Protein kinase C-dependent alpha-secretase competes with beta-secretase for cleavage of amyloid-beta precursor protein in the trans-golgi network

The release of amyloidogenic amyloid-beta peptide (Abeta) from amyloid-beta precursor protein (APP) requires cleavage by beta- and gamma-secretases. In contrast, alpha-secretase cleaves APP within the Abeta sequence and precludes amyloidogenesis. Regulated and unregulated alpha-secretase activities have been reported, and the fraction of cellular alpha-secretase activity regulated by protein kinase C (PKC) has been attributed to the ADAM (a disintegrin and metalloprotease) family members TACE and ADAM-10. Although unregulated alpha-secretase cleavage of APP has been shown to occur at the cell surface, we sought to identify the intracellular site of PKC-regulated alpha-secretase APP cleavage. To accomplish this, we measured levels of secreted ectodomains and C-terminal fragments of APP generated by alpha-secretase (sAPPalpha) (C83) versus beta-secretase (sAPPbeta) (C99) and secreted Abeta in cultured cells treated with PKC and inhibitors of TACE/ADAM-10. We found that PKC stimulation increased sAPPalpha but decreased sAPPbeta levels by altering the competition between alpha- versus beta-secretase for APP within the same organelle rather than by perturbing APP trafficking. Moreover, data implicating the trans-Golgi network (TGN) as a major site for beta-secretase activity prompted us to hypothesize that PKC-regulated alpha-secretase(s) also reside in this organelle. To test this hypothesis, we performed studies demonstrating proteolytically mature TACE intracellularly, and we also showed that regulated alpha-secretase APP cleavage occurs in the TGN using an APP mutant construct targeted specifically to the TGN. By detecting regulated alpha-secretase APP cleavage in the TGN by TACE/ADAM-10, we reveal ADAM activity in a novel location. Finally, the competition between TACE/ADAM-10 and beta-secretase for intracellular APP cleavage may represent a novel target for the discovery of new therapeutic agents to treat Alzheimer's disease.