Targeting γ-secretase triggers the selective enrichment of oligomeric APP-CTFs in brain extracellular vesicles from Alzheimer cell and mouse models
Abstract Background We recently demonstrated an endolysosomal accumulation of the β-secretase-derived APP C-terminal fragment (CTF) C99 in brains of Alzheimer disease (AD) mouse models. Moreover, we showed that the treatment with the γ-secretase inhibitor (D6) led to further increased endolysosomal...
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2019-12-01
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Series: | Translational Neurodegeneration |
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Online Access: | https://doi.org/10.1186/s40035-019-0176-6 |
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Article |
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English |
format |
Article |
sources |
DOAJ |
author |
Inger Lauritzen Anaïs Bécot Alexandre Bourgeois Raphaëlle Pardossi-Piquard Maria-Grazia Biferi Martine Barkats Fréderic Checler |
spellingShingle |
Inger Lauritzen Anaïs Bécot Alexandre Bourgeois Raphaëlle Pardossi-Piquard Maria-Grazia Biferi Martine Barkats Fréderic Checler Targeting γ-secretase triggers the selective enrichment of oligomeric APP-CTFs in brain extracellular vesicles from Alzheimer cell and mouse models Translational Neurodegeneration Extracellular vesicles C99 APP-CTFs Homo- and hetero-oligomerization Endosomes Lysosomes |
author_facet |
Inger Lauritzen Anaïs Bécot Alexandre Bourgeois Raphaëlle Pardossi-Piquard Maria-Grazia Biferi Martine Barkats Fréderic Checler |
author_sort |
Inger Lauritzen |
title |
Targeting γ-secretase triggers the selective enrichment of oligomeric APP-CTFs in brain extracellular vesicles from Alzheimer cell and mouse models |
title_short |
Targeting γ-secretase triggers the selective enrichment of oligomeric APP-CTFs in brain extracellular vesicles from Alzheimer cell and mouse models |
title_full |
Targeting γ-secretase triggers the selective enrichment of oligomeric APP-CTFs in brain extracellular vesicles from Alzheimer cell and mouse models |
title_fullStr |
Targeting γ-secretase triggers the selective enrichment of oligomeric APP-CTFs in brain extracellular vesicles from Alzheimer cell and mouse models |
title_full_unstemmed |
Targeting γ-secretase triggers the selective enrichment of oligomeric APP-CTFs in brain extracellular vesicles from Alzheimer cell and mouse models |
title_sort |
targeting γ-secretase triggers the selective enrichment of oligomeric app-ctfs in brain extracellular vesicles from alzheimer cell and mouse models |
publisher |
BMC |
series |
Translational Neurodegeneration |
issn |
2047-9158 |
publishDate |
2019-12-01 |
description |
Abstract Background We recently demonstrated an endolysosomal accumulation of the β-secretase-derived APP C-terminal fragment (CTF) C99 in brains of Alzheimer disease (AD) mouse models. Moreover, we showed that the treatment with the γ-secretase inhibitor (D6) led to further increased endolysosomal APP-CTF levels, but also revealed extracellular APP-CTF-associated immunostaining. We here hypothesized that this latter staining could reflect extracellular vesicle (EV)-associated APP-CTFs and aimed to characterize these γ-secretase inhibitor-induced APP-CTFs. Methods EVs were purified from cell media or mouse brains from vehicle- or D6-treated C99 or APPswedish expressing cells/mice and analyzed for APP-CTFs by immunoblot. Combined pharmacological, immunological and genetic approaches (presenilin invalidation and C99 dimerization mutants (GXXXG)) were used to characterize vesicle-containing APP-CTFs. Subcellular APP-CTF localization was determined by immunocytochemistry. Results Purified EVs from both AD cell or mouse models were enriched in APP-CTFs as compared to EVs from control cells/brains. Surprisingly, EVs from D6-treated cells not only displayed increased C99 and C99-derived C83 levels but also higher molecular weight (HMW) APP-CTF-immunoreactivities that were hardly detectable in whole cell extracts. Accordingly, the intracellular levels of HMW APP-CTFs were amplified by the exosomal inhibitor GW4869. By combined pharmacological, immunological and genetic approaches, we established that these HMW APP-CTFs correspond to oligomeric APP-CTFs composed of C99 and/or C83. Immunocytochemical analysis showed that monomers were localized mainly to the trans-Golgi network, whereas oligomers were confined to endosomes and lysosomes, thus providing an anatomical support for the selective recovery of HMW APP-CTFs in EVs. The D6-induced APP-CTF oligomerization and subcellular mislocalization was indeed due to γ-secretase blockade, since it similarly occurred in presenilin-deficient fibroblasts. Further, our data proposed that besides favoring APP-CTF oligomerization by preventing C99 proteolysis, γ-secretase inhibiton also led to a defective SorLA-mediated retrograde transport of HMW APP-CTFs from endosomal compartments to the TGN. Conclusions This is the first study to demonstrate the presence of oligomeric APP-CTFs in AD mouse models, the levels of which are selectively enriched in endolysosomal compartments including exosomes and amplified by γ-secretase inhibition. Future studies should evaluate the putative contribution of these exosome-associated APP-CTFs in AD onset, progression and spreading. |
