Interplays Between Covalent Modifications in the Endoplasmic Reticulum Increase Conformational Diversity in Nascent Prion Protein

Orsi, Andrea; Sitia, Roberto

doi:10.4161/pri.1.4.5727

Cited by 5 publications

(5 citation statements)

References 28 publications

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“…Of these post-translational modifications, glycosylation and disulfide bond formation depend on the cellular redox state (22,27). Impairing the ER oxidative environment or mutating the cysteines yields intracellular, diglycosylated PrP chains lacking the disulfide bond, which resembles the PrP␣3M mutants described here (22,27,30,32). In this work we demonstrated the interplay between oxidative folding and the formation of the toxic CtmPrP.…”

Section: Discussionmentioning

confidence: 88%

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Failure of Prion Protein Oxidative Folding Guides the Formation of Toxic Transmembrane Forms

Lisa

Domingo

Martínez

et al. 2012

Journal of Biological Chemistry

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Background:In vivo folding could play an essential role in prion neurodegenerations. Results: Artificial mutants causing labile PrP folds when expressed in cells originate toxic CtmPrP featured by the absence of the intramolecular disulfide bond. Conclusion: Oxidative folding impairment facilitates the formation of the toxic PrP forms. Significance: Unveiling the mechanism facilitating the formation of toxic PrP forms is crucial for the understanding and prevention of prion disorders.The mechanism by which pathogenic mutations in the globular domain of the cellular prion protein (PrP C ) increase the likelihood of misfolding and predispose to diseases is not yet known. Differences in the evidences provided by structural and metabolic studies of these mutants suggest that in vivo folding could be playing an essential role in their pathogenesis. To address this role, here we use the single or combined M206S and M213S artificial mutants causing labile folds and express them in cells. We find that these mutants are highly toxic, fold as transmembrane PrP, and lack the intramolecular disulfide bond. When the mutations are placed in a chain with impeded transmembrane PrP formation, toxicity is rescued. These results suggest that oxidative folding impairment, as on aging, can be fundamental for the genesis of intracellular neurotoxic intermediates key in prion neurodegenerations.Prion disorders are dominant gain-of-function neurodegenerations whose pathogenesis is linked to misfolded forms of the cellular prion protein (PrP C ), 3 including the prion PrP Sc and the neurotoxic CtmPrP (1-4). PrPSc is an aggregated and proteaseresistant ␤-sheet-enriched conformer of PrP C , which self-perpetuates by the templating the conversion of cell surface PrP C (1, 4). In contrast, CtmPrP is an intracellular transmembrane form generated at the ER with neurotoxic properties (1,5,6). Despite that CtmPrP formation was associated with features of the ER translocation process several pathogenic mutations in the C-terminal domain such as H187R and E200K enhance its levels, suggesting a yet unexplored in vivo interplay between folding and the accumulation and action of this neurotoxic form (6 -8).Since the enunciation of the prion hypothesis, research has focused on the mechanism by which a native PrP C structure reorganizes and acquires self-propagative features like those of PrP Sc (9 -11). The PrP C native state was assigned to the fold adopted by the chain lacking the signal sequences and containing the disulfide bond and used as reference for testing the effect of pathogenic mutations and its conversion into active prions (10,(12)(13)(14)(15)(16). However, the in vivo folding of proteins segregating into the secretory route such as PrP is a complex process participated by the ER folding machinery. This machinery coordinates processing (signal sequences removal, addition of covalent modifications, binding of cofactors, etc.), avoids undesired aggregations, and permits the acquisition of correct structure. This global process involves m...

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Section: Discussionmentioning

confidence: 88%

“…In these mice, truncated PrP was detergent-insoluble in detergent, PK-sensitive, and with migration properties resembling those of PrP␣3M mutants. Also, Cys mutants used in cellular studies resulted in PrP forms sharing key properties with PrP␣3M mutants, but their topology and toxicity were not addressed (22,27,30,32).…”

Section: Discussionmentioning

confidence: 99%

Failure of Prion Protein Oxidative Folding Guides the Formation of Toxic Transmembrane Forms

Lisa

Domingo

Martínez

et al. 2012

Journal of Biological Chemistry

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“…The mouse neuroblastoma N2a cell line was used to assess the effect of FKBP inactivation on endogenous PrP C expression. Under control conditions, an immunoblot of PrP C revealed three isoforms that differed in extent of Asn-linked glycosylation from zero to two glycans ( Figure 1A ; Orsi et al. , 2007 ).…”

Section: Resultsmentioning

confidence: 99%

“…PrP C was immunoisolated with mAb 3F4 and analyzed by SDS–PAGE and fluorography. Band doublets are due to the presence of both precursor and GPI-anchored forms of PrP C ( Orsi et al. , 2007 ).…”

Section: Resultsmentioning

confidence: 99%

Inhibition of the FKBP family of peptidyl prolyl isomerases induces abortive translocation and degradation of the cellular prion protein

Stocki

Sawicki

Mays

et al. 2016

MBoC

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“…PrP C is co-translationally translocated into the rough endoplasmic reticulum (ER), where the N-terminal signal peptide (SP) is cleaved, and the GPI anchor is added concurrently with removal of a C-terminal signal sequence. In the ER, the PrP polypeptide undergoes oxidative folding with formation of a single disulphide bond, and the protein is variably modified at two N-glycosylation sites, resulting in a mixture of di-, mono- and unglycosylated forms [6] . After transit in the mid-Golgi, where the immature, core-glycosylated molecules are complex-glycosylated, PrP is transported through the later compartments of the secretory pathway and delivered to the cell surface, where it resides in lipid rafts [7] .…”

Section: Introductionmentioning

confidence: 99%

Cell Type-Specific Neuroprotective Activity of Untranslocated Prion Protein

et al. 2010

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BackgroundA key pathogenic role in prion diseases was proposed for a cytosolic form of the prion protein (PrP). However, it is not clear how cytosolic PrP localization influences neuronal viability, with either cytotoxic or anti-apoptotic effects reported in different studies. The cellular mechanism by which PrP is delivered to the cytosol of neurons is also debated, and either retrograde transport from the endoplasmic reticulum or inefficient translocation during biosynthesis has been proposed. We investigated cytosolic PrP biogenesis and effect on cell viability in primary neuronal cultures from different mouse brain regions.Principal FindingsMild proteasome inhibition induced accumulation of an untranslocated form of cytosolic PrP in cortical and hippocampal cells, but not in cerebellar granules. A cyclopeptolide that interferes with the correct insertion of the PrP signal sequence into the translocon increased the amount of untranslocated PrP in cortical and hippocampal cells, and induced its synthesis in cerebellar neurons. Untranslocated PrP boosted the resistance of cortical and hippocampal neurons to apoptotic insults but had no effect on cerebellar cells.SignificanceThese results indicate cell type-dependent differences in the efficiency of PrP translocation, and argue that cytosolic PrP targeting might serve a physiological neuroprotective function.

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Interplays Between Covalent Modifications in the Endoplasmic Reticulum Increase Conformational Diversity in Nascent Prion Protein

Cited by 5 publications

References 28 publications

Failure of Prion Protein Oxidative Folding Guides the Formation of Toxic Transmembrane Forms

Failure of Prion Protein Oxidative Folding Guides the Formation of Toxic Transmembrane Forms

Inhibition of the FKBP family of peptidyl prolyl isomerases induces abortive translocation and degradation of the cellular prion protein

Cell Type-Specific Neuroprotective Activity of Untranslocated Prion Protein

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