APP functions at the endomembrane system

INTRODUCTION: APP is a type I transmembrane protein, and its intracellular domain (AICD) interacts with FE65, X11, and mDAB1, et al., through the YENPTY motif. Accordingly, it has been proposed that the functions of APP are similar to Notch, which regulates gene expression through releasing NICD after combining with its ligands. However, the detailed underlying mechanisms of how FE65 is regulated by APP, as well as their target genes, remain unknown. METHODS: We overexpressed APP695 in N2a cells, and deleted/mutated APP in H4 cells by CRISPR-Cas9, respectively. The impacts of APP overexpression and ablation were detected by proteome comparison and immunoblotting. Thereafter, the subcellular localizations of APP and its binding protein FE65 were examined by immunofluorescence. RESULTS: APP overexpression not only resulted in its retention in Golgi apparatus (GA) and Endoplasmic reticulum (ER), but also induced FE65 clustering. In addition, it significantly reduced the protein level of ATF4 in N2a cells. On the contrary, knockdown of FE65 restored the protein level of ATF4. More importantly, APP overexpression also impaired autophagy. In contrast, knockout of APP rescued LC3 conversion. Of note, the APP Swedish mutant did not impact ER stress or the autophagic flux. Nevertheless, it facilitated the cleavage of APP-CTFs by gamma-secretase. DISCUSSION: The evidence presented in the current study demonstrates that APP is involved in stabilizing the endomembrane system and the subcellular transport of FE65. More importantly, the role of APP in regulating autophagy indicated that full-length APP could be considered a therapeutic target for preventing the development of AD.