METABOLITES OF THE BETA-AMYLOID PRECURSOR PROTEIN GENERATED BY BETA-SECRETASE LOCALIZE TO THE TRANS-GOLGI NETWORK AND LATE ENDOSOME IN 293 CELLS

Deposition of beta-amyloid occurs in the brains of all sufferers of Al zheimer's disease, beta-amyloid is proteolytically derived from the be ta-amyloid precursor protein by as yet unidentified enzymes termed sec retases, We have generated and characterised antisera to the carboxy-t erminal domain and beta-secretase cleavage site of the Alzheimer's amy loid precursor protein, The beta-secretase cleavage event occurs at th e extreme N-terminus of the beta-amyloid peptide, Our antiserum to the N-terminus of the P-amyloid peptide (NT beta 4) specifically recognis es beta-secretase cleaved species as opposed to intact beta APP, NT be ta 4 specifically immuneprecipitates a 13 kDa fragment of beta APP (p1 3) which is potentially amyloidogenic. We have used these antisera in confocal laser scanning immunofluorescence microscopy to localise the intracellular location of potentially amyloidogenic beta APP processin g fragments such as p13, Using a number of marker antisera of known in tracellular location, we have defined the major location of beta APP f ragments possessing the Asp-1 N-terminus of beta-amyloid as the trans- Golgi network or late endosome on the basis of colocalisation with a m onoclonal antibody to the cation-independent mannose-6-phosphate recep tor, The colocalisation was further investigated using brefeldin A whi ch demonstrated that the p13 fragment and mannose-6-phosphate receptor are trafficked by alternative pathways from the trans-Golgi network. (C) 1996 Wiley-Liss, Inc.