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 tra nsported 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 api cal determinant by its interaction with sphingolipid-cholesterol rafts. We modified the rat growth hormone (rGH), an unglycosylated, unpolarized secre ted 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 t o an increase in apical delivery of the protein. However, addition of N-gly cans to GPI-anchored rGH resulted in predominant apical delivery, suggestin g 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-anc hored rGH, a transmembrane form of rGH which was not raft-associated accumu lated intracellularly. Addition of N-glycans to this chimeric protein preve nted intracellular accumulation and led to apical delivery.