The cytoplasmic domain of P-selectin contains a sorting determinant that mediates rapid degradation in lysosomes.
P-selectin is a cell adhesion molecule required transiently on the surface of activated platelets and endothelial cells as a receptor for leukocytes. It is stored in secretory granules in platelets, endothelial cells, and transfected neuroendocrine cells and is rapidly delivered to the plasma membrane upon exocytosis of the secretory granules. It is then rapidly internalized in endothelial cells and transfected cells. We find that in transfected neuroendocrine PC12 cells, the fraction of P-selectin that is not targeted to secretory granules is rapidly degraded. In transfected CHO fibroblasts, which lack secretory granules, P-selectin was degraded with a half time of 2.3 h in plated cells, while low density lipoprotein receptor (LDL-R) had a half life of 9 h. In cells cultured in ammonium chloride to inhibit lysosomal proteinases, P-selectin was protected from degradation and rapidly accumulated in vesicles enriched in lgp-B, a resident lysosomal membrane protein. The cytoplasmic domain of P-selectin was sufficient to confer rapid turnover on LDL-R. Deletion of 10 amino acids from the cytoplasmic domain of P-selectin extended the half life to 9.5 h and abrogated rapid lysosomal targeting in the presence of ammonium chloride, implicating this sequence as a necessary element of a novel lysosomal targeting signal. The rate limiting step in degradation occurred after internalization from the cell surface, indicating that sorting of P-selectin away from efficiently recycled proteins occurs in endosomes. We propose that this sorting event represents a constitutive equivalent of receptor down regulation, and may function to regulate the expression of P-selectin at the surface of activated endothelial cells.
Green
S A
SA
Department of Biochemistry and Biophysics, University of California, San Francisco 94143-0534.
Setiadi
H
H
McEver
R P
RP
Kelly
R B
RB
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D016466:CHO Cells; D015815:Cell Adhesion Molecules / Q000737:chemistry / Q000378:metabolism*; D006224:Cricetinae; D003593:Cytoplasm / Q000378:metabolism; D003594:Cytoplasmic Granules / Q000378:metabolism; D004247:DNA; D006207:Half-Life; D008247:Lysosomes / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D019007:P-Selectin; D016716:PC12 Cells; D010980:Platelet Membrane Glycoproteins / Q000737:chemistry / Q000378:metabolism*; D014162:Transfection
D015815:Cell Adhesion Molecules; D019007:P-Selectin; D010980:Platelet Membrane Glycoproteins; D004247:DNA