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Subcellular trafficking abnormalities of a prion protein with a disrupted disulfide loop
Author(s) -
Yanai Anat,
Meiner Zeev,
Gahali Inbar,
Gabizon Ruth,
Taraboulos Albert
Publication year - 1999
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/s0014-5793(99)01316-2
Subject(s) - mutant , proteolysis , gene isoform , chemistry , wild type , microbiology and biotechnology , mutant protein , prion protein , biology , biochemistry , gene , enzyme , medicine , disease , pathology
The single disulfide loop (Cys 178 ‐Cys 213 ) of the prion protein (PrP) may stabilize the conformation of this protein by bridging the C‐terminal α‐helices. The substitution mutant Cys 178 Ala fails to form the prion isoform PrP Sc when expressed in scrapie‐infected neuroblastoma ScN2a cells (Muramoto et al., Proc. Natl. Acad. Sci. USA 93, 15457–15462). To investigate the reasons for this failure, we introduced the C178A substitution in the full length mouse PrP gene as well as in its N‐terminally truncated Δ23‐88 version. The resulting mutants (C178A and ΔC178A, respectively) were transiently expressed in N2a and CHO cells. Wild‐type PrP, wild‐type Δ23‐88 and the point mutant E199K served as controls in these experiments. Compared to the wild‐type controls, the C178A mutants were markedly resistant to proteolysis and they were also vastly insoluble in sarcosyl. Studying the metabolic fate of the C178A mutants, we found that in contrast to control PrP molecules, these mutants (i) remained sensitive to the diagnostic endoglycosidase EndoH, (ii) failed to reach the cell surface and (iii) congregated in large juxtanuclear spots. We surmise that these severe trafficking abnormalities may contribute both to the spontaneous aggregation of the C178A mutants and to their reported inability to form PrP Sc .