pflugers Arch - Eur J Physiol (2009) 457: 1173. doi:10.1007/s00424-008-0573-7

Abstract

Platelet activation at sites of vascular injury is triggered through different signaling pathways leading to activation of phospholipase (PL) Cβ or PLCγ2. Active PLCs trigger Ca2+ mobilization and entry, which is a prerequisite for adhesion, secretion, and thrombus formation. PLCβ isoenzymes are activated downstream of G protein-coupled receptors (GPCRs), whereas PLCγ2 is activated downstream of immunoreceptor tyrosine-based activation motif (ITAM)-coupled receptors, such as the major platelet collagen receptor glycoprotein (GP) VI or CLEC-2. The mechanisms underlying PLC regulation are not fully understood. An involvement of small GTPases of the Rho family (Rho, Rac, Cdc42) in PLC activation has been proposed but this has not been investigated in platelets. We here show that murine platelets lacking Rac1 display severely impaired GPVI- or CLEC-2-dependent activation and aggregation. This defect was associated with impaired production of inositol 1,4,5-trisphosphate (IP3) and intracellular calcium mobilization suggesting inappropriate activation of PLCγ2 despite normal tyrosine phosphorylation of the enzyme. Rac1−/− platelets displayed defective thrombus formation on collagen under flow conditions which could be fully restored by co-infusion of ADP and the TxA2 analog U46619, indicating that impaired GPVI-, but not G-protein signaling, was responsible for the observed defect. In line with this, Rac1−/− mice were protected in two collagen-dependent arterial thrombosis models. Together, these results demonstrate that Rac1 is essential for ITAM-dependent PLCγ2 activation in platelets and that this is critical for thrombus formation in vivo.

Keywords

Ca2+ mobilizationG-proteinIP3Phospholipase CPlateletsRac1

Abbreviations

BSA

bovine serum albumin

CLEC-2

C-type lectin-like receptor

CRP

collagen related peptide

CVX

convulxin

FACS

fluoresence activated cell sorting

FcR

Fc receptor

FITC

fluoresceine isothiocyanate

GP

glycoprotein

HRP

horseradish peroxidase

Ig

immunoglobulin

PAGE

polyacrylamide gel electrophoresis

PLCγ2

phospholipase Cγ2

PMA

phorbol 12-myristate 13-acetate

prp

platelet rich plasma

PVDF

polyvinylidene difluoride

RC

rhodocytin

SDS

sodium dodecyl sulfate

TxA2

thromboxane A2

vWF

von Willebrand factor

Copyright information

© Springer-Verlag 2008