Characterization of prion protein-enriched domains, isolated from rat cerebellar granule cells in culture.
The biological functions of prion protein (PrP(C)) and its possible interaction with other specific molecular membrane partners remain largely unknown. The aim of this study is to gain information on the molecular environment of PrP(C) by analyzing the lipid and protein composition of a PrP(C)-enriched membrane subfraction, called prion domain, PrD. This domain was obtained by immunoprecipitation of detergent-resistant microdomains (DRM) of rat cerebellar granule cells under conditions designed to preserve lipid-mediated membrane organization. The electrophoretic pattern of PrD, after staining with Coomassie blue, showed the enrichment of some protein bands in comparison with DRM. microLiquid chromatography-electrospray ionization-mass spectrometry (microLC-ESI-MS)/MS analysis showed that Thy-1 and different types of myosin were strongly enriched in PrD and, in a lesser extent, also OBCAM, LSAMP and tubulin, present altogether in a single band. Experiments using the chemical cross-linker BS(3) suggested the existence of an interaction between PrP(C) and neural cell adhesion molecule (NCAM). Concerning lipids, the comparison between PrD and DRM showed a similar phospholipid/sphingolipid ratio, a phospholipid/cholesterol ratio doubled, and a strong decrease of plasmenilethanolamine (19.7 +/- 3.5% vs. 38.3 +/- 1.2%). In conclusion, the peculiar lipid composition and in particular the presence of proteins involved in synaptic plasticity, cell adhesion, cytoskeleton regulation and signalling, suggest an important physiological role in neurons of Prion Domain.
Farina
Francesca
F
Department of Experimental Medicine (DIMS), University of Milano-Bicocca, Monza, Italy.
Botto
Laura
L
Chinello
Clizia
C
Cunati
Diana
D
Magni
Fulvio
F
Masserini
Massimo
M
Palestini
Paola
P
D000818:Animals; D002448:Cell Adhesion / Q000502:physiology; D002478:Cells, Cultured; D002531:Cerebellum / Q000737:chemistry / Q000166:cytology* / Q000378:metabolism*; D003599:Cytoskeleton / Q000502:physiology; D008563:Membrane Lipids / Q000502:physiology; D009473:Neuronal Plasticity / Q000502:physiology; D011328:Prions / Q000737:chemistry* / Q000235:genetics / Q000502:physiology; D017434:Protein Structure, Tertiary / Q000502:physiology; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction / Q000502:physiology
D008563:Membrane Lipids; D011328:Prions