Vectorial apical delivery and slow endocytosis of a glycolipid-anchored fusion protein in transfected MDCK cells.
Academic Article
Overview
abstract
To characterize the mechanisms that determine the apical polarity of proteins anchored by glycosylphosphatidylinositol (GPI), we studied the targeting of a GPI-anchored form of a herpes simplex glycoprotein, gD-1, in transfected MDCK cells. Using a biotin-based targeting assay, we found that GPI-anchored gD-1 was sorted intracellularly and delivered directly to the apical surface. Endocytosis of GPI-anchored gD-1 occurred slowly and preferentially from the apical domain, while transcytosis of the basolateral fraction did not occur at a significant rate (incompatible with being a precursor to the apical pool). Prevention of tight junction formation by incubation in medium with micromolar Ca2+ resulted in expression of GPI-anchored gD-1 on the free surface, but not on the attached surface of the cell. Our results indicate that the apical polarity of a GPI-anchored protein is generated by vectorial delivery to the apical membrane, where its distribution is maintained by slow endocytosis and by a retention system not necessarily involving the tight junction.
