Exclusively Targeting β-Secretase to Lipid Rafts by GPI-Anchor Addition Up-Regulates β-Site Processing of the Amyloid Precursor Protein
β-Secretase (BACE, Asp-2) is a transmembrane aspartic proteinase responsible for cleaving the amyloid precursor protein (APP) to generate the soluble ectodomain sAPPβ and its C-terminal fragment$CTF\beta.\>CTF\beta$is subsequently cleaved by β-secretase to produce the neurotoxic/synaptotoxic amyl...
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Veröffentlicht in: | Proceedings of the National Academy of Sciences - PNAS 2003-09, Vol.100 (20), p.11735-11740 |
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creator | Cordy, Joanna M. Hussain, Ishrut Dingwall, Colin Hooper, Nigel M. Turner, Anthony J. |
description | β-Secretase (BACE, Asp-2) is a transmembrane aspartic proteinase responsible for cleaving the amyloid precursor protein (APP) to generate the soluble ectodomain sAPPβ and its C-terminal fragment$CTF\beta.\>CTF\beta$is subsequently cleaved by β-secretase to produce the neurotoxic/synaptotoxic amyloid-β peptide (Aβ) that accumulates in Alzheimer's disease. Indirect evidence has suggested that amyloidogenic APP processing may preferentially occur in lipid rafts. Here, we show that relatively little wild-type BACE is found in rafts prepared from a human neuroblastoma cell line (SH-SY5Y) by using Triton X-100 as detergent. To investigate further the significance of lipid rafts in APP processing, a glycosylphosphatidylinositol (GPI) anchor has been added to BACE, replacing the transmembrane and C-terminal domains. The GPI anchor targets the enzyme exclusively to lipid raft domains. Expression of GPI-BACE substantially up-regulates the secretion of both sAPPβ and amyloid-β peptide over levels observed from cells overexpressing wild-type BACE. This effect was reversed when the lipid rafts were disrupted by depleting cellular cholesterol levels. These results suggest that processing of APP to the amyloid-β peptide occurs predominantly in lipid rafts and that BACE is the rate-limiting enzyme in this process. The processing of the APP695isoform by GPI-BACE was up-regulated 20-fold compared with wild-type BACE, whereas only a 2-fold increase in the processing of APP751/770was seen, implying a differential compartmentation of the APP isoforms. Changes in the local membrane environment during aging may facilitate the cosegregation of APP and BACE leading to increased β-amyloid production. |
doi_str_mv | 10.1073/pnas.1635130100 |
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Indirect evidence has suggested that amyloidogenic APP processing may preferentially occur in lipid rafts. Here, we show that relatively little wild-type BACE is found in rafts prepared from a human neuroblastoma cell line (SH-SY5Y) by using Triton X-100 as detergent. To investigate further the significance of lipid rafts in APP processing, a glycosylphosphatidylinositol (GPI) anchor has been added to BACE, replacing the transmembrane and C-terminal domains. The GPI anchor targets the enzyme exclusively to lipid raft domains. Expression of GPI-BACE substantially up-regulates the secretion of both sAPPβ and amyloid-β peptide over levels observed from cells overexpressing wild-type BACE. This effect was reversed when the lipid rafts were disrupted by depleting cellular cholesterol levels. These results suggest that processing of APP to the amyloid-β peptide occurs predominantly in lipid rafts and that BACE is the rate-limiting enzyme in this process. The processing of the APP695isoform by GPI-BACE was up-regulated 20-fold compared with wild-type BACE, whereas only a 2-fold increase in the processing of APP751/770was seen, implying a differential compartmentation of the APP isoforms. Changes in the local membrane environment during aging may facilitate the cosegregation of APP and BACE leading to increased β-amyloid production.</description><identifier>ISSN: 0027-8424</identifier><identifier>EISSN: 1091-6490</identifier><identifier>DOI: 10.1073/pnas.1635130100</identifier><identifier>PMID: 14504402</identifier><language>eng</language><publisher>United States: National Academy of Sciences</publisher><subject>Alzheimer's disease ; Amino Acid Sequence ; Amyloid beta-Protein Precursor - metabolism ; Amyloid Precursor Protein Secretases ; Antibodies ; Aspartic Acid Endopeptidases ; Biological Sciences ; Cell lines ; Cell membranes ; Cholesterol ; Cholesterol - metabolism ; Cholesterols ; Endopeptidases - metabolism ; Enzymes ; Glycosylphosphatidylinositols - metabolism ; Humans ; Hydrolysis ; Immunoblotting ; Lipid Metabolism ; Lipids ; Molecular Sequence Data ; Neurology ; Peptides ; Protein isoforms ; Protein Processing, Post-Translational ; Proteins ; Rafts ; Tumor Cells, Cultured</subject><ispartof>Proceedings of the National Academy of Sciences - PNAS, 2003-09, Vol.100 (20), p.11735-11740</ispartof><rights>Copyright 1993-2003 National Academy of Sciences of the United States of America</rights><rights>Copyright National Academy of Sciences Sep 30, 2003</rights><rights>Copyright © 2003, The National Academy of Sciences 2003</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c526t-f0355abd4086be9703f6a2a53bae434772197e00af08481b1a71e701053be59f3</citedby><cites>FETCH-LOGICAL-c526t-f0355abd4086be9703f6a2a53bae434772197e00af08481b1a71e701053be59f3</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Uhttp://www.pnas.org/content/100/20.cover.gif</thumbnail><linktopdf>$$Uhttps://www.jstor.org/stable/pdf/3147861$$EPDF$$P50$$Gjstor$$H</linktopdf><linktohtml>$$Uhttps://www.jstor.org/stable/3147861$$EHTML$$P50$$Gjstor$$H</linktohtml><link.rule.ids>230,314,727,780,784,803,885,27924,27925,53791,53793,58017,58250</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/14504402$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Cordy, Joanna M.