Endoplasmic Reticulum and Trans-Golgi Network Generate Distinct Populations of Alzheimer β -Amyloid Peptides

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 96; no. 2; pp. 742 - 747
• Main Authors: Greenfield, Jeffrey P., Tsai, Julia, Gouras, Gunnar K., Hai, Bing, Thinakaran, Gopal, Checler, Frederic, Sisodia, Sangram S., Greengard, Paul, Xu, Huaxi
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences of the United States of America 19-01-1999; National Acad Sciences; National Academy of Sciences; The National Academy of Sciences
• Subjects: Alzheimer Disease - metabolism; Alzheimer's disease; Amyloid beta-Peptides - metabolism; Animals; Antibodies; Biological Sciences; Brain - metabolism; Cell Fractionation; Cell lines; Cells, Cultured; Cellular immunity; Centrifugation, Density Gradient; Endoplasmic reticulum; Endoplasmic Reticulum - metabolism; Enzymes; Fluorescence; Fluorescent antibody techniques; Fractionation; Golgi Apparatus - metabolism; Humans; Immunohistochemistry; Microscopy, Fluorescence; Neurology; Neurons; Peptide Fragments - metabolism; Peptides; Proteins; Rats; Trans golgi network
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.96.2.742

The excessive generation and accumulation of 40- and 42-aa β -amyloid peptides (Aβ40/Aβ42) in selectively vulnerable brain regions is a major neuropathological feature of Alzheimer's disease. Aβ , derived by proteolytic cleavage from the β -amyloid precursor protein (β APP) is normally secreted. However, recent evidence suggests that significant levels of Aβ also may remain inside cells. Here, we have investigated the subcellular compartments within which distinct amyloid species are generated and the compartments from which they are secreted. Three experimental approaches were used: (i) immunofluorescence performed in intact cortical neurons; (ii) sucrose gradient fractionation performed with mouse neuroblastoma cells stably expressing wild-type β APP695(N2a695); and (iii) cell-free reconstitution of Aβ generation and trafficking from N2a695cells. These studies demonstrate that: (i) Aβ40(Aβ1-40plus Aβx-40, where x is an NH2-terminal truncation) is generated exclusively within the trans-Golgi Network (TGN) and packaged into post-TGN secretory vesicles; (ii) Aβx-42is made and retained within the endoplasmic reticulum in an insoluble state; (iii) Aβ42(Aβ1-42plus Aβx-42) is made in the TGN and packaged into secretory vesicles; and (iv) the amyloid peptides formed in the TGN consist of two pools (a soluble population extractable with detergents and a detergent-insoluble form). The identification of the organelles in which distinct forms of Aβ are generated and from which they are secreted should facilitate the identification of the proteolytic enzymes responsible for their formation.