Chemical chaperones interfere with the formation of scrapie prion protein.

Tatzelt, Jörg; Prusiner, Stanley B.; Welch, William J.

doi:10.1002/j.1460-2075.1996.tb01027.x

Cited by 288 publications

(225 citation statements)

References 69 publications

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“…Solid media contained 2.3% agar (Difco). Mouse neuroblastoma (N2a) cells (31) were cultured as described previously (32). PrP was detected by the monoclonal anti-PrP antibody 3F4 (33) or the polyclonal anti-PrP antiserum A7 (10).…”

Section: Methodsmentioning

confidence: 99%

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Post-translational Import of the Prion Protein into the Endoplasmic Reticulum Interferes with Cell Viability

Heller

Winklhofer

Heske

et al. 2003

Journal of Biological Chemistry

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Aberrant folding of the mammalian prion protein (PrP) is linked to prion diseases in humans and animals. We show that during post-translational targeting of PrP to the endoplasmic reticulum (ER) the putative transmembrane domain induces misfolding of PrP in the cytosol and interferes with its import into the ER. Unglycosylated and misfolded PrP with an uncleaved N-terminal signal sequence associates with ER membranes, and, moreover, decreases cell viability. PrP expressed in the cytosol, lacking the N-terminal ER targeting sequence, also adopts a misfolded conformation; however, this has no adverse effect on cell growth. PrP processing, productive ER import, and cellular viability can be restored either by deleting the putative transmembrane domain or by using a N-terminal signal sequence specific for co-translational ER import. Our study reveals that the putative transmembrane domain features in the formation of misfolded PrP conformers and indicates that post-translational targeting of PrP to the ER can decrease cell viability.Prion diseases in humans and animals are characterized by the accumulation of PrP Sc , a partially protease-resistant isoform of the cellular prion protein PrP C . PrP Sc is generated through a conformational transformation of PrP C and represents the major component of infectious prions (reviewed in Refs. 1-4). PrP 1 is post-translationally modified by the attachment of two N-linked complex carbohydrate moieties (Asn 180 and Asn 196 ) (5-7) and a glycosylphosphatidylinositol (GPI) anchor at serine 231 (8) as well as by the formation of a disulfide bond between Cys 178 and Cys 213 . Studies with recombinant PrP purified from bacteria revealed that the formation of the disulfide bond is essential for the native folding of PrP (9).The co-and post-translational modifications of PrP C are initiated with the cleavage of the N-terminal signal peptide (aa 1-22) and the transfer of core glycans, whereas the nascent chain is still associated with the translocon. Shortly after the protein is fully translocated, the GPI anchor is attached to the acceptor amino acid close to the C terminus. The final maturation of PrP C includes the processing of the core glycans into complex-type glycans by a series of enzymatic reactions in the endoplasmic reticulum (ER) and Golgi compartment. Posttranslational modifications, like N-linked glycosylation and GPI anchor attachment, are often used as diagnostic markers to monitor efficient import into the ER. In the case of PrP, however, we and others have shown that PrP devoid of a GPI anchor remains mainly unglycosylated but is imported efficiently into the ER and transported through the secretory pathway (10 -14). It has been found that the only specific marker for ER import of PrP is a cleaved N-terminal signal sequence (10). 2 Misfolding of PrP C in the cytosol or in the ER can induce neurodegeneration in the absence of PrP Sc . Neurotoxic properties of cytosolic PrP aggregates were observed after proteasomal inhibition in cultured cells or after the forced expr...

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Section: Methodsmentioning

confidence: 99%

“…Western Blotting-Following SDS-PAGE, proteins were transferred onto a nitrocellulose membrane and analyzed as described previously (32).…”

Section: Methodsmentioning

confidence: 99%

Post-translational Import of the Prion Protein into the Endoplasmic Reticulum Interferes with Cell Viability

Heller

Winklhofer

Heske

et al. 2003

Journal of Biological Chemistry

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“…One example of this is the effect of the 'chemical chaperone' trimethylamide N-oxide (TMAO) on PrP misfolding. Experimentally, it was found that addition of TMAO efficiently reduced PrP Sc formation in mouse neuroblastoma cells (75% reduction with 120 mM TMAO) [167]. Simulations in the presence of 1M TMAO were performed with hamster recPrP (res.…”

