NR AHSB
AU Magalhaes,A.C.; Silva,J.A.; Lee,K.S.; Martins,V.R.; Prado,V.F.; Ferguson,S.S.; Gomez,M.V.; Brentani,R.R.; Prado,M.A.
TI Endocytic intermediates involved with the intracellular trafficking of a fluorescent cellular prion protein
QU The Journal of Biological Chemistry 2002 Sep 6; 277(36): 33311-8
PT journal article
AB We have investigated the intracellular traffic of PrPc, a glycosylphosphatidylinositol (GPI)-anchored protein implicated in spongiform encephalopathies. A fluorescent functional green fluorescent protein (GFP)-tagged version of PrPc is found at the cell surface and in intracellular compartments in SN56 cells. Confocal microscopy and organelle-specific markers suggest that the protein is found in both the Golgi and the recycling endosomal compartment. Perturbation of endocytosis with a dynamin I-K44A dominant-negative mutant altered the steady-state distribution of the GFP-PrPc, leading to the accumulation of fluorescence in unfissioned endocytic intermediates. These pre-endocytic intermediates did not seem to accumulate GFP-GPI, a minimum GPI-anchored protein, suggesting that PrPc trafficking does not depend solely on the GPI anchor. We found that internalized GFP-PrPc accumulates in Rab5-positive endosomes and that a Rab5 mutant alters the steady-state distribution of GFP-PrPc but not that of GFP-GPI between the plasma membrane and early endosomes. Therefore, we conclude that PrPc internalizes via a dynamin-dependent endocytic pathway and that the protein is targeted to the recycling endosomal compartment via Rab5-positive early endosomes. These observations indicate that traffic of GFP-PrPc is not determined predominantly by the GPI anchor and that, different from other GPI-anchored proteins, PrPc is delivered to classic endosomes after internalization.
MH Animal; Cell Nucleus/metabolism; Copper/metabolism; Dynamin I; Dynamins; *Endocytosis; Endosomes/metabolism; GTP Phosphohydrolases/metabolism; Genes, Dominant; Golgi Apparatus/*metabolism; Luminescent Proteins/metabolism; Mice; Microscopy, Confocal; Microscopy, Fluorescence; Mutation; Plasmids/metabolism; PrPc Proteins/*chemistry/*metabolism; Protein Binding; Protein Structure, Tertiary; Protein Transport; Recombinant Fusion Proteins/metabolism; Support, Non-U.S. Gov't; Time Factors; Transfection
AD Laboratorio de Neurofarmacologia, Departamento de Farmacologia, ICB, Universidade Federal de Minas Gerais, Belo Horizonte, Minas Gerais 31270-910, Brazil.
SP englisch
PO USA