ABSTRACT
Lymphatic vessels develop from specialized endothelial cells in preexisting blood vessels, but the molecular signals that regulate this separation are unknown. Here we identify a failure to separate emerging lymphatic vessels from blood vessels in mice lacking the hematopoietic signaling protein SLP-76 or Syk. Blood-lymphatic connections lead to embryonic hemorrhage and arteriovenous shunting. Expression of slp-76 could not be detected in endothelial cells, and blood-filled lymphatics also arose in wild-type mice reconstituted with SLP-76-deficient bone marrow. These studies reveal a hematopoietic signaling pathway required for separation of the two major vascular networks in mammals.
Subject(s)
Blood Vessels/embryology , Enzyme Precursors/metabolism , Fluorescein-5-isothiocyanate/analogs & derivatives , Lymphatic System/embryology , Phosphoproteins/metabolism , Protein-Tyrosine Kinases/metabolism , Signal Transduction , Adaptor Proteins, Signal Transducing , Animals , Animals, Newborn , Antigens, CD34/metabolism , Blood Vessels/abnormalities , Blood Vessels/cytology , Blood Vessels/metabolism , Bone Marrow Cells/cytology , Bone Marrow Cells/physiology , Bone Marrow Transplantation , Dextrans , Endothelium, Lymphatic/cytology , Endothelium, Lymphatic/metabolism , Endothelium, Vascular/cytology , Endothelium, Vascular/metabolism , Enzyme Precursors/genetics , Glycoproteins/metabolism , Hemorrhage , Homeodomain Proteins/genetics , Homeodomain Proteins/metabolism , Intestines/blood supply , Intracellular Signaling Peptides and Proteins , Lymphatic System/abnormalities , Lymphatic System/cytology , Lymphatic System/metabolism , Membrane Transport Proteins , Mesenteric Arteries/abnormalities , Mesenteric Veins/abnormalities , Mice , Phosphoproteins/genetics , Protein-Tyrosine Kinases/genetics , Stem Cells/physiology , Syk Kinase , Tumor Suppressor Proteins , Vascular Endothelial Growth Factor Receptor-3/metabolism , Veins/cytology , Veins/embryology , Veins/metabolismABSTRACT
Mice deficient in the adaptor Src homology 2 domain-containing leukocyte phosphoprotein of 76 kD (SLP-76) exhibit a bleeding disorder and lack T cells. Linker for activation of T cells (LAT)-deficient mice exhibit a similar T cell phenotype, but show no signs of hemorrhage. Both SLP-76 and LAT are important for optimal platelet activation downstream of the collagen receptor, GPVI. In addition, SLP-76 is involved in signaling mediated by integrin alphaIIbbeta3. Because SLP-76 and LAT function coordinately in T cell signal transduction, yet their roles appear to differ in hemostasis, we investigated in detail the functional consequences of SLP-76 and LAT deficiencies in platelets. Previously we have shown that LAT(-/-) platelets exhibit defective responses to the GPVI-specific agonist, collagen-related peptide (CRP). Consistent with this, we find that surface expression of P-selectin in response to high concentrations of GPVI ligands is reduced in both LAT- and SLP-76-deficient platelets. However, platelets from LAT(-/-) mice, but not SLP-76(-/-) mice, aggregate normally in response to high concentrations of collagen and convulxin. Additionally, unlike SLP-76, LAT is not tyrosine phosphorylated after fibrinogen binding to integrin alphaIIbbeta3, and collagen-stimulated platelets deficient in LAT spread normally on fibrinogen-coated surfaces. Together, these findings indicate that while LAT and SLP-76 are equally required for signaling via the T cell antigen receptor (TCR) and pre-TCR, platelet activation downstream of GPVI and alphaIIbbeta3 shows a much greater dependency on SLP-76 than LAT.
