DETECTION OF THE MEMBRANE-RETAINED CARBOXY-TERMINAL TAIL CONTAINING POLYPEPTIDES OF THE AMYLOID PRECURSOR PROTEIN IN TISSUE FROM ALZHEIMERS-DISEASE BRAIN

A major hallmark of Alzheimer's disease (AD) is the presence of extrac ellular amyloid plaques consisting primarily of amyloid beta peptide ( A beta) which is derived from a larger beta-amyloid precursor protein (APP). APP is processed via secretory and endosomal/lysosomal pathways by a group of proteases called secretases. During the processing of A PP, the carboxy-terminal tail fragment has been suggested to remain wi thin the cell. To investigate the fate of this fragment, we generated an antibody specific for a nine amino acid residue, the sequence of wh ich was derived from the carboxy-terminal putative cytoplasmic tail of APP. Computer analysis of the entire APP gene, searching for regions of greatest antigenicity, surface probability, hydrophilicity, and pre sence of beta turns, indicated that the cytoplasmic tail region is an immunodominant region of APP. The peptide coupled to keyhole limpet he mocyanin protein, produced avery high titer antibody (1:1 x 10(6)). To evaluate the specificity of the antibody, immunoprecipitation of in v itro transcribed and translated DNA encoding the carboxy-terminal amin o acids of APP in wheat germ extract was carried out. A single immunop recipitated band of the correct size was seen by SDS-PAGE. The antibod y was also able to specifically detect the accumulation of the stable C-terminal tail containing fragments of APP in neurites of the amygdal a and hippocampus regions of the human brain tissue from AD subjects, but did not react with age-matched control normal brain tissue. The lo calization of the C-terminal tail of APP within the brain tissue of AD patients underscores the likely importance of the C-terminus in the p athogenesis of AD.