The functional gamma-secretase inhibitor prevents production of amyloid beta 1-34 in human and murine cell lines.
The amyloid precursor protein (APP) undergoes two consecutive cleavages by different proteases, beta-secretase and gamma-secretase, leading to the release of an amyloidogenic 4 kDa fragment called amyloid beta (Abeta). Combining immunoprecipitation and mass spectrometry, we characterized soluble Abeta in cultured cell media of mouse neuroblastoma N2a cells and double hAPP/hBACE-1 transfected HEK293. The major Abeta isoforms detected were Abeta11-34, Abeta1-34, Abeta11-40 and Abeta1-40. In this study, we demonstrate that overexpression of human beta-secretase (BACE-1) in HEK293 cells resulted in predominant Abeta cleavage at position Glu(11) rather than Asp(1), as well as increased production of Abeta(x)-34, but not Abeta(x)-40. Incubation of cells with a specific gamma-secretase inhibitor suggests that cleavage of APP at Leu(34) could be mediated by gamma-secretase itself or by a gamma-secretase dependent process.
Vandermeeren
M
M
CNS Discovery Research, Janssen Research Foundation, Janssen Pharmaceutica, B-2340 Beerse, Belgium.
Geraerts
M
M
Pype
S
S
Dillen
L
L
Van Hove
C
C
Mercken
M
M
D053829:Amyloid Precursor Protein Secretases; D016229:Amyloid beta-Peptides / Q000032:analysis / Q000037:antagonists & inhibitors / Q000096:biosynthesis* / Q000737:chemistry; D016564:Amyloid beta-Protein Precursor / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D000818:Animals; D016282:Aspartic Acid Endopeptidases / Q000235:genetics / Q000494:pharmacology; D002460:Cell Line; D003470:Culture Media / Q000737:chemistry; D010450:Endopeptidases / Q000187:drug effects / Q000502:physiology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D051379:Mice; D010446:Peptide Fragments / Q000032:analysis / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D020033:Protein Isoforms / Q000032:analysis; D012995:Solubility; D014162:Transfection
D016229:Amyloid beta-Peptides; D016564:Amyloid beta-Protein Precursor; D003470:Culture Media; D004791:Enzyme Inhibitors; D010446:Peptide Fragments; D020033:Protein Isoforms; C444321:amyloid beta-protein (1-34); D053829:Amyloid Precursor Protein Secretases; D010450:Endopeptidases; D016282:Aspartic Acid Endopeptidases; C509280:BACE1 protein, human; C509282:Bace1 protein, mouse