Enhanced Production and Oligomerization of the 42-residue Amyloid β-Protein by Chinese Hamster Ovary Cells Stably Expressing Mutant Presenilins

doi:10.1074/jbc.272.12.7977

Enhanced Production and Oligomerization of the 42-residue Amyloid β-Protein by Chinese Hamster Ovary Cells Stably Expressing Mutant Presenilins*

From the Department of ^‡ Neurology and
** Pathology, Harvard Medical School and Center for Neurologic Diseases, Brigham and Women's Hospital, Boston, Massachusetts 02115,
^¶ Athena Neurosciences, Inc., South San Francisco, California 94080, and the
^∥ University of Heidelberg, Central Institute of Mental Health, J5-D-68159 Mannheim, Germany

^§§ To whom correspondence should be addressed. Tel.: 617-732-6454; Fax: 617-732-7787; E-mail: selkoe{at}cnd.bwh.harvard.edu

↵^‡‡ Current address: Dept. of Neuroscience, University of California, San Diego, La Jolla, CA 92093.

Abstract

Mutations in the presenilin 1 (PS1) and presenilin 2 (PS2) genes cause the most common and aggressive form of early onset familial Alzheimer's disease. To elucidate their pathogenic mechanism, wild-type (wt) or mutant (M146L, C410Y) PS1 and wt or mutant (M239V) PS2 genes were stably transfected into Chinese hamster ovary cells that overexpress the β-amyloid precursor protein (APP). The identity of the 43-45-kDa PS1 holoproteins was confirmed by N-terminal radiosequencing. PS1 was rapidly processed (t_1/2 = 40 min) in the endoplasmic reticulum into stable fragments. Wild-type and mutant PS2 holoproteins exhibited similar half lives (1.5 h); however, their endoproteolytic fragments showed both mutation-specific and cell type-specific differences. Mutant PS1 or PS2 consistently induced a 1.4-2.5-fold increase (p < 0.001) in the relative production of the highly amyloidogenic 42-residue form of amyloid β-protein (Aβ₄₂) as determined by quantitative immunoprecipitation and by enzyme-linked immunosorbent assay. In mutant PS1 and PS2 cell lines with high increases in Aβ₄₂/Aβ_total ratios, spontaneous formation of low molecular weight oligomers of Aβ₄₂ was observed in media, suggesting enhanced Aβ aggregation from the elevation of Aβ₄₂. We conclude that mutant PS1 and PS2 proteins enhance the proteolysis of β-amyloid precursor protein by the γ-secretase cleaving at Aβ residue 42, thereby promoting amyloidogenesis.

Footnotes

↵^§ These authors contributed equally to this work.
↵* This work was supported by National Institutes of Health Grants AG06173 (to D. J. S.) and AG12376 (to E. H. K.) and the Foundation for Neurologic Diseases. The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¹ The abbreviations used are:

AD

Alzheimer's disease

Aβ

amyloid β-protein

CHO

Chinese hamster ovary

PS

presenilin

APP

β-amyloid precursor protein

ER

endoplasmic reticulum

BFA

brefeldin A

ELISA

enzyme-linked immunosorbent assay

wt

wild-type

CHAPS

3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonic acid.
↵³ M. Citron and D. J. Selkoe, unpublished data.
- Received December 4, 1996.
- Revision received January 14, 1997.