N-Glycans and Glycosylphosphatidylinositol-Anchor Act on Polarized Sorting of Mouse PrPC in Madin-Darby Canine Kidney Cells

The cellular prion protein (PrPC) plays a fundamental role in prion disease. PrPC is a glycosylphosphatidylinositol (GPI)-anchored protein with two variably occupied N-glycosylation sites. In general, GPI-anchor and N-glycosylation direct proteins to apical membranes in polarized cells whereas the m...

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Bibliographic Details
Published in:PloS one 2011-09, Vol.6 (9), p.e24624
Main Authors: Puig, Berta, Altmeppen, Hermann C., Thurm, Dana, Geissen, Markus, Conrad, Catharina, Braulke, Thomas, Glatzel, Markus
Format: Article
Language:English
Subjects:
Amino acids
Biochemistry
Biology
Biotinylation
Cell surface
Chromatography
Confocal microscopy
Disease
Glycoproteins
Glycosylation
Glycosylphosphatidylinositol
Laboratories
Mass spectrometry
Medicine
Membranes
Microscopy
Mutation
N-glycans
Neurodegeneration
Neuropathology
Polysaccharides
Prion protein
Proteins
Scientific imaging
Veterinary Science
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Summary:The cellular prion protein (PrPC) plays a fundamental role in prion disease. PrPC is a glycosylphosphatidylinositol (GPI)-anchored protein with two variably occupied N-glycosylation sites. In general, GPI-anchor and N-glycosylation direct proteins to apical membranes in polarized cells whereas the majority of mouse PrPC is found in basolateral membranes in polarized Madin-Darby canine kidney (MDCK) cells. In this study we have mutated the first, the second, and both N-glycosylation sites of PrPC and also replaced the GPI-anchor of PrPC by the Thy-1 GPI-anchor in order to investigate the role of these signals in sorting of PrPC in MDCK cells. Cell surface biotinylation experiments and confocal microscopy showed that lack of one N-linked oligosaccharide leads to loss of polarized sorting of PrPC. Exchange of the PrPC GPI-anchor for the one of Thy-1 redirects PrPC to the apical membrane. In conclusion, both N-glycosylation and GPI-anchor act on polarized sorting of PrPC, with the GPI-anchor being dominant over N-glycans.
ISSN:1932-6203
1932-6203
DOI:10.1371/journal.pone.0024624