Oligomerized Tie2 localizes to clathrin-coated pits in response to angiopoietin-1.
The tyrosine kinase receptor Tie2 is expressed on endothelial cells, and together with its ligand angiopoietin-1 (Ang1), is important for angiogenesis and vascular stability. Upon activation by Ang1, Tie2 is rapidly internalized and degraded, a mechanism most likely necessary to attenuate receptor activity. Using immunogold electron microscopy, we show that on the surface of endothelial cells, Tie2 is arranged in variably sized clusters containing dimers and higher order oligomers. Clusters of Tie2 were expressed on the apical and basolateral plasma membranes, and on the tips of microvilli. Upon activation by Ang1, Tie2 co-localized with the clathrin heavy chain at the apical and basolateral plasma membranes and within endothelial cells indicating that Tie2 internalizes through clathrin-coated pits. Inhibiting cellular endocytosis by depleting cellular potassium or by acidifying the cytosol blocked the internalization of Tie2 in response to Ang1. Our results suggest that one pathway mediating the internalization of Tie2 in response to Ang1 is through clathrin-coated pits.
Molecular and Cellular Biology Research, Sunnybrook Research Institute, 2075 Bayview Avenue, Research Building, S-218, Toronto, ON, M4N 3M5, Canada.
Elena
E
Bogdanovic
Neil
N
Coombs
Daniel J
DJ
Dumont
D042683:Angiopoietin-1 / Q000819:agonists / Q000378:metabolism*; D002478:Cells, Cultured; D003034:Coated Pits, Cell-Membrane / Q000201:enzymology* / Q000648:ultrastructure; D004730:Endothelium, Vascular / Q000201:enzymology* / Q000648:ultrastructure; D006801:Humans; D042787:Receptor, TIE-2 / Q000378:metabolism*
D042683:Angiopoietin-1; D042787:Receptor, TIE-2