title: Cytosolic prion protein is not toxic and protects against Bax-mediated cell death in human primary neurons.
abstract: Recently, it was observed that reverse-translocated cytosolic PrP and PrP expressed in the cytosol induce rapid death in neurons (Ma, J., Wollmann, R., and Lindquist, S. (2002) Science 298, 1781-1785). In this study, we investigated whether accumulation of prion protein (PrP) in the cytosol is toxic to human neurons in primary culture. We show that in these neurons, a single PrP isoform lacking signal peptide accumulates in the cytosol of neurons treated with epoxomicin, a specific proteasome inhibitor. Therefore, endogenously expressed PrP is subject to the endoplasmic reticulum-associated degradation (ERAD) pathway and is degraded by the proteasome in human primary neurons. In contrast to its toxicity in N2a cells, reverse-translocated PrP (ERAD-PrP) is not toxic even when neurons are microinjected with cDNA constructs to overexpress either wild-type PrP or mutant PrPD178N. We found that ERAD-PrP in human neurons remains detergent-soluble and proteinase K-sensitive, in contrast to its detergent-insoluble and proteinase K-resistant state in N2a cells. Furthermore, not only is microinjection of a cDNA construct expressing CyPrP not toxic, it protects these neurons against Bax-mediated cell death. We conclude that in human neurons, ERAD-PrP is not converted naturally into a form reminiscent of scrapie PrP and that PrP located in the cytosol retains its protective function against Bax. Thus, it is unlikely that simple accumulation of PrP in the cytosol can cause neurodegeneration in prion diseases.
authors.affiliation: Bloomfield Center for Research in Aging, Lady Davis Institute for Medical Research, Jewish General Hospital.
authors.forename: Xavier
authors.initials: X
authors.lastname: Roucou
authors.forename: Qi
authors.initials: Q
authors.lastname: Guo
authors.forename: Yan
authors.initials: Y
authors.lastname: Zhang
authors.forename: Cynthia G
authors.initials: CG
authors.lastname: Goodyer
authors.forename: Andrea C
authors.initials: AC
authors.lastname: LeBlanc
mesh_terms: D015153:Blotting, Western; D016923:Cell Death; D002478:Cells, Cultured; D003001:Cloning, Molecular; D003600:Cytosol / Q000378:metabolism*; D018076:DNA, Complementary / Q000378:metabolism; D019286:Endopeptidase K / Q000494:pharmacology; D004721:Endoplasmic Reticulum / Q000378:metabolism; D017353:Gene Deletion; D006801:Humans; D020287:In Situ Nick-End Labeling; D009447:Neuroblastoma / Q000378:metabolism; D009474:Neurons / Q000378:metabolism*; D011328:Prions / Q000378:metabolism*; D021381:Protein Transport; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D019253:Proto-Oncogene Proteins c-bcl-2; D013347:Subcellular Fractions; D014162:Transfection; D051028:bcl-2-Associated X Protein
chemical_list: C494826:BAX protein, human; D018076:DNA, Complementary; D011328:Prions; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; D019286:Endopeptidase K