A cell-permeable tool for analysing APP intracellular domain function and manipulation of PIKfyve activity


The mechanisms for regulating PIKfyve complex activity are currently emerging. The PIKfyve complex, consisting of the phosphoinositide kinase PIKfyve (also known as FAB1), VAC14 and FIG4, is required for the production of phosphatidylinositol-3,5-bisphosphate (PI(3,5)P2). PIKfyve function is required for homeostasis of the endo/lysosomal system and is crucially implicated in neuronal function and integrity, as loss of function mutations in the PIKfyve complex lead to neurodegeneration in mouse models and human patients. Our recent work has shown that the intracellular domain of the Amyloid Precursor Protein (APP), a molecule central to the aetiology of Alzheimer's disease binds to VAC14 and enhances PIKfyve function. Here we utilise this recent advance to create an easy-to-use tool for increasing PIKfyve activity in cells. We fused APP's intracellular domain (AICD) to the HIV TAT domain, a cell permeable peptide allowing proteins to penetrate cells. The resultant TAT-AICD fusion protein is cell permeable and triggers an increase of PI(3,5)P2. Using the PI(3,5)P2 specific GFP-ML1Nx2 probe we show that cell-permeable AICD alters PI(3,5)P2 dynamics. TAT-AICD also provides partial protection from pharmacological inhibition of PIKfyve. All three lines of evidence show that the APP intracellular domain activates the PIKfyve complex in cells, a finding that is important for our understanding of the mechanism of neurodegeneration in Alzheimer's disease.

Publication DOI: https://doi.org/10.1042/BSR20160040
Dataset DOI: https://doi.org/10.17036/f49288df-d8f3-47da-aa64-e4e4e85c8633
Divisions: College of Health & Life Sciences > School of Biosciences
College of Health & Life Sciences
College of Health & Life Sciences > School of Biosciences > Cellular and Molecular Biomedicine
Additional Information: -© 2016 The Author(s). This is an open access article published by Portland Press Limited on behalf of the Biochemical Society and distributed under the Creative Commons Attribution Licence 4.0 (CC BY). Funding: Alzheimer’s Research UK (ARUK-PhD-2012/13); and BBSRC (BBK014862/1).
Uncontrolled Keywords: Alzheimer's disease,FAB1,FIG4,Neurodegeneration,Phosphoinositide,VAC14,Vacuolar H-ATPase (V-ATPase),Biochemistry,Biophysics,Cell Biology,Molecular Biology
Publication ISSN: 1573-4935
Last Modified: 22 Apr 2024 07:13
Date Deposited: 08 Mar 2016 16:00
Full Text Link: http://www.bios ... ent/36/2/e00319
Related URLs: http://www.scop ... tnerID=8YFLogxK (Scopus URL)
PURE Output Type: Article
Published Date: 2016-04-15
Published Online Date: 2016-03-02
Accepted Date: 2016-03-02
Authors: Guscott, Benjamin
Balklava, Zita (ORCID Profile 0000-0001-9039-9710)
Safrany, Stephen T.
Wassmer, Thomas



Version: Published Version

License: Creative Commons Attribution


Version: Accepted Version

Access Restriction: Restricted to Repository staff only

License: Creative Commons Attribution

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