title: Growth and maturation of megakaryocytes is regulated by Lnk/Sh2b3 adaptor protein through crosstalk between cytokine- and integrin-mediated signals.
abstract: OBJECTIVE
  Various cytokines and growth factors control the differentiation and maturation of megakaryocytes (MKs). However, the mechanism regulating platelet release from MKs is not well understood. Here, we investigated a role of Lnk/Sh2b3, an intracellular adaptor protein, in megakaryopoiesis.
  
  
  METHODS
  Number of MK progenitor in bone marrow (BM) of wild-type or Lnk(-/-) mice and their sensitivity to thrombopoietin (TPO) were determined in colony-forming unit assay. Using BM-derived wild-type or Lnk(-/-) MKs stimulated with TPO, activation of the signaling molecules was biochemically analyzed and effect of integrin stimulation on TPO signals was studied by addition of vascular cell adhesion molecule (VCAM-1). Platelet production from MKs in the presence of VCAM-1 was counted by flow cytometry and their morphological change was observed by time-lapse microscopy.
  
  
  RESULTS
  Lnk(-/-) mice showed elevated platelets and mature MKs due to enhanced sensitivity of progenitors to TPO. Erk1/2 phosphorylation induced by TPO was augmented and prolonged in Lnk(-/-) MKs while activation of signal transducers and activators of transcription (Stat)3, Stat5, and Akt was normal. Wild-type MKs, but not in Lnk(-/-) MKs on VCAM-1 showed reduced Stat5 phosphorylation and mitogen-activated protein kinases activation upon stimulation with TPO. Additionally, the presence of VCAM in culture accelerated spontaneous platelet release from mature wild-type MKs, but not from Lnk(-/-) MKs.
  
  
  CONCLUSIONS
  Results suggest that contact of MKs with adhesion molecules via integrins might contribute to platelet release, which is under Lnk-mediated regulation of Stat-5 activation and show that Lnk functions in responses controlled by cell adhesion and in crosstalk between integrin- and cytokine-mediated signaling.
authors.affiliation: Division of Immunology, Department of Microbiology and Immunology, The University of Tokyo, Tokyo, Japan.
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Takizawa
authors.forename: Koji
authors.initials: K
authors.lastname: Eto
authors.forename: Atsuko
authors.initials: A
authors.lastname: Yoshikawa
authors.forename: Hiromitsu
authors.initials: H
authors.lastname: Nakauchi
authors.forename: Kiyoshi
authors.initials: K
authors.lastname: Takatsu
authors.forename: Satoshi
authors.initials: S
authors.lastname: Takaki
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D001792:Blood Platelets / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects / Q000502:physiology; D006801:Humans; D016023:Integrins / Q000378:metabolism*; D047908:Intracellular Signaling Peptides and Proteins; D020935:MAP Kinase Signaling System / Q000187:drug effects / Q000502:physiology*; D008533:Megakaryocytes / Q000166:cytology / Q000378:metabolism*; D008565:Membrane Proteins; D051379:Mice; D018345:Mice, Knockout; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D050796:STAT3 Transcription Factor / Q000378:metabolism; D050799:STAT5 Transcription Factor / Q000378:metabolism; D013926:Thrombopoietin / Q000378:metabolism / Q000494:pharmacology*; D019010:Vascular Cell Adhesion Molecule-1 / Q000378:metabolism*
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D016023:Integrins; D047908:Intracellular Signaling Peptides and Proteins; C490777:Lnk protein, mouse; D008565:Membrane Proteins; D011506:Proteins; D050796:STAT3 Transcription Factor; D050799:STAT5 Transcription Factor; C494087:Stat3 protein, mouse; D019010:Vascular Cell Adhesion Molecule-1; D013926:Thrombopoietin; D051057:Proto-Oncogene Proteins c-akt; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3