title: Signal-dependent distribution of cell surface P-selectin in clathrin-coated pits affects leukocyte rolling under flow.
abstract: Flowing leukocytes roll on P-selectin that is mobilized from secretory granules to the surfaces of endothelial cells after stimulation with histamine or thrombin. Before it is internalized, P-selectin clusters in clathrin-coated pits, which enhances its ability to support leukocyte rolling. We found that thrombin and histamine induced comparable exocytosis of P-selectin on endothelial cells. However, compared with histamine, thrombin decreased the recruitment of P-selectin into clathrin-coated pits, slowed the internalization of P-selectin, and reduced the number and stability of neutrophils rolling on P-selectin. Significantly more RhoA was activated in thrombin- than in histamine-stimulated endothelial cells. Inhibitors of RhoA or its effector, Rho kinase, reversed thrombin's ability to inhibit the internalization and adhesive function of P-selectin in endothelial cells. Experiments with transfected cells confirmed that the inhibitory actions of thrombin and Rho kinase on P-selectin required its cytoplasmic domain. Thus, a signaling event affects both the function and clearance of a protein that enters the constitutive clathrin-mediated endocytic pathway.
authors.affiliation: Cardiovascular Biology Research Program, Oklahoma Medical Research Foundation, Oklahoma City, OK 73104, USA.
authors.forename: Hendra
authors.initials: H
authors.lastname: Setiadi
authors.forename: Rodger P
authors.initials: RP
authors.lastname: McEver
mesh_terms: D000818:Animals; D016466:CHO Cells; D002966:Clathrin / Q000378:metabolism*; D003034:Coated Pits, Cell-Membrane / Q000378:metabolism*; D006224:Cricetinae; D004705:Endocytosis / Q000187:drug effects / Q000502:physiology; D042783:Endothelial Cells / Q000187:drug effects / Q000378:metabolism; D005089:Exocytosis / Q000187:drug effects / Q000502:physiology; D006632:Histamine / Q000494:pharmacology; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D036904:Leukocyte Rolling / Q000502:physiology*; D019007:P-Selectin / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D017434:Protein Structure, Tertiary / Q000187:drug effects / Q000502:physiology; D012039:Regional Blood Flow / Q000502:physiology; D015398:Signal Transduction / Q000502:physiology*; D013917:Thrombin / Q000494:pharmacology; D054460:rho-Associated Kinases; D020742:rhoA GTP-Binding Protein / Q000037:antagonists & inhibitors / Q000378:metabolism
chemical_list: D002966:Clathrin; D047908:Intracellular Signaling Peptides and Proteins; D019007:P-Selectin; D006632:Histamine; D017346:Protein Serine-Threonine Kinases; D054460:rho-Associated Kinases; D013917:Thrombin; D020742:rhoA GTP-Binding Protein