title: The Phosphatidylinositol 3,4,5-trisphosphate (PI(3,4,5)P3) Binder Rasa3 Regulates Phosphoinositide 3-kinase (PI3K)-dependent Integrin αIIbβ3 Outside-in Signaling.
abstract: The class I PI3K family of lipid kinases plays an important role in integrin αIIbβ3function, thereby supporting thrombus growth and consolidation. Here, we identify Ras/Rap1GAP Rasa3 (GAP1IP4BP) as a major phosphatidylinositol 3,4,5-trisphosphate-binding protein in human platelets and a key regulator of integrin αIIbβ3outside-in signaling. We demonstrate that cytosolic Rasa3 translocates to the plasma membrane in a PI3K-dependent manner upon activation of human platelets. Expression of wild-type Rasa3 in integrin αIIbβ3-expressing CHO cells blocked Rap1 activity and integrin αIIbβ3-mediated spreading on fibrinogen. In contrast, Rap1GAP-deficient (P489V) and Ras/Rap1GAP-deficient (R371Q) Rasa3 had no effect. We furthermore show that two Rasa3 mutants (H794L and G125V), which are expressed in different mouse models of thrombocytopenia, lack both Ras and Rap1GAP activity and do not affect integrin αIIbβ3-mediated spreading of CHO cells on fibrinogen. Platelets from thrombocytopenic mice expressing GAP-deficient Rasa3 (H794L) show increased spreading on fibrinogen, which in contrast to wild-type platelets is insensitive to PI3K inhibitors. Together, these results support an important role for Rasa3 in PI3K-dependent integrin αIIbβ3-mediated outside-in signaling and cell spreading.
authors.affiliation: From the School of Physiology, Pharmacology and Neuroscience, University of Bristol, Bristol, BS8 1TD, United Kingdom.
authors.forename: Anthony M
authors.initials: AM
authors.lastname: Battram
authors.affiliation: From the School of Physiology, Pharmacology and Neuroscience, University of Bristol, Bristol, BS8 1TD, United Kingdom.
authors.forename: Tom N
authors.initials: TN
authors.lastname: Durrant
authors.affiliation: From the School of Physiology, Pharmacology and Neuroscience, University of Bristol, Bristol, BS8 1TD, United Kingdom.
authors.forename: Ejaife O
authors.initials: EO
authors.lastname: Agbani
authors.affiliation: School of Biochemistry, University of Bristol, Bristol, BS8 1TD, United Kingdom.
authors.forename: Kate J
authors.initials: KJ
authors.lastname: Heesom
authors.affiliation: the McAllister Heart Institute, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27514.
authors.forename: David S
authors.initials: DS
authors.lastname: Paul
authors.affiliation: the McAllister Heart Institute, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27514.
authors.forename: Raymond
authors.initials: R
authors.lastname: Piatt
authors.affiliation: From the School of Physiology, Pharmacology and Neuroscience, University of Bristol, Bristol, BS8 1TD, United Kingdom.
authors.forename: Alastair W
authors.initials: AW
authors.lastname: Poole
authors.affiliation: School of Biochemistry, University of Bristol, Bristol, BS8 1TD, United Kingdom.
authors.forename: Peter J
authors.initials: PJ
authors.lastname: Cullen
authors.affiliation: the McAllister Heart Institute, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27514; Department of Biochemistry and Biophysics, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27514.
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Bergmeier
authors.affiliation: From the School of Physiology, Pharmacology and Neuroscience, University of Bristol, Bristol, BS8 1TD, United Kingdom.
authors.forename: Samantha F
authors.initials: SF
authors.lastname: Moore
authors.affiliation: From the School of Physiology, Pharmacology and Neuroscience, University of Bristol, Bristol, BS8 1TD, United Kingdom. Electronic address: i.hers@bristol.ac.uk.
authors.forename: Ingeborg
authors.initials: I
authors.lastname: Hers
mesh_terms: D019943:Amino Acid Substitution / Q000235:genetics; D000818:Animals; D001792:Blood Platelets / Q000378:metabolism / Q000473:pathology; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D004195:Disease Models, Animal; D020690:GTPase-Activating Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D051379:Mice; D008817:Mice, Mutant Strains; D020125:Mutation, Missense; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics / Q000378:metabolism*; D018129:Phosphatidylinositol Phosphates / Q000235:genetics / Q000378:metabolism*; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000235:genetics; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology*; D013921:Thrombocytopenia / Q000235:genetics / Q000378:metabolism / Q000473:pathology
chemical_list: D020690:GTPase-Activating Proteins; D018129:Phosphatidylinositol Phosphates; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; C484658:RAP1GAP protein, human; C583390:RASA3 protein, mouse; C539504:Rap1GAP protein, mouse; D018160:Receptors, Cytoplasmic and Nuclear; C061107:inositol-1,3,4,5-tetrakisphosphate receptor; C060974:phosphatidylinositol 3,4,5-triphosphate; D019869:Phosphatidylinositol 3-Kinases
keywords: GAP1IP4BP; Ras-related protein 1 (Rap1); Rasa3; cell signaling; integrin; phosphatidylinositide 3-kinase (PI3K); platelet