title: Characterization of presenilin complexes from mouse and human brain using Blue Native gel electrophoresis reveals high expression in embryonic brain and minimal change in complex mobility with pathogenic presenilin mutations.
abstract: The presenilin proteins are required for intramembrane cleavage of a subset of type 1 membrane proteins including the Alzheimer's disease amyloid precursor protein. Previous studies indicate presenilin proteins form enzymatically active high molecular mass complexes consisting of heterodimers of N- and C-terminal fragments in association with nicastrin, presenilin enhancer-2 and anterior pharynx defective-1 proteins. Using Blue Native gel electrophoresis (BN/PAGE) we have studied endogenous presenilin 1 complex mass, stability and association with nicastrin, presenilin enhancer-2 and anterior pharynx defective-1. Solubilization of mouse or human brain membranes with dodecyl-d-maltoside produced a 360-kDa species reactive with antibodies to presenilin 1. Presenilin 1 complex levels were high in embryonic brain. Complex integrity was sensitive to Triton X-100 and SDS, but stable to reducing agent. Addition of 5 M urea caused complex dissolution and nicastrin to migrate as a subcomplex. Nicastrin and presenilin enhancer-2 were detected in the presenilin 1 complex following BN/PAGE, electroelution and second-dimension analysis. Anterior pharynx defective-1 was detected as an 18-kDa form and 9-kDa C-terminal fragment by standard SDS/PAGE of mouse tissues, and as a predominant 36-kDa band after presenilin 1 complex second-dimension analysis. Membranes from brain cortex of Alzheimer's disease patients, or from cases with presenilin 1 missense mutations, indicated no change in presenilin 1 complex mobility. Higher molecular mass presenilin 1-reactive species were detected in brain containing presenilin 1 exon 9 deletion mutation. This abnormality was confirmed using cells transfected with the same presenilin deletion mutation.
authors.affiliation: Department of Pathology, The University of Melbourne, Australia. janetta@unimelb.edu.au
authors.forename: Janetta G
authors.initials: JG
authors.lastname: Culvenor
authors.forename: Nancy T
authors.initials: NT
authors.lastname: Ilaya
authors.forename: Michael T
authors.initials: MT
authors.lastname: Ryan
authors.forename: Louise
authors.initials: L
authors.lastname: Canterford
authors.forename: David E
authors.initials: DE
authors.lastname: Hoke
authors.forename: Nicholas A
authors.initials: NA
authors.lastname: Williamson
authors.forename: Catriona A
authors.initials: CA
authors.lastname: McLean
authors.forename: Colin L
authors.initials: CL
authors.lastname: Masters
authors.forename: Geneviève
authors.initials: G
authors.lastname: Evin
mesh_terms: D000544:Alzheimer Disease / Q000378:metabolism* / Q000473:pathology; D053829:Amyloid Precursor Protein Secretases; D016564:Amyloid beta-Protein Precursor / Q000378:metabolism; D000818:Animals; D016282:Aspartic Acid Endopeptidases; D001921:Brain / Q000378:metabolism*; D002462:Cell Membrane; D002478:Cells, Cultured; D003598:Cytoskeletal Proteins / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases / Q000378:metabolism; D005091:Exons / Q000235:genetics; D006801:Humans; D008562:Membrane Glycoproteins / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics* / Q000378:metabolism*; D051379:Mice; D020125:Mutation, Missense; D010447:Peptide Hydrolases; D053764:Presenilin-1; D017384:Sequence Deletion; D015534:Trans-Activators / Q000378:metabolism; D051176:beta Catenin
chemical_list: D016564:Amyloid beta-Protein Precursor; C495270:CTNNB1 protein, human; C495267:CTNNB1 protein, mouse; D003598:Cytoskeletal Proteins; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; C508853:PSEN1 protein, human; C469349:PSENEN protein, human; D053764:Presenilin-1; D015534:Trans-Activators; D051176:beta Catenin; 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; C509282:Bace1 protein, mouse