Dataset Information

Increased H-Bond Stability Relates to Altered ε-Cleavage Efficiency and Aβ Levels in the I45T Familial Alzheimer's Disease Mutant of APP.

ABSTRACT: Cleavage of the amyloid precursor protein's (APP) transmembrane domain (TMD) by γ-secretase is a crucial step in the aetiology of Alzheimer's Disease (AD). Mutations in the APP TMD alter cleavage and lead to familial forms of AD (FAD). The majority of FAD mutations shift the preference of initial cleavage from ε49 to ε48, thus raising the AD-related Aβ42/Aβ40 ratio. The I45T mutation is among the few FAD mutations that do not alter ε-site preference, while it dramatically reduces the efficiency of ε-cleavage. Here, we investigate the impact of the I45T mutation on the backbone dynamics of the substrate TMD. Amide exchange experiments and molecular dynamics simulations in solvent and a lipid bilayer reveal an increased stability of amide hydrogen bonds at the ζ- and γ-cleavage sites. Stiffening of the H-bond network is caused by an additional H-bond between the T45 side chain and the TMD backbone, which alters dynamics within the cleavage domain. In particular, the increased H-bond stability inhibits an upward movement of the ε-sites in the I45T mutant. Thus, an altered presentation of ε-sites to the active site of γ-secretase as a consequence of restricted local flexibility provides a rationale for reduced ε-cleavage efficiency of the I45T mutant.

SUBMITTER: Gotz A

PROVIDER: S-EPMC6440955 | biostudies-literature | 2019 Mar

REPOSITORIES: biostudies-literature

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Publications

Increased H-Bond Stability Relates to Altered ε-Cleavage Efficiency and Aβ Levels in the I45T Familial Alzheimer's Disease Mutant of APP.

Götz Alexander A Högel Philipp P Silber Mara M Chaitoglou Iro I Luy Burkhard B Muhle-Goll Claudia C Scharnagl Christina C Langosch Dieter D

Scientific reports 20190329 1

Cleavage of the amyloid precursor protein's (APP) transmembrane domain (TMD) by γ-secretase is a crucial step in the aetiology of Alzheimer's Disease (AD). Mutations in the APP TMD alter cleavage and lead to familial forms of AD (FAD). The majority of FAD mutations shift the preference of initial cleavage from ε49 to ε48, thus raising the AD-related Aβ42/Aβ40 ratio. The I45T mutation is among the few FAD mutations that do not alter ε-site preference, while it dramatically reduces the efficiency ...[more]

PMID: 30926830

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Project description:Genome-wide association studies have identified BIN1 (Bridging integrator 1) and RIN3 (Ras and Rab interactor 3) as genetic risk factors for late-onset Alzheimer's disease (LOAD). The neuronal isoform of BIN1 (BIN1V1), but not the non-neuronal isoform (BIN1V9), has been shown to regulate tau-pathology and Aβ generation via RAB5-mediated endocytosis in neurons. BIN1 directly interacts with RIN3 to initiate RAB5-mediated endocytosis, which is essential for β-secretase (BACE1)-mediated β-secretase cleavage of β-amyloid precursor protein (APP) to generate Amyloid-β (Aβ), the key component of senile plaques in AD. Understanding the regulatory roles of BIN1 (neuronal BIN1V1) and RIN3 in β-secretase mediated cleavage of APP and Aβ generation is key to developing novel therapeutics to delay or prevent AD progression. Neuronal and non-neuronal isoforms of BIN1 (BIN1V1 and BIN1V9, respectively) were introduced with RIN3 into an in vitro cell-based system to test RIN3-dependent effects of neuronal BIN1V1 and non-neuronal BIN1V9 on β-secretase-mediated cleavage of APP and Aβ generation. Confocal microscopy was performed to examine RIN3-dependent subcellular localization of BIN1V1 and BIN1V9. Western blot analysis was performed to assess the effects of RIN3 and BIN1V1/BIN1V9 on β-secretase mediated processing of APP. We enriched cells expressing BIN1V1 without or with RIN3 via FACS to measure Aβ generation using Aβ ELISA assay, and to evaluate APP internalization by chasing biotinylated or antibody-labeled cell surface APP. Neuronal BIN1V1 containing the CLAP domain and non-neuronal BIN1V9 lacking the CLAP domain are the major isoforms present in the brain. Employing confocal microscopy, we showed that RIN3 differentially regulates the recruitment of both BIN1V1 and BIN1V9 into RAB5-endosomes. We further showed that BIN1V1, but not BIN1V9, downregulates β-secretase (BACE1)-mediated processing of APP in a RIN3-dependent manner. Overexpression of BIN1V1 also attenuated Aβ generation in a RIN3-dependent manner. Using cell-based internalization assays, we show BIN1V1, but not BIN1V9, delays the endocytosis of APP, but not of BACE1, into early endosomes, thereby spatially and temporally separating these two proteins into different cellular compartments, resulting in reduced cleavage of APP by BACE1 and reduced Aβ generation-all in a RIN3-dependent manner. Finally, we show that RIN3 sequesters BIN1V1 in RAB5-positive early endosomes, likely via the CLAP-domain, resulting in attenuated β-secretase processing of APP and Aβ generation by delaying endocytosis of APP. Our findings provide new mechanistic data on how two AD-associated molecules, RIN3 and BIN1 (neuronal BIN1V1), interact to govern Aβ production, implicating these two proteins as potential therapeutic targets for the prevention and treatment of AD.

Dataset Information

Increased H-Bond Stability Relates to Altered ε-Cleavage Efficiency and Aβ Levels in the I45T Familial Alzheimer's Disease Mutant of APP.

Publications

Increased H-Bond Stability Relates to Altered ε-Cleavage Efficiency and Aβ Levels in the I45T Familial Alzheimer's Disease Mutant of APP.

Similar Datasets

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