Amyloid plaques, found in characteristically large numbers in specific brain areas of Alzheimer's disease (AD) and Down's Syndrome (DS) patients, are composed of a 417–43 amino acid peptide, A4, derived from a transmembrane glycoprotein, amyloid precursor protein (APP). In transformed cells APP has been shown to be cleaved within the extracellular portion of the A4 region causing the release of 100–120 kDa soluble N‐terminal APP products. If this cleavage occurs in human tissue, neither the soluble product nor the remaining 10–12 kDa transmembrane fragment could be further degraded to yield A4. It has been hypothesized that an alternate APP cleavage product containing the intact A4 region is released in increased amounts in AD and DS brain where subsequent extracellular degradation produces the amy‐loidogenic A4 peptide. In support of this hypothesis, we have found that PC‐12 cells maintained in serum‐free media with or without additional injurious agents release a 60 kDa protein which has been detected by immunoprecipitation and immunoblot analyses with 9 antisera elicited by 4 distinct peptides within the carboxyl‐terminal half of APP. Six of these antisera, elicited by peptides corresponding to the carboxyl‐terminal 20 amino acids of APP, or the A4 peptide itself, do not bind the normally released 120 kDa APP product which is detected by 11 other antisera elicited by peptides with the N‐terminal portion of APP. Controls in which two 60 kDa‐detecting antisera were preabsorbed with the peptides used to elicit them, produced markedly reduced 60 kd bands on immunoblots. Thus, the 60 kDa protein appears to be the first reported APP product released from neural cells which, containing the intact A4 region, is potentially amyloidogenic. Similar analyses of proteins released into the serum‐free media of PC‐12 cells maintained in 0.0030.005% SDS, 60 mM chloroquine, or other membrane damaging or lysosomal inhibiting agents revealed increased 60 kDa band intensities. Cultures grown with other classes of injurious agents did not demonstrate increased release. Together these data indicate that neural cells maintained under selected stressful or injurious conditions release increased quantities of a potentially amyloidogenic APP product, supporting the importance of a similar induction of alternative APP cleavage in AD and DS brain amy‐loidogenesis.
- protein cleavage
ASJC Scopus subject areas
- Cellular and Molecular Neuroscience