topic |
Extracellular vesicles C99 APP-CTFs Homo- and hetero-oligomerization Endosomes Lysosomes |
url |
https://doi.org/10.1186/s40035-019-0176-6 |
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doaj-027cf11112fc4c188a93980afe9655952020-12-06T12:55:17ZengBMCTranslational Neurodegeneration2047-91582019-12-018111710.1186/s40035-019-0176-6Targeting γ-secretase triggers the selective enrichment of oligomeric APP-CTFs in brain extracellular vesicles from Alzheimer cell and mouse modelsInger Lauritzen0Anaïs Bécot1Alexandre Bourgeois2Raphaëlle Pardossi-Piquard3Maria-Grazia Biferi4Martine Barkats5Fréderic Checler6Institut de Pharmacologie Moléculaire et Cellulaire, CNRS-UMR7275, team labeled «Fondation pour la Recherche Médicale» et «Laboratoire d’excellence Distalz», Université de Nice-Sophia-AntipolisInstitut de Pharmacologie Moléculaire et Cellulaire, CNRS-UMR7275, team labeled «Fondation pour la Recherche Médicale» et «Laboratoire d’excellence Distalz», Université de Nice-Sophia-AntipolisInstitut de Pharmacologie Moléculaire et Cellulaire, CNRS-UMR7275, team labeled «Fondation pour la Recherche Médicale» et «Laboratoire d’excellence Distalz», Université de Nice-Sophia-AntipolisInstitut de Pharmacologie Moléculaire et Cellulaire, CNRS-UMR7275, team labeled «Fondation pour la Recherche Médicale» et «Laboratoire d’excellence Distalz», Université de Nice-Sophia-AntipolisInstitut-MyologieInstitut-MyologieInstitut de Pharmacologie Moléculaire et Cellulaire, CNRS-UMR7275, team labeled «Fondation pour la Recherche Médicale» et «Laboratoire d’excellence Distalz», Université de Nice-Sophia-AntipolisAbstract Background We recently demonstrated an endolysosomal accumulation of the β-secretase-derived APP C-terminal fragment (CTF) C99 in brains of Alzheimer disease (AD) mouse models. Moreover, we showed that the treatment with the γ-secretase inhibitor (D6) led to further increased endolysosomal APP-CTF levels, but also revealed extracellular APP-CTF-associated immunostaining. We here hypothesized that this latter staining could reflect extracellular vesicle (EV)-associated APP-CTFs and aimed to characterize these γ-secretase inhibitor-induced APP-CTFs. Methods EVs were purified from cell media or mouse brains from vehicle- or D6-treated C99 or APPswedish expressing cells/mice and analyzed for APP-CTFs by immunoblot. Combined pharmacological, immunological and genetic approaches (presenilin invalidation and C99 dimerization mutants (GXXXG)) were used to characterize vesicle-containing APP-CTFs. Subcellular APP-CTF localization was determined by immunocytochemistry. Results Purified EVs from both AD cell or mouse models were enriched in APP-CTFs as compared to EVs from control cells/brains. Surprisingly, EVs from D6-treated cells not only displayed increased C99 and C99-derived C83 levels but also higher molecular weight (HMW) APP-CTF-immunoreactivities that were hardly detectable in whole cell extracts. Accordingly, the intracellular levels of HMW APP-CTFs were amplified by the exosomal inhibitor GW4869. By combined pharmacological, immunological and genetic approaches, we established that these HMW APP-CTFs correspond to oligomeric APP-CTFs composed of C99 and/or C83. Immunocytochemical analysis showed that monomers were localized mainly to the trans-Golgi network, whereas oligomers were confined to endosomes and lysosomes, thus providing an anatomical support for the selective recovery of HMW APP-CTFs in EVs. The D6-induced APP-CTF oligomerization and subcellular mislocalization was indeed due to γ-secretase blockade, since it similarly occurred in presenilin-deficient fibroblasts. Further, our data proposed that besides favoring APP-CTF oligomerization by preventing C99 proteolysis, γ-secretase inhibiton also led to a defective SorLA-mediated retrograde transport of HMW APP-CTFs from endosomal compartments to the TGN. Conclusions This is the first study to demonstrate the presence of oligomeric APP-CTFs in AD mouse models, the levels of which are selectively enriched in endolysosomal compartments including exosomes and amplified by γ-secretase inhibition. Future studies should evaluate the putative contribution of these exosome-associated APP-CTFs in AD onset, progression and spreading.https://doi.org/10.1186/s40035-019-0176-6Extracellular vesiclesC99APP-CTFsHomo- and hetero-oligomerizationEndosomesLysosomes |