Amyloid precursor protein–mediated endocytic pathway disruption induces axonal dysfunction and neurodegeneration
Wei Xu, April M. Weissmiller, Joseph A. White II, Fang Fang, Xinyi Wang, Yiwen Wu, Matthew L. Pearn, Xiaobei Zhao, Mariko Sawa, Shengdi Chen, Shermali Gunawardena, Jianqing Ding, William C. Mobley, Chengbiao Wu
Wei Xu, April M. Weissmiller, Joseph A. White II, Fang Fang, Xinyi Wang, Yiwen Wu, Matthew L. Pearn, Xiaobei Zhao, Mariko Sawa, Shengdi Chen, Shermali Gunawardena, Jianqing Ding, William C. Mobley, Chengbiao Wu
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Research Article Neuroscience

Amyloid precursor protein–mediated endocytic pathway disruption induces axonal dysfunction and neurodegeneration

Abstract

The endosome/lysosome pathway is disrupted early in the course of both Alzheimer’s disease (AD) and Down syndrome (DS); however, it is not clear how dysfunction in this pathway influences the development of these diseases. Herein, we explored the cellular and molecular mechanisms by which endosomal dysfunction contributes to the pathogenesis of AD and DS. We determined that full-length amyloid precursor protein (APP) and its β-C-terminal fragment (β-CTF) act though increased activation of Rab5 to cause enlargement of early endosomes and to disrupt retrograde axonal trafficking of nerve growth factor (NGF) signals. The functional impacts of APP and its various products were investigated in PC12 cells, cultured rat basal forebrain cholinergic neurons (BFCNs), and BFCNs from a mouse model of DS. We found that the full-length wild-type APP (APPWT) and β-CTF both induced endosomal enlargement and disrupted NGF signaling and axonal trafficking. β-CTF alone induced atrophy of BFCNs that was rescued by the dominant-negative Rab5 mutant, Rab5S34N. Moreover, expression of a dominant-negative Rab5 construct markedly reduced APP-induced axonal blockage in Drosophila. Therefore, increased APP and/or β-CTF impact the endocytic pathway to disrupt NGF trafficking and signaling, resulting in trophic deficits in BFCNs. Our data strongly support the emerging concept that dysregulation of Rab5 activity contributes importantly to early pathogenesis of AD and DS.

Authors

Wei Xu, April M. Weissmiller, Joseph A. White II, Fang Fang, Xinyi Wang, Yiwen Wu, Matthew L. Pearn, Xiaobei Zhao, Mariko Sawa, Shengdi Chen, Shermali Gunawardena, Jianqing Ding, William C. Mobley, Chengbiao Wu

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Figure 5

APP, APP mutants, or C99 induced hyperactivation of Rab5 in PC12M cells.

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APP, APP mutants, or C99 induced hyperactivation of Rab5 in PC12M cells....
PC12M cells were cultured and transfected with the indicated constructs. The levels of GTP-Rab5 were assayed as in Figure 2. The levels of total Rab5 were blotted as loading controls (A). The results were quantitated and are presented in B. The level of GTP-Rab5 in untransfected cells was set at 100%. (C) PC12M cells were treated with either vehicle or 1 μM GSI for 24 hours, and cell lysates were analyzed by SDS-PAGE/immunoblotting with the indicated antibodies. Both full-length APP (fl APP) and APP CTFs are shown. The levels of activated Rab5 and total Rab5 are also shown. (D) PC12M cells were transfected with mCherry-Rab5WT for 24 hours and were treated with either 1 μM GSI or vehicle for another 48 hours. Cells were fixed and analyzed by use of a Leica TCS SPE confocal microscope with a ×63 oil objective lens. Representative images of mCherry-Rab5WT puncta are shown. Enlarged mCherry-Rab5WT puncta by GSI treatment are highlighted in the inset (×1.6; arrowheads). Scale bars: 10 μm. The sizes of mCherry-Rab5WT puncta from 25–30 transfected cells were quantitated, and the size distribution pattern is shown in E. Data represent mean ± SEM of at least 3 independent experiments. All corresponding P values were calculated using Student’s t test.