title: Conformation dependent pro-apoptotic activity of the recombinant human prion protein fragment 90-231.
abstract: The transition of prion protein from a mainly alpha-structured isoform (PrPC) to a beta sheet-containing protein (PrPSc) represents a major pathogenetic mechanism in prion diseases. To study the role of PrP structural conformation in prion-dependent neurodegeneration, we analysed the neurotoxicity of PrP in alpha and beta conformations, using a recombinant protein encompassing amino acids 90-231 of the human PrP (hPrP90-231). Using controlled thermal denaturation (53 degrees C, 1h) we converted hPrP90-231 in a structural isoform displaying PrPSc-related characteristics: high beta sheet content, increased aggregability and a slight increase in the resistance to protease K. In virtue of these structural changes, hPrP90-231 powerfully affected the survival of SH-SY5Y cells, inducing a caspase-3 and p38- dependent apoptosis. Conversely, in the native alpha-helix-rich conformation, hPrP90-231 did not show significant cell toxicity. The relationship between the structural state of hPrP90-231 and its neurotoxicity was demonstrated, inducing the thermal denaturation of the peptide in the presence of Congo red that prevented both the transition of hPrP90-231 into a beta-rich isoform and the acquisition of toxic properties. In conclusion, we report that the toxicity of hPrP90-231 is dependent on its three-dimensional structure, as is supposed to occur for the pathogen PrP during TSE.
authors.affiliation: Section of Pharmacology, Dept. Oncology Biology and Genetics, University of Genoa, Viale Benedetto XV 2, 16132 Genoa, Italy.
authors.forename: A
authors.initials: A
authors.lastname: Corsaro
authors.forename: D
authors.initials: D
authors.lastname: Paludi
authors.forename: V
authors.initials: V
authors.lastname: Villa
authors.forename: C
authors.initials: C
authors.lastname: D'Arrigo
authors.forename: K
authors.initials: K
authors.lastname: Chiovitti
authors.forename: S
authors.initials: S
authors.lastname: Thellung
authors.forename: C
authors.initials: C
authors.lastname: Russo
authors.forename: D
authors.initials: D
authors.lastname: Di Cola
authors.forename: P
authors.initials: P
authors.lastname: Ballerini
authors.forename: E
authors.initials: E
authors.lastname: Patrone
authors.forename: G
authors.initials: G
authors.lastname: Schettini
authors.forename: A
authors.initials: A
authors.lastname: Aceto
authors.forename: T
authors.initials: T
authors.lastname: Florio
mesh_terms: D000682:Amyloid / Q000096:biosynthesis; D017209:Apoptosis / Q000187:drug effects*; D052160:Benzothiazoles; D053148:Caspase 3; D053179:Caspase 7; D020169:Caspases / Q000378:metabolism; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D002942:Circular Dichroism; D019286:Endopeptidase K / Q000737:chemistry; D005456:Fluorescent Dyes; D006801:Humans; D006868:Hydrolysis; D015151:Immunoblotting; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008854:Microscopy, Electron; D009336:Necrosis; D018621:PrPC Proteins / Q000737:chemistry* / Q000494:pharmacology*; D011487:Protein Conformation; D011489:Protein Denaturation; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry / Q000494:pharmacology; D013329:Structure-Activity Relationship; D013778:Tetrazolium Salts; D013844:Thiazoles / Q000378:metabolism; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
chemical_list: D000682:Amyloid; D052160:Benzothiazoles; D005456:Fluorescent Dyes; D018621:PrPC Proteins; D011994:Recombinant Proteins; D013778:Tetrazolium Salts; D013844:Thiazoles; C009462:thioflavin T; D007770:L-Lactate Dehydrogenase; D048051:p38 Mitogen-Activated Protein Kinases; D019286:Endopeptidase K; C505407:CASP3 protein, human; C505520:CASP7 protein, human; D053148:Caspase 3; D053179:Caspase 7; D020169:Caspases; C022616:thiazolyl blue