Prion Protein Contains a Second Endoplasmic Reticulum Targeting Signal Sequence Located at Its C Terminus

Hölscher, Christina; Bach, Ute; Dobberstein, Bernhard

doi:10.1074/jbc.m007331200

Cited by 45 publications

(38 citation statements)

References 42 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…We have also observed that deletion of the signal peptide prevents translocation, resulting in cytosolic molecules that are fully accessible to added protease. This result, which confirms other published reports (8,10), indicates that the hydrophobic, transmembrane segment cannot by itself target the polypeptide chain to the translocon. Thus, in contrast to the case for other type II membrane-spanning proteins (whose N termini are cytoplasmic), the transmembrane domain of PrP is not capable of functioning as a signal-anchor sequence.…”

Section: Topological Determinants In Prprelative Amount Ofsupporting

confidence: 82%

“…Another potential topological determinant, the C-terminal, GPI addition signal, does not appear to play a significant role. Although the GPI signal can mediate post-translational translocation of PrP molecules lacking an N-terminal signal sequence (8), deletion or substitution of this segment has no effect on the membrane topology of the full-length protein (this paper and see Ref. 11).…”

Section: Topological Determinants In Prprelative Amount Ofmentioning

confidence: 99%

“…This segment, which is normally cleaved off during addition of the anchor structure in the ER, is capable of inserting into the membrane post-translationally when the N-terminal signal sequence and transmembrane domains have been deleted (8). To test the effect of the GPI addition sequence on the membrane topology of PrP in the context of the full-length protein, we analyzed constructs in which this sequence was deleted.…”

Section: Residues On the C-terminal Side Of The Signal Peptide Cleavamentioning

confidence: 99%

See 2 more Smart Citations

Mutational Analysis of Topological Determinants in Prion Protein (PrP) and Measurement of Transmembrane and Cytosolic PrP during Prion Infection

Stewart

Harris

2003

Journal of Biological Chemistry

View full text Add to dashboard Cite

The prion protein (PrP) can adopt multiple membrane topologies, including a fully translocated form ( Sec PrP), two transmembrane forms ( Ntm PrP and Ctm PrP), and a cytosolic form. It is important to understand the factors that influence production of these species, because two of them, Ctm PrP and cytosolic PrP, have been proposed to be key neurotoxic intermediates in certain prion diseases. In this paper, we perform a mutational analysis of PrP synthesized using an in vitro translation system in order to further define sequence elements that influence the formation of Ctm PrP. We find that substitution of charged residues in the hydrophobic core of the signal peptide increases synthesis of Ctm PrP and also reduces the efficiency of translocation into microsomes. Combining these mutations with substitutions in the transmembrane domain causes the protein to be synthesized exclusively with the Ctm PrP topology. Reducing the spacing between the signal peptide and the transmembrane domain also increases Ctm PrP. In contrast, topology is not altered by mutations that prevent signal peptide cleavage or by deletion of the C-terminal signal for glycosylphosphatidylinositol anchor addition. Removal of the signal peptide completely blocks translocation. Taken together, our results are consistent with a model in which the signal peptide and transmembrane domain function in distinct ways as determinants of PrP topology. We also present characterization of an antibody that selectively recognizes Ctm PrP and cytosolic PrP by virtue of their uncleaved signal peptides. By using this antibody, as well as the distinctive gel mobility of Ctm PrP and cytosolic PrP, we show that the amounts of these two forms in cultured cells and rodent brain are not altered by infection with scrapie prions. We conclude that Ctm PrP and cytosolic PrP are unlikely to be obligate neurotoxic intermediates in familial or infectiously acquired prion diseases.

show abstract

Section: Topological Determinants In Prprelative Amount Ofsupporting

confidence: 82%

Section: Topological Determinants In Prprelative Amount Ofmentioning

confidence: 99%

Section: Residues On the C-terminal Side Of The Signal Peptide Cleavamentioning

confidence: 99%

See 1 more Smart Citation

Mutational Analysis of Topological Determinants in Prion Protein (PrP) and Measurement of Transmembrane and Cytosolic PrP during Prion Infection

Stewart

Harris

2003

Journal of Biological Chemistry

View full text Add to dashboard Cite

show abstract

“…The last domain or C-terminal globular domain (Donne et al, 1997;Riek et al, 1997) contains the -helix and two parallel stranded -sheets (Riek et al, 1996). PrP c presents at least three distinct topological orientations: the fully extracellular form (or (sec)PrP) (Holscher et al, 2001), and two transmembrane isoforms (called Ntm-PrP and Ctm-PrP) with opposite sequence orientations with respect to the lumen of the endoplasmic reticulum (Hegde et al, 1998).…”

Section: Introductionmentioning

confidence: 99%

New insights into cellular prion protein (PrPc) functions: The “ying and yang” of a relevant protein

Nicolas

Gavı́n

Rı́o

2009

Brain Research Reviews

View full text Add to dashboard Cite

show abstract

“…In mammalian cells, most secretory proteins, including PrP, are targeted to the ER co-translationally via binding of the signal sequence to the signal recognition particle (SRP), which temporarily arrests translation until the ribosome docks at the ER membrane (41,64). In contrast, yeast have two pathways for ER translocation: an SRP-dependent, cotranslational pathway and an SRP-independent, post-translational pathway (42).…”

Section: Discussionmentioning

confidence: 99%

Cell Surface Expression of the Prion Protein in Yeast Does Not Alter Copper Utilization Phenotypes

Dong

Harris

2004

Journal of Biological Chemistry

View full text Add to dashboard Cite

Prion diseases are fatal neurodegenerative disorders that result from conversion of a normal, cell surface glycoprotein (PrP C ) into a conformationally altered isoform (PrP Sc ) that is thought to be infectious. Although a great deal is known about the role of PrP Sc in the disease process, the physiological function of PrP C has remained enigmatic. In this report, we have used the yeast Saccharomyces cerevisiae to test one hypothesized function of PrP C , as a receptor for the uptake or efflux of copper ions. We first modified the PrP signal peptide by replacing its hydrophobic core with the signal sequence from the yeast protein dipeptidyl aminopeptidase B, so that the resulting protein was targeted cotranslationally to the secretory pathway when synthesized in yeast. PrP molecules with the modified signal peptide were efficiently glycosylated, glycolipid-anchored, and localized to the plasma membrane. We then tested whether PrP expression altered the growth deficiency phenotypes of yeast strains harboring deletions in genes that encode key components of copper utilization pathways, including transporters, chaperones, pumps, reductases, and cuproenzymes. We found that PrP did not rescue any of these mutant phenotypes, arguing against a direct role for the protein in copper utilization. Our results provide further clarification of the physiological function of PrP C , and lay the groundwork for using PrPexpressing yeast to study other aspects of prion biology.

show abstract

Prion Protein Contains a Second Endoplasmic Reticulum Targeting Signal Sequence Located at Its C Terminus

Cited by 45 publications

References 42 publications

Mutational Analysis of Topological Determinants in Prion Protein (PrP) and Measurement of Transmembrane and Cytosolic PrP during Prion Infection

Mutational Analysis of Topological Determinants in Prion Protein (PrP) and Measurement of Transmembrane and Cytosolic PrP during Prion Infection

New insights into cellular prion protein (PrPc) functions: The “ying and yang” of a relevant protein

Cell Surface Expression of the Prion Protein in Yeast Does Not Alter Copper Utilization Phenotypes

Contact Info

Product

Resources

About