Inhibition of dynamin-dependent endocytosis increases shedding of the amyloid precursor protein ectodomain and reduces generation of amyloid beta protein

• Published in: BMC cell biology 2005-08, Vol.6 (1), p.30-30, Article 30
• Main Authors: Carey, Robyn M, Balcz, Brigitte A, Lopez-Coviella, Ignacio, Slack, Barbara E
• Format: Article
• Language: English
• Subjects: Amyloid beta-Peptides - biosynthesis; Amyloid beta-Protein Precursor - chemistry; Amyloid beta-Protein Precursor - metabolism; Amyloid Precursor Protein Secretases; Aspartic Acid Endopeptidases; Cell Line; Dynamin I - antagonists & inhibitors; Dynamin I - genetics; Dynamin I - physiology; Endocytosis - physiology; Endopeptidases - metabolism; Humans; Mutation; Protein Kinase C - metabolism; Protein Processing, Post-Translational; Protein Structure, Tertiary; Transfection
• ISSN: 1471-2121; 1471-2121
• DOI: 10.1186/1471-2121-6-30

The amyloid precursor protein (APP) is transported via the secretory pathway to the cell surface, where it may be cleaved within its ectodomain by alpha-secretase, or internalized within clathrin-coated vesicles. An alternative proteolytic pathway occurs within the endocytic compartment, where the sequential action of beta- and gamma-secretases generates the amyloid beta protein (Abeta). In this study, we investigated the effects of modulators of endocytosis on APP processing. Human embryonic kidney cells were transfected with a dominant negative mutant of dynamin I, an important mediator of clathrin-dependent endocytosis, and APP proteolysis was analyzed. Overexpression of the mutant dynamin (dyn I K44A) resulted in increased shedding of the APP ectodomain (sAPPalpha), accumulation of the C-terminal alpha-secretase product C83, and a reduction in the release of Abeta. Levels of mature APP on the cell surface were increased in cells expressing dyn I K44A, and internalization of surface-immunolabeled APP, assessed by fluorescence microscopy, was inhibited. Dynamin is a substrate for protein kinase C (PKC), and it was hypothesized that activators of PKC, which are known to stimulate alpha-secretase-mediated cleavage of APP, might exert their effects by inhibiting dynamin-dependent endocytosis. However, the internalization of surface-biotinylated APP was unaffected by treatment of cells with phorbol 12-myristate 13-acetate in the presence of the alpha-secretase inhibitor TAPI-1. The results indicate that APP is internalized by a dynamin-dependent process, and suggest that alterations in the activity of proteins that mediate endocytosis might lead to significant changes in Abeta production.