title: Rap1b is critical for glycoprotein VI-mediated but not ADP receptor-mediated alpha2beta1 activation.
abstract: BACKGROUND
  The platelet alpha2beta1 integrin functions as both an adhesion and signaling receptor upon exposure to collagen. Recent studies have indicated that alpha2beta1 function can be activated via inside-out signaling, similar to the prototypical platelet integrin alphaIIbbeta3. However, signaling molecules that regulate alpha2beta1 activation in platelets are not well defined. A strong candidate molecule is the small GTPase Rap1b, the dominant platelet isoform of Rap1, which regulates alphaIIbbeta3 activation.
  
  
  OBJECTIVE
  We hypothesized that Rap1b positively regulates alpha2beta1 during agonist-induced platelet activation.
  
  
  METHODS
  To test whether Rap1b activates alpha2beta1 downstream of glycoprotein (GP)VI or other platelet receptors, we stimulated platelets purified from Rap1b-/- or wild-type mice with diverse agonists and measured alpha2beta1 activation using fluorescein isothiocyanate-labeled monomeric collagen. We also examined the role of Rap1b in outside-in signaling pathways by analyzing adhesion and spreading of Rap1b-/- or wild-type platelets on monomeric, immobilized collagen. Finally, we monitored the activation status of related Rap GTPases to detect changes in signaling pathways potentially associated with Rap1b-mediated events.
  
  
  RESULTS
  Rap1b-/- platelets displayed comparable ADP-induced or thrombin-induced alpha2beta1 activation as wild-type platelets, but reduced convulxin-dependent alpha2beta1 activation. Rap1b-/- platelets exhibited increased spreading on immobilized collagen but similar adhesion to immobilized collagen compared to wild-type platelets. Rap1b-/- platelets also showed Rap1a and Rap2 activation upon agonist stimulation, possibly revealing functional compensation among Rap family members.
  
  
  CONCLUSIONS
  Rap1b is required for maximal GPVI-induced but not ADP-induced activation of alpha2beta1 in murine platelets.
authors.affiliation: Department of Pharmacology, The University of North Carolina at Chapel Hill, Chapel Hill, NC 27599-7260, USA.
authors.forename: Z
authors.initials: Z
authors.lastname: Wang
authors.forename: S P
authors.initials: SP
authors.lastname: Holly
authors.forename: M K
authors.initials: MK
authors.lastname: Larson
authors.forename: J
authors.initials: J
authors.lastname: Liu
authors.forename: W
authors.initials: W
authors.lastname: Yuan
authors.forename: M
authors.initials: M
authors.lastname: Chrzanowska-Wodnicka
authors.forename: G C
authors.initials: GC
authors.lastname: White
authors.forename: L V
authors.initials: LV
authors.lastname: Parise
mesh_terms: D000818:Animals; D001792:Blood Platelets / Q000166:cytology / Q000378:metabolism; D048430:Cell Shape; D003094:Collagen; D038982:Integrin alpha2beta1 / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D010973:Platelet Adhesiveness; D010980:Platelet Membrane Glycoproteins / Q000378:metabolism*; D018048:Receptors, Purinergic P2 / Q000378:metabolism*; D015398:Signal Transduction; D020668:rap GTP-Binding Proteins / Q000172:deficiency / Q000502:physiology*
chemical_list: D038982:Integrin alpha2beta1; D010980:Platelet Membrane Glycoproteins; D018048:Receptors, Purinergic P2; C098341:platelet membrane glycoprotein VI; D003094:Collagen; C492136:Rap1b protein, mouse; D020668:rap GTP-Binding Proteins