N-Glycans mediate the apical sorting of a GPI-anchored, raft-associated protein in Madin-Darby canine kidney cells.
Glycosyl-phosphatidylinositol (GPI)- anchored proteins are preferentially transported to the apical cell surface of polarized Madin-Darby canine kidney (MDCK) cells. It has been assumed that the GPI anchor itself acts as an apical determinant by its interaction with sphingolipid-cholesterol rafts. We modified the rat growth hormone (rGH), an unglycosylated, unpolarized secreted protein, into a GPI-anchored protein and analyzed its surface delivery in polarized MDCK cells. The addition of a GPI anchor to rGH did not lead to an increase in apical delivery of the protein. However, addition of N-glycans to GPI-anchored rGH resulted in predominant apical delivery, suggesting that N-glycans act as apical sorting signals on GPI-anchored proteins as they do on transmembrane and secretory proteins. In contrast to the GPI-anchored rGH, a transmembrane form of rGH which was not raft-associated accumulated intracellularly. Addition of N-glycans to this chimeric protein prevented intracellular accumulation and led to apical delivery.
Benting
J H
JH
European Molecular Biology Laboratory (EMBL), Cell Biology and Biophysics Programme, D-69012 Heidelberg, Germany.
Rietveld
A G
AG
Simons
K
K
D000818:Animals; D001692:Biological Transport; D018958:CD55 Antigens / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D016764:Cell Polarity; D002784:Cholesterol / Q000378:metabolism*; D004285:Dogs; D006031:Glycosylation; D017261:Glycosylphosphatidylinositols / Q000378:metabolism*; D013006:Growth Hormone / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007668:Kidney / Q000166:cytology / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011134:Polysaccharides / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D015202:Protein Engineering; D051381:Rats; D011973:Receptors, LDL / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism; D012995:Solubility; D013107:Sphingolipids / Q000378:metabolism*; D014162:Transfection
D018958:CD55 Antigens; D017261:Glycosylphosphatidylinositols; D008565:Membrane Proteins; D011134:Polysaccharides; D011973:Receptors, LDL; D011993:Recombinant Fusion Proteins; D013107:Sphingolipids; D013006:Growth Hormone; D002784:Cholesterol