Section: The Effect Of Small Molecule Ligandsmentioning

confidence: 99%

Molecular Dynamics as an Approach to Study Prion Protein Misfolding and the Effect of Pathogenic Mutations

Kamp

Daggett

2011

Topics in Current Chemistry

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“…43 and 44). Several in vitro and cell culture studies revealed that chaperones have an impact on the conversion of the prion protein and the aggregation of polyglutamine proteins (45)(46)(47)(48)(49)(50)(51)(52)(53)(54). The potential of chaperones in preventing or reducing the toxicity of misfolded proteins has finally been established in animal models showing that increased expression of Hsp70 and Hsp40 suppressed the toxicity of ␣-synuclein and polyglutamine proteins (55)(56)(57)(58)(59).…”

Section: Inactivation Of a Pathogenic Parkin Mutant By Spontaneousmentioning

confidence: 99%

Inactivation of Parkin by Oxidative Stress and C-terminal Truncations

Winklhofer

Henn

Kay-Jackson

et al. 2003

Journal of Biological Chemistry

127

111

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Loss of parkin function is linked to autosomal recessive juvenile parkinsonism. Here we show that proteotoxic stress and short C-terminal truncations induce misfolding of parkin. As a consequence, wild-type parkin was depleted from a high molecular weight complex and inactivated by aggregation. Similarly, the pathogenic parkin mutant W453Stop, characterized by a Cterminal deletion of 13 amino acids, spontaneously adopted a misfolded conformation. Mutational analysis indicated that C-terminal truncations exceeding 3 amino acids abolished formation of detergent-soluble parkin. In the cytosol scattered aggregates of misfolded parkin contained the molecular chaperone Hsp70. Moreover, increased expression of chaperones prevented aggregation of wild-type parkin and promoted folding of the W453Stop mutant. Analyzing parkin folding in vitro indicated that parkin is aggregation-prone and that its folding is dependent on chaperones. Our study demonstrates that C-terminal truncations impede parkin folding and reveal a new mechanism for inactivation of parkin.Autosomal recessive juvenile parkinsonism (AR-JP), 1 the major cause of early onset parkinsonism, is characterized by mutations within the parkin gene. Parkin, a 465-amino acid protein, shows homology to ubiquitin at the N terminus and harbors a RING box near the C terminus, consisting of two RING finger motifs that flank a cysteine-rich domain (in-between RING fingers domain) (1, 2). Functional studies established that parkin acts as a ubiquitin-protein isopeptide ligase and that pathogenic mutations compromise this activity (3-6). As a consequence, substrates destined for proteasomal degradation via parkin might accumulate in parkin-deficient cells. Indeed, recent studies with cell culture models provide experimental evidence for such a scenario. It was shown that disease-related mutations in the parkin gene impair protein interactions of parkin with either a parkin substrate or another component of the ubiquitin ligase complex. Accumulation of Pael-R, one of the identified parkin substrates, causes endoplasmic reticulum (ER) stress, indicating that parkin has the potential to suppress unfolded protein stress-induced cell death (3, 4). Recent studies (7) revealed that parkin deficiency potentiates the accumulation of cyclin E and promotes apoptosis in neuronal cells exposed to excitotoxic stress. Interestingly, parkin is a significant component of Lewy bodies, the histopathologic hallmark of PD (5,8,9). Furthermore, it has been shown that parkin is protective against the toxic effects of proteasomal dysfunction and mutant ␣-synuclein (10), implying that the impact of parkin function and dysfunction might not be restricted to the entity of AR-JP.It still remains enigmatic why dopaminergic neurons in the substantia nigra are highly vulnerable in AR-JP, as parkin as well as its substrates identified so far are not selectively expressed in these cells. However, an inherent feature of dopaminergic neurons is an elevated level of reactive oxygen and nitrogen species due to...

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Chemical chaperones interfere with the formation of scrapie prion protein.

Cited by 288 publications

References 69 publications

Post-translational Import of the Prion Protein into the Endoplasmic Reticulum Interferes with Cell Viability

Post-translational Import of the Prion Protein into the Endoplasmic Reticulum Interferes with Cell Viability

Molecular Dynamics as an Approach to Study Prion Protein Misfolding and the Effect of Pathogenic Mutations

Inactivation of Parkin by Oxidative Stress and C-terminal Truncations

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