Doppel and PrPC co-immunoprecipitate in detergent-resistant membrane domains of epithelial FRT cells
| Autor: | Chiara Zurzolo, Maddalena Costanzo, Alessandro Negro, Vincenza Campana, Daniela Sarnataro, M. Catia Sorgato, Anna Caputo |
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| Přispěvatelé: | Caputo, A., Sarnataro, Daniela, Campana, V., Costanzo, M., Negro, A., Sorgato, C. M., Zurzolo, Chiara, Dipartimento di Biologia e Patologia Cellulare e Moleculare, University of Naples Federico II = Università degli studi di Napoli Federico II, Trafic membranaire et Pathogénèse, Institut Pasteur [Paris] (IP), Department of Biological Chemistry and Consiglio Nazionale delle Ricerche, Università degli Studi di Padova = University of Padua (Unipd), MURST (Ministero dell'Istruzione, dell'Universit`a e della Ricerca) [grant numbers PRIN 2005, PRIN 2006, FIRB 2003 to C.Z. and M.C.S.], the Telethon Foundation [grant numbers GGP0414, GTF03001], and from the European Union [grant number LSHB-CT-2006–019090 to C.Z.]. D.S. is the recipient of a fellowship from the Research Programme FIRB 2003., Università degli studi di Napoli Federico II, Institut Pasteur [Paris], Universita degli Studi di Padova |
| Rok vydání: | 2009 |
| Předmět: |
DRM
detergent-resistant membrane animal diseases TSE transmissible spongiform encephalopathy Thyroid Gland MESH: Membrane Microdomains Scrapie Plasma protein binding Biochemistry PrPC cellular prion protein 0302 clinical medicine MβCD methyl-β-cyclodextrin Dpl/ PrP/lipid rafts/protein trafficking/prions/epithelial cells MESH: Animals FRT Fischer rat thyroid Lipid raft Cells Cultured GFP green fluorescent protein 0303 health sciences MDCK Medin–Darby canine kidney TRITC tetramethylrhodamine β-isothiocyanate PNGase F peptide N-glycosidase F HRP horseradish peroxidase Transfection Cell biology PrPSc scrapie prion protein [SDV.MP]Life Sciences [q-bio]/Microbiology and Parasitology MESH: Epithelial Cells protein trafficking MESH: Membrane Proteins mo murine Research Article Protein Binding MESH: Cells Cultured GPI glycosylphosphatidylinositol Cell signaling MESH: Rats Prions doppel (Dpl) epithelial cell lipid raft prion protein (PrP) Biology CNS central nervous system GPI-Linked Proteins Endocytosis ER endoplasmic reticulum 03 medical and health sciences Membrane Microdomains MESH: Prions FBS fetal bovine serum MEV mevinolin Animals Immunoprecipitation MESH: Protein Binding PrPC Proteins MESH: PrPC Proteins Dpl doppel Molecular Biology CNX calnexin 030304 developmental biology hu human MESH: Rats Inbred F344 MESH: Immunoprecipitation Membrane Proteins Epithelial Cells Cell Biology endo H endoglycosidase H Molecular biology Rats Inbred F344 Rats MESH: Thyroid Gland nervous system diseases Membrane protein EEA1 early endosome antigen 1 Ectopic expression 030217 neurology & neurosurgery |
| Zdroj: | Biochemical Journal Biochemical Journal, 2010, 425 (2), pp.341-51. ⟨10.1042/BJ20091050⟩ Biochemical Journal, Portland Press, 2010, 425 (2), pp.341-51. ⟨10.1042/BJ20091050⟩ |
| ISSN: | 1470-8728 0264-6021 |
| DOI: | 10.1042/bj20091050 |
| Popis: | International audience; Dpl (doppel) is a paralogue of the PrPC (cellular prion protein), whose misfolded conformer (the scrapie prion protein, PrPSc) is responsible for the onset of TSEs (transmissible spongiform encephalopathies) or prion diseases. It has been shown that the ectopic expression of Dpl in the brains of some lines of PrP-knockout mice provokes cerebellar ataxia, which can be rescued by the reintroduction of the PrP gene, suggesting a functional interaction between the two proteins. It is, however, still unclear where, and under which conditions, this event may occur. In the present study we addressed this issue by analysing the intracellular localization and the interaction between Dpl and PrPC in FRT (Fischer rat thyroid) cells stably expressing the two proteins separately or together. We show that both proteins localize prevalently on the basolateral surface of FRT cells, in both singly and doubly transfected clones. Interestingly we found that they associate with DRMs (detergent-resistant membranes) or lipid rafts, from where they can be co-immunoprecipitated in a cholesterol-dependent fashion. Although the interaction between Dpl and PrPC has been suggested before, our results provide the first clear evidence that this interaction occurs in rafts and is dependent on the integrity of these membrane microdomains. Furthermore, both Dpl and PrPC could be immunoprecipitated with flotillin-2, a raft protein involved in endocytosis and cell signalling events, suggesting that they share the same lipid environment. |
| Databáze: | OpenAIRE |
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