Polarity of endogenous and exogenous glycosyl-phosphatidylinositol-anchored membrane proteins in Madin-Darby canine kidney cells

Jean Wilson, Nicolas Fasel, Jean Pierre Kraehenbuhl

Research output: Contribution to journalArticle

23 Citations (Scopus)

Abstract

Madin-Darby canine kidney cells (MDCK) were transfected with a cDNA encoding the glycosyl-phosphatidylinositol (GPI)-anchored protein mouse Thy-1 in order to study the steady-state surface distribution of exogenous and endogenous GPI-linked proteins. Immunofluorescence of transfected cells grown on collagen-coated coverslips showed that expression of Thy-1 was variable throughout the epithelium, with some cells expressing large amounts of Thy-1 adjacent to very faintly staining cells. Selective surface iodination of cells grown on collagen-coated or uncoated transwell filters followed by immunoprecipitation of Thy-1 demonstrated that all the Thy-1 was present exclusively in the apical plasma membrane. Although cells grown on uncoated filters had much smaller amounts of Thy-1, it was consistently localized on the apical surfaces. Immunofluorescent localization of Thy-1 on 1 μm frozen sections of filter-grown cells demonstrated that all the Thy-1 was on the apical surface and there was no detectable intracellular pool.Phosphatidylinositol-specific phospholipase C digestion of intact iodinated monolayers released Thy-1 only into the apical medium, indicating that Thy-1 was processed normally in transfected cells and was anchored by a GPI-tail) In agreement with previous findings, endogenous GPI-linked proteins were found only on the apical plasma membrane. These results suggest that there is a common mechanism for sorting and targeting of GPI-linked proteins in polarized epithelial cells.

Original languageEnglish (US)
Pages (from-to)143-149
Number of pages7
JournalJournal of Cell Science
Volume96
Issue number1
StatePublished - May 1990

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Glycosylphosphatidylinositols
Madin Darby Canine Kidney Cells
Membrane Proteins
Proteins
Collagen
Cell Membrane
Phosphoinositide Phospholipase C
Halogenation
Frozen Sections
Immunoprecipitation
Fluorescent Antibody Technique
Tail
Digestion
Epithelium
Complementary DNA
Epithelial Cells
Staining and Labeling

Keywords

  • Epithelial cell polarity
  • Glycolipid anchor
  • Thy-1

ASJC Scopus subject areas

  • Cell Biology

Cite this

Polarity of endogenous and exogenous glycosyl-phosphatidylinositol-anchored membrane proteins in Madin-Darby canine kidney cells. / Wilson, Jean; Fasel, Nicolas; Kraehenbuhl, Jean Pierre.

In: Journal of Cell Science, Vol. 96, No. 1, 05.1990, p. 143-149.

Research output: Contribution to journalArticle

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