title: Molecular basis of platelet activation by an alphaIIbbeta3-CHAMPS peptide.
abstract: BACKGROUND
  A novel method, known as computed helical anti-membrane protein (CHAMP), for the design of peptides that bind with high affinity and selectivity to transmembrane helices was recently described and illustrated using peptides that bind alphaIIb- and alphav-integrin subunits, which induce selective activation of integrins alphaIIbbeta3 and alphavbeta3, respectively.
  
  
  OBJECTIVE
  In the present study, we have investigated the ability of an alphaIIb-CHAMPS peptide (termed integrin-activatory-peptide or IAP) to stimulate protein tyrosine phosphorylation and aggregation in human and mouse platelets.
  
  
  METHODS
  The ability of IAP to stimulate platelet aggregation and dense granule secretion was measured in washed preparations of human and mouse platelets. Samples were taken for measurement of tyrosine phosphorylation.
  
  
  RESULTS
  IAP stimulates robust tyrosine phosphorylation of the tyrosine kinase Syk and the FcR gamma-chain, but only weak phosphorylation of PLCgamma2. Aggregation to low but not high concentrations of IAP is reduced in the presence of the Src kinase inhibitor, PP1, or by inhibitors of the two feedback agonists, ADP and thromboxane A(2) (TxA(2)) suggesting that activation is reinforced by Src kinase-driven release of ADP and TxA(2). Unexpectedly, aggregation by IAP is only partially inhibited in human and mouse platelets deficient in integrin alphaIIbbeta3. Further, IAP induces partial aggregation of formaldehyde-fixed platelets.
  
  
  CONCLUSIONS
  The present study demonstrates that the alphaIIb-CHAMPS peptide induces platelet activation through integrin alphaIIbbeta3-dependent and independent pathways with the former mediating tyrosine phosphorylation of FcR gamma-chain and Syk. The use of the alphaIIb-CHAMPS peptide to study integrin alphaIIbbeta3 function is compromised by non-integrin-mediated effects.
authors.affiliation: Centre for Cardiovascular Sciences, Institute for Biomedical Research, Division of Medical Sciences, The Medical School, University of Birmingham, Birmingham, UK.
authors.forename: B
authors.initials: B
authors.lastname: Grygielska
authors.forename: C E
authors.initials: CE
authors.lastname: Hughes
authors.forename: S P
authors.initials: SP
authors.lastname: Watson
mesh_terms: D000818:Animals; D015195:Drug Design; D006801:Humans; D039302:Integrin alphaVbeta3 / Q000819:agonists*; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism; D051379:Mice; D010455:Peptides / Q000737:chemistry / Q000494:pharmacology*; D010766:Phosphorylation / Q000187:drug effects; D015539:Platelet Activation / Q000187:drug effects*; D010974:Platelet Aggregation / Q000187:drug effects; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000819:agonists*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D017452:Receptors, IgG / Q000378:metabolism; D000072377:Syk Kinase
chemical_list: D039302:Integrin alphaVbeta3; D047908:Intracellular Signaling Peptides and Proteins; D010455:Peptides; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D017452:Receptors, IgG; D011505:Protein-Tyrosine Kinases; C000606896:SYK protein, human; D000072377:Syk Kinase; C000606897:Syk protein, mouse