title: Lnk regulates integrin alphaIIbbeta3 outside-in signaling in mouse platelets, leading to stabilization of thrombus development in vivo.
abstract: The nature of the in vivo cellular events underlying thrombus formation mediated by platelet activation remains unclear because of the absence of a modality for analysis. Lymphocyte adaptor protein (Lnk; also known as Sh2b3) is an adaptor protein that inhibits thrombopoietin-mediated signaling, and as a result, megakaryocyte and platelet counts are elevated in Lnk-/- mice. Here we describe an unanticipated role for Lnk in stabilizing thrombus formation and clarify the activities of Lnk in platelets transduced through integrin alphaIIbbeta3-mediated outside-in signaling. We equalized platelet counts in wild-type and Lnk-/- mice by using genetic depletion of Lnk and BM transplantation. Using FeCl3- or laser-induced injury and in vivo imaging that enabled observation of single platelet behavior and the multiple steps in thrombus formation, we determined that Lnk is an essential contributor to the stabilization of developing thrombi within vessels. Lnk-/- platelets exhibited a reduced ability to fully spread on fibrinogen and mediate clot retraction, reduced tyrosine phosphorylation of the beta3 integrin subunit, and reduced binding of Fyn to integrin alphaIIbbeta3. These results provide new insight into the mechanism of alphaIIbbeta3-based outside-in signaling, which appears to be coordinated in platelets by Lnk, Fyn, and integrins. Outside-in signaling modulators could represent new therapeutic targets for the prevention of cardiovascular events.
authors.affiliation: Research Institute, International Medical Center of Japan, 1-21-1 Toyama, Shinjuku-ku, Tokyo, Japan.
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Takizawa
authors.forename: Satoshi
authors.initials: S
authors.lastname: Nishimura
authors.forename: Naoya
authors.initials: N
authors.lastname: Takayama
authors.forename: Atsushi
authors.initials: A
authors.lastname: Oda
authors.forename: Hidekazu
authors.initials: H
authors.lastname: Nishikii
authors.forename: Yohei
authors.initials: Y
authors.lastname: Morita
authors.forename: Sei
authors.initials: S
authors.lastname: Kakinuma
authors.forename: Satoshi
authors.initials: S
authors.lastname: Yamazaki
authors.forename: Satoshi
authors.initials: S
authors.lastname: Okamura
authors.forename: Noriko
authors.initials: N
authors.lastname: Tamura
authors.forename: Shinya
authors.initials: S
authors.lastname: Goto
authors.forename: Akira
authors.initials: A
authors.lastname: Sawaguchi
authors.forename: Ichiro
authors.initials: I
authors.lastname: Manabe
authors.forename: Kiyoshi
authors.initials: K
authors.lastname: Takatsu
authors.forename: Hiromitsu
authors.initials: H
authors.lastname: Nakauchi
authors.forename: Satoshi
authors.initials: S
authors.lastname: Takaki
authors.forename: Koji
authors.initials: K
authors.lastname: Eto
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D001792:Blood Platelets / Q000378:metabolism*; D016466:CHO Cells; D019556:COS Cells; D000081247:CSK Tyrosine-Protein Kinase; D002522:Chlorocebus aethiops; D006224:Cricetinae; D003412:Cricetulus; D047908:Intracellular Signaling Peptides and Proteins; D008297:Male; D008565:Membrane Proteins; D051379:Mice; D008810:Mice, Inbred C57BL; D010766:Phosphorylation; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000502:physiology*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011506:Proteins / Q000502:physiology*; D051076:Proto-Oncogene Proteins c-fyn / Q000502:physiology; D015398:Signal Transduction / Q000502:physiology*; D013927:Thrombosis / Q000209:etiology*; D019061:src-Family Kinases
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D047908:Intracellular Signaling Peptides and Proteins; C490777:Lnk protein, mouse; D008565:Membrane Proteins; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D011506:Proteins; D011505:Protein-Tyrosine Kinases; D000081247:CSK Tyrosine-Protein Kinase; C494960:Fyn protein, mouse; D051076:Proto-Oncogene Proteins c-fyn; D019061:src-Family Kinases