Downregulation of A??PP enhances both calcium content of endoplasmic reticulum and acidic stores and the dynamics of store operated calcium channel activity

摘要

The amyloid-?? protein precursor (A??PP) is a type-1 transmembrane protein involved in Alzheimer's disease (AD). It has become increasingly evident that A??PP, its protein-protein interactions, and its proteolytical fragments may affect calcium homeostasis and vice versa. In addition, there is evidence that calcium dysregulation contributes to AD. To study the role of A??PP in calcium homeostasis, we downregulated its expression in SH-SY5Y cells using shRNA (SH-SY5Y/A??PP-) or increased expression of A??PP695 by transfection (SH-SY5Y/A??PP+). The levels of cytosolic Ca2+ after treatment with thapsigargin, monensin, activation of capacitative calcium entry (CCE), and treatment with SKF, a store operated channel (SOCs) inhibitor, were measured by fura-2AM fluorimetry. SH-SY5Y/A??PP+ cells show reduced response to thapsigargin and reduced CCE, although this reduction is not statistically significant. On the other hand, we found that, relative to SH-SY5Y, SH-SY5Y/A??PP-cells show a significant increase in the response to thapsigargin but not in CCE and their SOCs were more susceptible to SKF inhibition. Additionally, downregulation of A??PP resulted in increased response to monensin that induces calcium release from acidic stores. The increase of calcium release from the endoplasmic reticulum and the acidic stores, when A??PP is downregulated, could be attributed to elevated Ca 2+ content or to a dysregulation of Ca2+ transfer through their membranes. These data, along with already existing evidence regarding the role of A??PP in calcium homeostasis and the early occurring structural and functional abnormalities of endosomes, further substantiate the role of A??PP in calcium homeostasis and in AD. ? 2013 - IOS Press and the authors. All rights reserved.