The Metabolism and Imaging in Live Cells of the Bovine Prion Protein in Its Native Form or Carrying Single Amino Acid Substitutions

Negro, Alessandro; Ballarin, Cristina; Bertoli, Alessandro; Massimino, Maria Lina; Sorgato, Maria Catia

doi:10.1006/mcne.2000.0953

Cited by 64 publications

(55 citation statements)

References 66 publications

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“…Pulse-chase labeling experiments indicated that mutant PrP molecules misfold very soon after synthesis in the endoplasmic reticulum (ER) (6), raising the possibility that they are recognized as abnormal by the ER quality control machinery, and diverted to ER-associated degradation (ERAD). Consistent with this hypothesis, it was observed that several mutant PrPs were present at low levels on the cell surface, and localized in intracellular compartments, including the ER and cytoplasm (7)(8)(9)(10). It was also found that the proteasome inhibitor ALLN (Ac-Leu-Leu-NorLeu-al) affected the metabolism and cellular localization of PrP molecules carrying the amber mutation Y145stop, and the Q217R substitution linked to GSS (11,12).…”

mentioning

confidence: 56%

Cytosolic Prion Protein (PrP) Is Not Toxic in N2a Cells and Primary Neurons Expressing Pathogenic PrP Mutations

Fioriti

Dossena

Stewart

et al. 2005

Journal of Biological Chemistry

111

117

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Inherited prion diseases are linked to mutations in the prion protein (PrP) gene, which favor conversion of PrP into a conformationally altered, pathogenic isoform. The cellular mechanism by which this process causes neurological dysfunction is unknown. It has been proposed that neuronal death can be triggered by accumulation of PrP in the cytosol because of impairment of proteasomal degradation of misfolded PrP molecules retrotranslocated from the endoplasmic reticulum (Ma, J., Wollmann, R., and Lindquist, S. (2002) Science 298, 1781-1785). To test whether this neurotoxic mechanism is operative in inherited prion diseases, we evaluated the effect of proteasome inhibitors on the viability of transfected N2a cells and primary neurons expressing mouse PrP homologues of the D178N and nine octapeptide mutations. We found that the inhibitors caused accumulation of an unglycosylated, aggregated form of PrP exclusively in transfected N2a expressing PrP from the cytomegalovirus promoter. This form contained an uncleaved signal peptide, indicating that it represented polypeptide chains that had failed to translocate into the ER lumen during synthesis, rather than retrogradely translocated PrP. Quantification of N2a viability in the presence of proteasome inhibitors demonstrated that accumulation of this form was not toxic. No evidence of cytosolic PrP was found in cerebellar granule neurons from transgenic mice expressing wild-type or mutant PrPs from the endogenous promoter, nor were these neurons more susceptible to proteasome inhibitor toxicity than neurons from PrP knock-out mice. Our analysis fails to confirm the previous observation that mislocation of PrP in the cytosol is neurotoxic, and argues against the hypothesis that perturbation of PrP metabolism through the proteasomal pathway plays a pathogenic role in prion diseases.

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mentioning

confidence: 56%

Cytosolic Prion Protein (PrP) Is Not Toxic in N2a Cells and Primary Neurons Expressing Pathogenic PrP Mutations

Fioriti

Dossena

Stewart

et al. 2005

Journal of Biological Chemistry

111

117

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“…PrP C continuously cycles between the plasma membrane and endocytic compartments. This process is related to the availability of the protein at the cell surface, and involves internalization pathways that may be critical for PrP C physiological functions [72]. Trafficking of PrP C seems to be a complex cellular event and may involve more than one internalization mechanism.…”

Section: Cellular Localization and Traffickingmentioning

confidence: 99%

Physiological role of the cellular prion protein

et al. 2007

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“…Although PrP C is a highly conserved protein in vertebrates, its actual physiological role remains elusive. Recent work has shown that PrP C plays pleiotrophic roles in neuronal and non-neuronal cells, such as cellular trafficking, and also participates in copper uptake (Lee et al, 2001;Negro et al, 2001;Perera and Hooper, 2001), cell adhesion/differentiation (Luckenbill-Edds, 1997), cell signaling (Martins et al, 1997), and neuronal survival (Bounhar et al, 2001). Interestingly, some reports have shown that PrP C also has the ability to protect neural tissue against oxidative stress (Brown et al, 1999.…”

