Different GPI-attachment signals affect the oligomerisation of GPI-anchored proteins and their apical sorting.
To understand the mechanism involved in the apical sorting of glycosylphosphatidylinositol (GPI)-anchored proteins (GPI-APs) we fused to the C-terminus of GFP the GPI-anchor-attachment signal of the folate receptor (FR) or of the prion protein (PrP), two native GPI-anchored proteins that are sorted apically or basolaterally, respectively, in MDCK cells. We investigated the behaviour of the resulting fusion proteins GFP-FR and GFP-PrP by analysing three parameters: their association with DRMs, their oligomerisation and their apical sorting. Strikingly, we found that different GPI-attachment signals differently modulate the ability of the resulting GFP-fusion protein to oligomerise and to be apically sorted. This is probably owing to differences in the GPI anchor and/or in the surrounding lipid microenvironment. Accordingly, we show that addition of cholesterol to the cells is necessary and sufficient to drive the oligomerisation and consequent apical sorting of GFP-PrP, which under control conditions does not oligomerise and is basolaterally sorted.
Paladino
Simona
S
Dipartimento di Biologia e Patologia Cellulare e Molecolare, Università degli Studi di Napoli Federico II.
Lebreton
Stephanie
S
Tivodar
Simona
S
Campana
Vincenza
V
Tempre
Rosaria
R
Zurzolo
Chiara
C
D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism; D002460:Cell Line; D002784:Cholesterol / Q000378:metabolism; D004285:Dogs; D004847:Epithelial Cells / Q000378:metabolism; D058973:Folate Receptors, GPI-Anchored; D017261:Glycosylphosphatidylinositols / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism*; D011328:Prions / Q000378:metabolism; D021381:Protein Transport; D011956:Receptors, Cell Surface / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism
D002352:Carrier Proteins; D058973:Folate Receptors, GPI-Anchored; D017261:Glycosylphosphatidylinositols; D008565:Membrane Proteins; D011328:Prions; D011956:Receptors, Cell Surface; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D002784:Cholesterol