Immunoseparation of Prion protein-enriched domains from other detergent-resistant membrane fractions, isolated from neuronal cells.
The possibility of coexistence of different subtypes of membrane lipid rafts has been investigated in cerebellar granule cells, by submitting detergent-resistant membrane fractions to immunoprecipitation. Among the proteins and lipids present in detergent-resistant fractions, almost all Prion protein, GAP43 and PKC were present in the immunoprecipitate obtained with anti-GAP43 or anti-Prion protein antibody at 4 degrees C, together with a small fraction of cholesterol and sphingolipids, suggesting that they belong to a distinct subset of membranes. On the contrary, all Fyn and almost all MARCKS remained in the supernatant. Fluorescence microscopy experiments showed that Fyn and Prion protein were mostly not colocalized within a single neuron. Our results suggest that granule cells membranes contains different subtypes of detergent-resistant fractions, possibly deriving from different lipid rafts.
Botto
Laura
L
Department of Experimental, Environmental Medicine and Biotechnologies (DIMESAB), Medical School, University of Milano-Bicocca, Via Cadore 48, 20052 Monza, Italy.
Masserini
Massimo
M
Cassetti
Arianna
A
Palestini
Paola
P
D000818:Animals; D002531:Cerebellum / Q000166:cytology; D003902:Detergents; D005455:Fluorescent Antibody Technique; D019922:GAP-43 Protein / Q000302:isolation & purification; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins / Q000302:isolation & purification*; D008856:Microscopy, Fluorescence; D000076250:Myristoylated Alanine-Rich C Kinase Substrate; D009419:Nerve Tissue Proteins / Q000302:isolation & purification; D009474:Neurons / Q000737:chemistry* / Q000166:cytology; D011328:Prions / Q000302:isolation & purification*; D011493:Protein Kinase C / Q000302:isolation & purification; D011506:Proteins / Q000302:isolation & purification; D051381:Rats; D017207:Rats, Sprague-Dawley
D003902:Detergents; D019922:GAP-43 Protein; D047908:Intracellular Signaling Peptides and Proteins; C000619712:Marcks protein, rat; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D011328:Prions; D011506:Proteins; D000076250:Myristoylated Alanine-Rich C Kinase Substrate; D011493:Protein Kinase C