title: Gamma-secretase exists on the plasma membrane as an intact complex that accepts substrates and effects intramembrane cleavage.
abstract: Research on Alzheimer's disease led to the identification of a novel proteolytic mechanism in all metazoans, the presenilin/gamma-secretase complex. This unique intramembrane-cleaving aspartyl protease is required for the normal processing of Notch, Jagged, beta-amyloid precursor protein (APP), E-cadherin, and many other receptor-like proteins. We recently provided indirect evidence of gamma-secretase activity at the cell surface in HeLa cells following inhibition of receptor-mediated endocytosis. Here, we directly identify and isolate gamma-secretase as an intact complex (Presenilin, Nicastrin, Aph-1, and Pen-2) from the plasma membrane, both in overexpressing cell lines and endogenously. Inhibition of its proteolytic activity allowed cell surface gamma-secretase to be captured in association with its plasma membrane-localized APP substrates (C83 and C99). Moreover, non-denaturing isolation of the intact enzyme complex revealed that cell surface gamma-secretase can specifically generate amyloid beta-protein from an APP substrate and similarly cleave a Notch substrate. These data directly establish the proteolytic function of gamma-secretase on the plasma membrane, independent of a hypothesized substrate trafficking role. We conclude that presenilin/gamma-secretase exists as a mature complex at the cell surface, where it interacts with and can cleave its substrates, consistent with an essential function in processing many adhesion molecules and receptors required for cell-cell interaction or intercellular signaling.
authors.affiliation: Center for Neurologic Diseases, Brigham & Women's Hospital and Harvard Medical School, Boston, Massachusetts 02115, USA.
authors.forename: Jay H
authors.initials: JH
authors.lastname: Chyung
authors.forename: Daniel M
authors.initials: DM
authors.lastname: Raper
authors.forename: Dennis J
authors.initials: DJ
authors.lastname: Selkoe
mesh_terms: D053829:Amyloid Precursor Protein Secretases; D016564:Amyloid beta-Protein Precursor / Q000737:chemistry; D000818:Animals; D016282:Aspartic Acid Endopeptidases; D019921:Biotinylation; D015153:Blotting, Western; D015820:Cadherins / Q000378:metabolism; D002450:Cell Communication; D002460:Cell Line; D002462:Cell Membrane / Q000201:enzymology* / Q000378:metabolism; D006224:Cricetinae; D004591:Electrophoresis, Polyacrylamide Gel; D004705:Endocytosis; D010450:Endopeptidases / Q000096:biosynthesis*; D004797:Enzyme-Linked Immunosorbent Assay; D006367:HeLa Cells; D006801:Humans; D047468:Immunoprecipitation; D008562:Membrane Glycoproteins / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D010447:Peptide Hydrolases; D053764:Presenilin-1; D053766:Presenilin-2; D011485:Protein Binding; D051880:Receptors, Notch; D015398:Signal Transduction
chemical_list: D016564:Amyloid beta-Protein Precursor; D015820:Cadherins; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; C508853:PSEN1 protein, human; C508861:PSEN2 protein, human; D053764:Presenilin-1; D053766:Presenilin-2; D051880:Receptors, Notch; C413604:nicastrin protein; C475154:APH1A protein, human; D053829:Amyloid Precursor Protein Secretases; D010450:Endopeptidases; D010447:Peptide Hydrolases; D016282:Aspartic Acid Endopeptidases; C509280:BACE1 protein, human