title: Gas6-induced tissue factor expression in endothelial cells is mediated through caveolin-1-enriched microdomains.
abstract: BACKGROUND
  Gas6 has been shown to interact with Axl in endothelial cells and to induce several signaling pathways involved in cell survival and proliferation. However, the interaction of Gas6/Axl with lipid raft/caveolin-1 in endothelial cells and its role in thrombosis are unknown.
  
  
  OBJECTIVE
  We tested whether Axl and/or caveolin-1 is involved in Gas6-induced Akt, ERK1/2, and c-Src activation leading to altered tissue factor expression in endothelial cells.
  
  
  METHODS
  Gas6-treated endothelial cells were transfected with small interfering RNA (siRNA) for Axl, caveolin-1, c-Src, and Akt or treated with pharmacological inhibitors of c-Src and ERK1/2. Sucrose gradient centrifugation and confocal microscopy were used to study lipid raft/caveolin-1-enriched fractions. Akt, ERK1/2, p38, and c-Src activation was analyzed by Western blot analysis. Tissue factor expression was assessed by real-time quantitative polymerase chain reaction and immunofluorescence.
  
  
  CONCLUSIONS
  Gas6 induced Axl and c-Src localization into lipid raft/caveolin-1-enriched fractions. Gas6 increased the phosphorylation of Akt, ERK1/2, and c-Src but not p38. Using siRNA, we demonstrated that Axl is required for Akt, ERK1/2, and c-Src activation after Gas6 stimulation. siRNA for caveolin-1 blocked Gas6-induced phosphorylation of Akt, ERK1/2, and c-Src. c-Src downregulation inhibited Gas6-induced Akt but not ERK1/2 phosphorylation. Finally, Gas6 increased tissue factor mRNA and protein expression in endothelial cells. Tissue factor expression was blocked by siRNA for Axl, caveolin-1, or Akt as well as c-Src inhibition. These data demonstrate that the signaling pathway Gas6/Axl/caveolin-1/c-Src/Akt is required for tissue factor expression in endothelial cells, providing mechanistic insight into how Gas6 exerts its prothrombotic role in the vasculature.
authors.affiliation: Lady Davis Institute for Medical Research, McGill University, Montreal, QC, Canada.
authors.forename: S
authors.initials: S
authors.lastname: Laurance
authors.forename: M N
authors.initials: MN
authors.lastname: Aghourian
authors.forename: Z
authors.initials: Z
authors.lastname: Jiva Lila
authors.forename: C A
authors.initials: CA
authors.lastname: Lemarié
authors.forename: M D
authors.initials: MD
authors.lastname: Blostein
mesh_terms: D000081247:CSK Tyrosine-Protein Kinase; D051242:Caveolin 1 / Q000378:metabolism*; D049109:Cell Proliferation; D002470:Cell Survival; D042783:Endothelial Cells / Q000166:cytology*; D061307:Human Umbilical Vein Endothelial Cells; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000378:metabolism*; D021962:Membrane Microdomains / Q000737:chemistry; D010766:Phosphorylation; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D020794:Receptor Protein-Tyrosine Kinases / Q000378:metabolism; D015398:Signal Transduction; D013925:Thromboplastin / Q000378:metabolism*; D013927:Thrombosis / Q000378:metabolism; D019061:src-Family Kinases / Q000378:metabolism; D000094963:Axl Receptor Tyrosine Kinase; D000099255:Growth Arrest-Specific Protein 6
chemical_list: D000081247:CSK Tyrosine-Protein Kinase; D051242:Caveolin 1; D036341:Intercellular Signaling Peptides and Proteins; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D020794:Receptor Protein-Tyrosine Kinases; D013925:Thromboplastin; D019061:src-Family Kinases; D000094963:Axl Receptor Tyrosine Kinase; D000099255:Growth Arrest-Specific Protein 6; C495540:CAV1 protein, human; C000631346:CSK protein, human; C000722289:AXL protein, human
keywords: axl receptor tyrosine kinase; c-src kinase; caveolin-1; growth arrest-specific protein 6; tissue factor