Preferred apical distribution of glycosyl-phosphatidylinositol (GPI) anchored proteins: a highly conserved feature of the polarized epithelial cell phenotype.
We use a sensitive biotin polarity assay to survey the surface distribution of glycosyl-phosphatidylinositol (GPI) anchored proteins in five model epithelial cell lines derived from different species (dog, pig, man) and tissues, i.e., kidney (MDCK I, MDCK II, LLC-PK1) and intestine (Caco-2 and SK-CO15). After biotinylation of apical or basolateral surfaces of confluent monolayers grown on polycarbonate filters, GPI-anchored proteins are identified by their shift from a Triton X-114 detergent-rich phase to a detergent-poor phase in the presence of phosphatidylinositol-specific phospholipase C. All GPI-anchored proteins detected (3-9 per cell type, at least 13 different proteins) are found to be apically polarized; no GPI-anchored protein is observed preferentially localized to the basal surface. One of the GPI-anchored proteins is identified as carcinoembryonic antigen (CEA). Survey of MDCK II-RCAr, a mutant cell line with a pleiotropic defect in galactosylation of glycoproteins and glycolipids (that presumably affects GPI anchors) also reveals an apical polarization of all GPI-anchored proteins. In contrast, analysis of MDCK II-ConAr (a mutant cell line with an unknown defect in glycosylation) revealed five GPI-anchored proteins, two of which appeared relatively unpolarized. Our results indicate that the polarized apical distribution of GPI-anchored proteins is highly conserved across species and tissue-type and may depend on glycosylation.
Lisanti
M P
MP
Department of Cell Biology and Anatomy, Cornell University Medical College, New York, New York 10021.
Le Bivic
A
A
Saltiel
A R
AR
Rodriguez-Boulan
E
E
D000818:Animals; D002460:Cell Line; D004285:Dogs; D004848:Epithelium / Q000032:analysis* / Q000648:ultrastructure; D006017:Glycolipids / Q000032:analysis*; D017261:Glycosylphosphatidylinositols; D006801:Humans; D015151:Immunoblotting; D007413:Intestinal Mucosa / Q000032:analysis; D007668:Kidney / Q000032:analysis; D008565:Membrane Proteins / Q000032:analysis*; D010641:Phenotype; D010716:Phosphatidylinositols / Q000032:analysis*
D006017:Glycolipids; D017261:Glycosylphosphatidylinositols; D008565:Membrane Proteins; D010716:Phosphatidylinositols