Inhibition of endocytosis activates alternative degradation pathway of βAPP in cultured cells.

Background: Alzheimer’s disease associated βAPP is sequentially endoproteolyzed by α/β-secretase and γ-cleavage. In the process, extracellular shedding by α-secretase (ADAM 9/10/17) or β-secretase (BACE 1/2) at position L17 or D1 (Aβ numbering) are prerequisites for the generation of P3 or Aβ, respectively. In addition, several alternative extracellular cleavage sites in βAPP have been reported at position I−6, V−3, R5, E11, F20, and A21. Among these sites, position R5 is considered to be cleaved by α-secretase-like activity, whereas position E11, F20 and A21 are cleaved by β-secretase. Therefore, extracellular shedding of βAPP is thought to be mediated exclusively by α/β-secretase activities. However, so far the characteristics of cleavages at position V−3 and I−6 are not well understood. The aim of this study is to characterize these two cleavages of βAPP. Methods: We analyzed the conditioned media of βAPP wt or sw expressing cells with or without pharmacological agents. Results: Here, we show that the cleavage at position I−6 of βAPP has characteristics distinct from that of α/β-secretase, while the cleavage at V−3 seems to be mediated by β-secretase. Although inhibition of endocytosis enhances the cleavages at both V−3 and I−6, PMA, an α-secretase stimulator, treatment enhances neither of these cleavages. Interestingly, a β-secretase inhibitor, z-VLL-CHO, suppressed V−3 but not I−6 cleavage. The pathological βAPP Swedish mutant adjacent to the cleavage sites shows similar effects. Conclusions: Our data demonstrate that neither α nor β-secretase undergoes extracellular shedding at I−6 of βAPP. Therefore, our data may indicate a novel alternative βAPP degradation pathway which is up-regulated upon low level of endocytosis. [ABSTRACT FROM AUTHOR]