title: RGT, a synthetic peptide corresponding to the integrin beta 3 cytoplasmic C-terminal sequence, selectively inhibits outside-in signaling in human platelets by disrupting the interaction of integrin alpha IIb beta 3 with Src kinase.
abstract: Mutational analysis has established that the cytoplasmic tail of the integrin beta 3 subunit binds c-Src (termed as Src in this study) and is critical for bidirectional integrin signaling. Here we show in washed human platelets that a cell-permeable, myristoylated RGT peptide (myr-RGT) corresponding to the integrin beta 3 C-terminal sequence dose-dependently inhibited stable platelet adhesion and spreading on immobilized fibrinogen, and fibrin clot retraction as well. Myr-RGT also inhibited the aggregation-dependent platelet secretion and secretion-dependent second wave of platelet aggregation induced by adenosine diphosphate, ristocetin, or thrombin. Thus, myr-RGT inhibited integrin outside-in signaling. In contrast, myr-RGT had no inhibitory effect on adenosine diphosphate-induced soluble fibrinogen binding to platelets that is dependent on integrin inside-out signaling. Furthermore, the RGT peptide induced dissociation of Src from integrin beta 3 and dose-dependently inhibited the purified recombinant beta 3 cytoplasmic domain binding to Src-SH3. In addition, phosphorylation of the beta 3 cytoplasmic tyrosines, Y(747) and Y(759), was inhibited by myr-RGT. These data indicate an important role for beta 3-Src interaction in outside-in signaling. Thus, in intact human platelets, disruption of the association of Src with beta 3 and selective blockade of integrin alpha IIb beta 3 outside-in signaling by myr-RGT suggest a potential new antithrombotic strategy.
authors.affiliation: State Key Laboratory of Medical Genomics, Shanghai Institute of Hematology, Ruijin Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China.
authors.forename: Xiaoyu
authors.initials: X
authors.lastname: Su
authors.forename: Jianqing
authors.initials: J
authors.lastname: Mi
authors.forename: Jinsong
authors.initials: J
authors.lastname: Yan
authors.forename: Panagiotis
authors.initials: P
authors.lastname: Flevaris
authors.forename: Yuanjing
authors.initials: Y
authors.lastname: Lu
authors.forename: Hongchen
authors.initials: H
authors.lastname: Liu
authors.forename: Zheng
authors.initials: Z
authors.lastname: Ruan
authors.forename: Xuefeng
authors.initials: X
authors.lastname: Wang
authors.forename: Nelly
authors.initials: N
authors.lastname: Kieffer
authors.forename: Saijuan
authors.initials: S
authors.lastname: Chen
authors.forename: Xiaoping
authors.initials: X
authors.lastname: Du
authors.forename: Xiaodong
authors.initials: X
authors.lastname: Xi
mesh_terms: D003018:Clot Retraction / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D006801:Humans; D039661:Integrin beta3; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000494:pharmacology*; D010766:Phosphorylation / Q000187:drug effects; D010973:Platelet Adhesiveness / Q000187:drug effects; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000378:metabolism*; D011485:Protein Binding / Q000187:drug effects; D015398:Signal Transduction / Q000187:drug effects*; D019061:src-Family Kinases / Q000378:metabolism*
chemical_list: D039661:Integrin beta3; D010446:Peptide Fragments; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D019061:src-Family Kinases