</creatorcontrib><creatorcontrib>Hussain, Ishrut</creatorcontrib><creatorcontrib>Dingwall, Colin</creatorcontrib><creatorcontrib>Hooper, Nigel M.</creatorcontrib><creatorcontrib>Turner, Anthony J.</creatorcontrib><title>Exclusively Targeting β-Secretase to Lipid Rafts by GPI-Anchor Addition Up-Regulates β-Site Processing of the Amyloid Precursor Protein</title><title>Proceedings of the National Academy of Sciences - PNAS</title><addtitle>Proc Natl Acad Sci U S A</addtitle><description>β-Secretase (BACE, Asp-2) is a transmembrane aspartic proteinase responsible for cleaving the amyloid precursor protein (APP) to generate the soluble ectodomain sAPPβ and its C-terminal fragment$CTF\beta.\>CTF\beta$is subsequently cleaved by β-secretase to produce the neurotoxic/synaptotoxic amyloid-β peptide (Aβ) that accumulates in Alzheimer's disease. Indirect evidence has suggested that amyloidogenic APP processing may preferentially occur in lipid rafts. Here, we show that relatively little wild-type BACE is found in rafts prepared from a human neuroblastoma cell line (SH-SY5Y) by using Triton X-100 as detergent. To investigate further the significance of lipid rafts in APP processing, a glycosylphosphatidylinositol (GPI) anchor has been added to BACE, replacing the transmembrane and C-terminal domains. The GPI anchor targets the enzyme exclusively to lipid raft domains. Expression of GPI-BACE substantially up-regulates the secretion of both sAPPβ and amyloid-β peptide over levels observed from cells overexpressing wild-type BACE. This effect was reversed when the lipid rafts were disrupted by depleting cellular cholesterol levels. These results suggest that processing of APP to the amyloid-β peptide occurs predominantly in lipid rafts and that BACE is the rate-limiting enzyme in this process. The processing of the APP695isoform by GPI-BACE was up-regulated 20-fold compared with wild-type BACE, whereas only a 2-fold increase in the processing of APP751/770was seen, implying a differential compartmentation of the APP isoforms. 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Indirect evidence has suggested that amyloidogenic APP processing may preferentially occur in lipid rafts. Here, we show that relatively little wild-type BACE is found in rafts prepared from a human neuroblastoma cell line (SH-SY5Y) by using Triton X-100 as detergent. To investigate further the significance of lipid rafts in APP processing, a glycosylphosphatidylinositol (GPI) anchor has been added to BACE, replacing the transmembrane and C-terminal domains. The GPI anchor targets the enzyme exclusively to lipid raft domains. Expression of GPI-BACE substantially up-regulates the secretion of both sAPPβ and amyloid-β peptide over levels observed from cells overexpressing wild-type BACE. This effect was reversed when the lipid rafts were disrupted by depleting cellular cholesterol levels. These results suggest that processing of APP to the amyloid-β peptide occurs predominantly in lipid rafts and that BACE is the rate-limiting enzyme in this process. The processing of the APP695isoform by GPI-BACE was up-regulated 20-fold compared with wild-type BACE, whereas only a 2-fold increase in the processing of APP751/770was seen, implying a differential compartmentation of the APP isoforms. Changes in the local membrane environment during aging may facilitate the cosegregation of APP and BACE leading to increased β-amyloid production.</abstract><cop>United States</cop><pub>National Academy of Sciences</pub><pmid>14504402</pmid><doi>10.1073/pnas.1635130100</doi><tpages>6</tpages><oa>free_for_read</oa></addata></record> |
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subjects | Alzheimer's disease Amino Acid Sequence Amyloid beta-Protein Precursor - metabolism Amyloid Precursor Protein Secretases Antibodies Aspartic Acid Endopeptidases Biological Sciences Cell lines Cell membranes Cholesterol Cholesterol - metabolism Cholesterols Endopeptidases - metabolism Enzymes Glycosylphosphatidylinositols - metabolism Humans Hydrolysis Immunoblotting Lipid Metabolism Lipids Molecular Sequence Data Neurology Peptides Protein isoforms Protein Processing, Post-Translational Proteins Rafts Tumor Cells, Cultured |
title | Exclusively Targeting β-Secretase to Lipid Rafts by GPI-Anchor Addition Up-Regulates β-Site Processing of the Amyloid Precursor Protein |
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