Section: Introductionmentioning

confidence: 99%

Overexpression of PrP^Cby Adenovirus-Mediated Gene Targeting Reduces Ischemic Injury in a Stroke Rat Model

Shyu¹,

Lin²,

Chiang³

et al. 2005

J. Neurosci.

116

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Prion diseases are induced by pathologically misfolded prion protein (PrPSc ), which recruit normal sialoglycoprotein PrP C by a templatedirected process. In this study, we investigated the expression of PrP C in a rat model of cerebral ischemia to more fully understand its physiological role. Immunohistochemical analysis demonstrated that PrP C -immunoreactive cells increased significantly in the penumbra of ischemic rat brain compared with the untreated brain. Western blot analysis showed that PrP C protein expression increased in ischemic brain tissue in a time-dependent manner. In addition, PrP C protein expression was seen to colocalize with neuron, glial, and vascular endothelial cells in the penumbric region of the ischemic brain. Overexpression of PrP C by injection of rAd (replication-defective recombinant adenoviral)-PGK (phosphoglycerate kinase)-PrP C -Flag into ischemic rat brain improved neurological behavior and reduced the volume of cerebral infarction, which is supportive of a role for PrP C in the neuroprotective adaptive cellular response to ischemic lesions. Concomitant upregulation of PrP C and activated extracellular signal-regulated kinase (ERK1/2) under hypoxia-reoxygenation in primary cortical cultures was shown to be dependent on ERK1/2 phosphorylation. During hypoxia-reoxygenation, mouse neuroblastoma cell line N18 cells transfected with luciferase rat PrP C promoter reporter constructs, containing the heat shock element (HSE), expressed higher luciferase activities (3-to 10-fold) than those cells transfected with constructs not containing HSE. We propose that HSTF-1 (hypoxia-activated transcription factor), phosphorylated by ERK1/2, may in turn interact with HSE in the promoter of PrP C resulting in gene expression of the prion gene. In summary, we conclude that upregulation of PrP C expression after cerebral ischemia and hypoxia exerts a neuroprotective effect on injured neural tissue. This study suggests that PrP C has physiological relevance to cerebral ischemic injury and could be useful as a therapeutic target for the treatment of cerebral ischemia.

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The Metabolism and Imaging in Live Cells of the Bovine Prion Protein in Its Native Form or Carrying Single Amino Acid Substitutions

Cited by 64 publications

References 66 publications

Cytosolic Prion Protein (PrP) Is Not Toxic in N2a Cells and Primary Neurons Expressing Pathogenic PrP Mutations

Cytosolic Prion Protein (PrP) Is Not Toxic in N2a Cells and Primary Neurons Expressing Pathogenic PrP Mutations

Physiological role of the cellular prion protein

Overexpression of PrP^Cby Adenovirus-Mediated Gene Targeting Reduces Ischemic Injury in a Stroke Rat Model

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The Metabolism and Imaging in Live Cells of the Bovine Prion Protein in Its Native Form or Carrying Single Amino Acid Substitutions

Cited by 64 publications

References 66 publications

Cytosolic Prion Protein (PrP) Is Not Toxic in N2a Cells and Primary Neurons Expressing Pathogenic PrP Mutations

Cytosolic Prion Protein (PrP) Is Not Toxic in N2a Cells and Primary Neurons Expressing Pathogenic PrP Mutations

Physiological role of the cellular prion protein

Overexpression of PrPCby Adenovirus-Mediated Gene Targeting Reduces Ischemic Injury in a Stroke Rat Model

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Overexpression of PrP^Cby Adenovirus-Mediated Gene Targeting Reduces Ischemic Injury in a Stroke Rat Model