Dimerization of P-selectin in platelets and endothelial cells.
P-selectin is a leukocyte adhesion receptor stored in platelets and endothelial cells and is translocated to the surface upon cell activation. Purified P-selectin is oligomeric and has increased avidity for its ligand relative to the monomeric form, but whether P-selectin self-associates in the membrane of intact cells is not known. A chemical cross-linking approach was used to show that P-selectin is present as noncovalent dimers in resting platelets, human umbilical vein endothelial cells, and heterologous RIN5F cells expressing P-selectin. The results of 2-dimensional isoelectric focusing are consistent in showing P-selectin dimers as homodimers, but they are composed of a more basic subset of P-selectin than the monomers. This suggests that the dimers are a biochemically distinct subset of P-selectin. P-selectin dimers form in the endoplasmic reticulum and Golgi compartments of human umbilical vein endothelial cells only after synthesis of the mature P-selectin subunit, and are not preferentially stored in Weibel-Palade bodies as compared with the monomeric form. Platelet activation with thrombin receptor-activating peptide leads to the presence of P-selectin monomers and homodimers on the cell surface as well as P-selectin heterodimers, which are composed of P-selectin and an unidentified protein of approximately 81 kd molecular weight. In summary, these studies demonstrate that P-selectin is homodimeric in situ and that platelet activation leads to the formation of an additional activation-specific heterodimeric species. In addition, the homodimer has unique biochemical characteristics compared with the monomeric form, and dimerization occurs in the endoplasmic reticulum and Golgi compartments of endothelial cells.
Barkalow
F J
FJ
Department of Pathology and the Department of Medicine, Brigham and Women's Hospital and Harvard Medical School, Boston, MA 02115, USA.
Barkalow
K L
KL
Mayadas
T N
TN
D019943:Amino Acid Substitution; D001792:Blood Platelets / Q000502:physiology*; D002460:Cell Line; D002478:Cells, Cultured; D003432:Cross-Linking Reagents; D019281:Dimerization; D004721:Endoplasmic Reticulum / Q000502:physiology; D004730:Endothelium, Vascular / Q000502:physiology*; D006056:Golgi Apparatus / Q000502:physiology; D006801:Humans; D066298:In Vitro Techniques; D004915:Leukemia, Erythroblastic, Acute; D015388:Organelles / Q000502:physiology; D019007:P-Selectin / Q000097:blood / Q000737:chemistry* / Q000378:metabolism*; D017354:Point Mutation; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D014407:Tumor Cells, Cultured; D014471:Umbilical Veins
D003432:Cross-Linking Reagents; D019007:P-Selectin; D011994:Recombinant Proteins