CELL BIOLOGY AND METABOLISM
Cleavage of Alzheimer's Amyloid Precursor Protein (APP) by Secretases Occurs after O-Glycosylation of APP in the Protein Secretory Pathway: IDENTIFICATION OF INTRACELLULAR COMPARTMENTS IN WHICH APP CLEAVAGE OCCURS WITHOUT USING TOXIC AGENTS THAT INTERFERE WITH PROTEIN METABOLISM*

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β-Amyloid peptide (Aβ) is a principal component of parenchymal amyloid deposits in Alzheimer's disease. Aβ is derived from amyloid precursor protein (APP) by proteolytic cleavage. APP is subject to N- and O-glycosylation and potential tyrosine sulfation, following protein synthesis, and is then thought to be cleaved in an intracellular secretory pathway after or during these post-translational modifications. Studies utilizing agents that affect a series of steps in the protein secretory pathway have identified the possible intracellular sites of APP cleavage and Aβ generation within the protein secretory pathway. In the present study, using cells with normal protein metabolism, but expressing mutant APP with defectiveO-glycosylation, we demonstrated that the majority of APP cleavage by α-, β-, and γ-secretases occurs afterO-glycosylation. Cells expressing the mutant APP noticeably decreased the generation of the intracellular APP carboxyl-terminal fragment (αAPPCOOH), a product of α-secretase, and both Aβ40 and Aβ42 in medium, a product of β- and γ-secretases. Furthermore, we found that the cells accumulate the mutant APP in intracellular reticular compartments such as the endoplasmic reticulum. Agents that could ambiguously affect the function of specific intracellular organelles and that may be toxic were not used. The present results indicate that APP is cleaved by α-, β-, and γ-secretases in step(s) during the transport of APP through Golgi complex, where O-glycosylation occurs, or in compartments subsequent to trans-Golgi of the APP secretory pathway.

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This research was supported in part by a grant from Program for Promotion of Basic Research Activity for Innovative Bioscience.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.

Recipient of Japan Society for the Promotion of Science Research Fellowships for Young Scientists.