Untersuchungen zur intrazellulären Lokalisation der γ-Sekretase

Alzheimer´s disease is characterized by brain deposition of extracellular amyloid β-peptide containing plaques. The cellular site of γ-secretase activity, which releases amyloid β-peptide and the corresponding APP intracellular domain (AICD) remains controversial. Proposed cleavage sites range from the endoplasmic reticulum, the Golgi apparatus, the cell surface to endosomal compartments. We now used C99-GFP, a fluorescent reporter substrate for γ-secretase activity and monitored AICD production in living cells. C99-GFP is efficiently cleaved by γ-secretase and AICD-GFP is released into the cytosol. Inhibiting γ-secretase results in accumulation of C99-GFP in early endosomes. By blocking selective transport steps along the secretory pathway we demonstrate that γ-secretase does not cleave its substrates in the endoplasmic reticulum, the Golgi/trans-Golgi network or in secretory vesicles. In contrast, inhibition of endocytosis did not inhibit cleavage of C99-GFP. Similar results were obtained for another γ-secretase substrate, NotchΔE. Our results suggest that intracellular domains are generated by γ-secretase at the plasma membrane and/or early endosomes