Carboxyl-terminal Fragments of Alzheimer β-Amyloid Precursor Protein Accumulate in Restricted and Unpredicted Intracellular Compartments in Presenilin 1-deficient Cells

Fusheng Chen,Toshitaka Kawarai,Richard Rozmahel,David Westaway,Sam Gandy,Jorge Ghiso,Gang Yu,Suzana Petanceska,Julia Mills,Anurag Tandon,Ekaterina Rogaeva,Dong Mei Zhang,Paul E Fraser,Peter St George-Hyslop,Howard T.j Mount ,Austin J Yang ,Lyne Lévesque ,You‐Qiang Song ,Georges Lévesque ,Dun Sheng Yang ,Masaki Nakagawa

doi:10.1074/jbc.m006986200

Abstract

Absence of functional presenilin 1 (PS1) protein leads to loss of gamma-secretase cleavage of the amyloid precursor protein (betaAPP), resulting in a dramatic reduction in amyloid beta peptide (Abeta) production and accumulation of alpha- or beta-secretase-cleaved COOH-terminal fragments of betaAPP (alpha- or beta-CTFs). The major COOH-terminal fragment (CTF) in brain was identified as betaAPP-CTF-(11-98), which is consistent with the observation that cultured neurons generate primarily Abeta-(11-40). In PS1(-/-) murine neurons and fibroblasts expressing the loss-of-function PS1(D385A) mutant, CTFs accumulated in the endoplasmic reticulum, Golgi, and lysosomes, but not late endosomes. There were some subtle differences in the subcellular distribution of CTFs in PS1(-/-) neurons as compared with PS1(D385A) mutant fibroblasts. However, there was no obvious redistribution of full-length betaAPP or of markers of other organelles in either mutant. Blockade of endoplasmic reticulum-to-Golgi trafficking indicated that in PS1(-/-) neurons (as in normal cells) trafficking of betaAPP to the Golgi compartment is necessary before alpha- and beta-secretase cleavages occur. Thus, although we cannot exclude a specific role for PS1 in trafficking of CTFs, these data argue against a major role in general protein trafficking. These results are more compatible with a role for PS1 either as the actual gamma-secretase catalytic activity or in other functions indirectly related to gamma-secretase catalysis (e.g. an activator of gamma-secretase, a substrate adaptor for gamma-secretase, or delivery of gamma-secretase to betaAPP-containing compartments).

Highlights

The presenilin 1 (PS1)1 and presenilin 2 (PS2) genes encode polytopic transmembrane proteins (1, 2), which are components of high molecular weight, multimeric protein complexes predominantly located in the nuclear envelope, endoplasmic reticulum (ER), Golgi, and selected intracellular vesicular structures (3–9)
These experiments indicate that trafficking of full-length ␤APP (FL␤APP) at least as far as the Golgi apparatus is required before ␤APP-COOH-terminal fragment (CTF) begin to accumulate
A similar increase in steady state levels of CTFs was observed in cells overexpressing the PS1D385A mutation (Fig. 2) (18)

Summary

Introduction

The presenilin 1 (PS1) and presenilin 2 (PS2) genes encode polytopic transmembrane proteins (1, 2), which are components of high molecular weight, multimeric protein complexes predominantly located in the nuclear envelope, endoplasmic reticulum (ER), Golgi, and selected intracellular vesicular structures (3–9). These proteins play major roles in controlling the proteolytic processing of Notch and the ␤-amyloid precursor protein (␤APP) (10 –15). It is possible that PS1 serves as an adaptor for the ␤APP/␥-secretase reaction or that it is necessary for the activation or trafficking of ␥-secretase itself To address these questions, we have investigated the intracellular distribution of COOH-terminal ␤APP derivatives in homogenates from PS1Ϫ/Ϫ mouse. Presenilin deficiency causes no obvious abnormality in the subcellular distribution of FL␤APP or organelle marker proteins

Methods

Results

Conclusion