title: GPIbalpha-selective activation of platelets induces platelet signaling events comparable to GPVI activation events.
abstract: Platelet glycoprotein (GP)Ib-IX-V, which binds von Willebrand factor (VWF), and GPVI, which binds collagen, form an adhesion-signaling complex on platelets and mediate platelet adhesion in flowing blood. Platelet activation following engagement of GPIb-IX-V/GPVI by VWF/collagen is critical for initiation and development of a protective thrombus across a site of damaged or exposed endothelium. We examined platelet aggregation and signaling following selective engagement of platelet GPIbalpha (the major ligand-binding subunit of GPIb-IX-V) by a multivalent surface-expressed GPIbalpha-binding VWF-A1 domain on COS-7 cells. COS-7 cells expressing the VWF-A1 domain containing an R543W mutation (a gain-of-function mutation found in Type 2B von Willebrand's Disease) were used as a selective agonist for GPIb-IX-V. When incubated in a cell-to-platelet ratio of up to 1 : 1200, VWF-A1/R543W cells caused rapid, spontaneous aggregation of washed platelets that was GPIbalpha- and alpha(IIb)beta(3)-dependent (blocked by inhibitory anti-VWF-A1, anti-GPIbalpha and anti-alpha(IIb)beta(3) antibodies). Platelet aggregation was also sensitive to inhibitors of Src, phosphoinositide 3-kinase (PI3-kinase) or Syk, confirming a role for these proteins in GPIbalpha-mediated signal transduction. Platelet tyrosine phosphorylation patterns and specific tyrosine phosphorylation of Syk after GPIbalpha engagement by VWF-A1/R543W was comparable to that induced by engagement of GPVI by collagen or collagen-related peptide (CRP). These data indicate signaling events triggered by specific ligation of GPIbalpha can lead to robust platelet activation and help define GPIb-IX-V as both an adhesion and signaling receptor on platelets.
authors.affiliation: The Australian Centre for Blood Diseases, Monash University, Melbourne, Australia. elizabeth.gardiner@med.monash.edu.au
authors.forename: Elizabeth E
authors.initials: EE
authors.lastname: Gardiner
authors.forename: Jane F
authors.initials: JF
authors.lastname: Arthur
authors.forename: Yang
authors.initials: Y
authors.lastname: Shen
authors.forename: Denuja
authors.initials: D
authors.lastname: Karunakaran
authors.forename: Lynley A
authors.initials: LA
authors.lastname: Moore
authors.forename: Jan Schulte
authors.initials: JS
authors.lastname: Am Esch
authors.forename: Robert K
authors.initials: RK
authors.lastname: Andrews
authors.forename: Michael C
authors.initials: MC
authors.lastname: Berndt
mesh_terms: D000818:Animals; D001792:Blood Platelets / Q000378:metabolism*; D019556:COS Cells; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D002522:Chlorocebus aethiops; D004791:Enzyme Inhibitors / Q000378:metabolism; D006801:Humans; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D010455:Peptides / Q000235:genetics / Q000378:metabolism; D015539:Platelet Activation; D010974:Platelet Aggregation / Q000502:physiology; D019038:Platelet Glycoprotein GPIb-IX Complex; D010980:Platelet Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D015398:Signal Transduction / Q000502:physiology*; D014841:von Willebrand Factor / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D002352:Carrier Proteins; D004791:Enzyme Inhibitors; D008562:Membrane Glycoproteins; D010455:Peptides; D019038:Platelet Glycoprotein GPIb-IX Complex; D010980:Platelet Membrane Glycoproteins; C094990:adhesion receptor; C435908:collagen-related peptide; C098341:platelet membrane glycoprotein VI; D014841:von Willebrand Factor