Role of the glycosyl-phosphatidylinositol anchor in the intracellular transport of a transmembrane protein in Madin-Darby canine kidney cells

In order to compare the trafficking of proteins with different membrane anc hors, we have constructed and expressed three different recombinant forms o f neutral endopeptidase (NEP) in MDCK cells. The wild type form of NEP (WT- NEP) is attached to the plasma membrane by a single N-terminal membrane spa nning domain, whereas the glycosylphosphatidylinositol-anchored form of the protein (GPI-NEP) contains a C-terminal GPI anchor. A double anchored form of NEP (DA-NEP) was also constructed, that contains both the original N-te rminal membrane spanning domain and a C-terminal GPI anchor. We show here t hat WT-NEP, GPI-NEP and DA-NEP, which are all apically targeted in MDCK cel ls, behave differently when subjected to Triton X-100 solubilisation: despi te the presence of the transmembrane anchor DA-NEP behaves as a GPI-anchore d protein, This suggests that the GPI anchor of DA-NEP is dominant over the transmembrane anchor of the native protein to determine its pattern of sol ubility in Triton X-100. (C) 1998 Elsevier Science B.V. All rights reserved .