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A Bioengineering Strategy to Control ADAM10 Activity in Living Cells

A Disintegrin and Metalloprotease 10, also known as ADAM10, is a cell surface protease ubiquitously expressed in mammalian cells where it cuts several membrane proteins implicated in multiple physiological processes. The dysregulation of ADAM10 expression and function has been implicated in patholog...

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Autores principales: Pastore, Francesco, Battistoni, Martina, Sollazzo, Raimondo, Renna, Pietro, Paciello, Fabiola, Li Puma, Domenica Donatella, Barone, Eugenio, Dagliyan, Onur, Ripoli, Cristian, Grassi, Claudio
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9863580/
https://www.ncbi.nlm.nih.gov/pubmed/36674432
http://dx.doi.org/10.3390/ijms24020917
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author Pastore, Francesco
Battistoni, Martina
Sollazzo, Raimondo
Renna, Pietro
Paciello, Fabiola
Li Puma, Domenica Donatella
Barone, Eugenio
Dagliyan, Onur
Ripoli, Cristian
Grassi, Claudio
author_facet Pastore, Francesco
Battistoni, Martina
Sollazzo, Raimondo
Renna, Pietro
Paciello, Fabiola
Li Puma, Domenica Donatella
Barone, Eugenio
Dagliyan, Onur
Ripoli, Cristian
Grassi, Claudio
author_sort Pastore, Francesco
collection PubMed
description A Disintegrin and Metalloprotease 10, also known as ADAM10, is a cell surface protease ubiquitously expressed in mammalian cells where it cuts several membrane proteins implicated in multiple physiological processes. The dysregulation of ADAM10 expression and function has been implicated in pathological conditions, including Alzheimer’s disease (AD). Although it has been suggested that ADAM10 is expressed as a zymogen and the removal of the prodomain results in its activation, other potential mechanisms for the ADAM10 proteolytic function and activation remain unclear. Another suggested mechanism is post-translational modification of the cytoplasmic domain, which regulates ADAM10-dependent protein ectodomain shedding. Therefore, the precise and temporal activation of ADAM10 is highly desirable to reveal the fine details of ADAM10-mediated cleavage mechanisms and protease-dependent therapeutic applications. Here, we present a strategy to control prodomain and cytosolic tail cleavage to regulate ADAM10 shedding activity without the intervention of small endogenous molecule signaling pathways. We generated a series of engineered ADAM10 analogs containing Tobacco Etch Virus protease (TEV) cleavage site (TEVcs), rendering ADAM10 cleavable by TEV. This strategy revealed that, in the absence of other stimuli, the TEV-mediated removal of the prodomain could not activate ADAM10. However, the TEV-mediated cleavage of the cytosolic domain significantly increased ADAM10 activity. Then, we generated ADAM10 with a minimal constitutively catalytic activity that increased significantly in the presence of TEV or after activating a chemically activatable TEV. Our results revealed a bioengineering strategy for controlling the ADAM10 activity in living cells, paving the way to obtain spatiotemporal control of ADAM10. Finally, we proved that our approach of controlling ADAM10 promoted α-secretase activity and the non-amyloidogenic cleavage of amyloid-β precursor protein (APP), thereby increasing the production of the neuroprotective soluble ectodomain (sAPPα). Our bioengineering strategy has the potential to be exploited as a next-generation gene therapy for AD.
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spelling pubmed-98635802023-01-22 A Bioengineering Strategy to Control ADAM10 Activity in Living Cells Pastore, Francesco Battistoni, Martina Sollazzo, Raimondo Renna, Pietro Paciello, Fabiola Li Puma, Domenica Donatella Barone, Eugenio Dagliyan, Onur Ripoli, Cristian Grassi, Claudio Int J Mol Sci Article A Disintegrin and Metalloprotease 10, also known as ADAM10, is a cell surface protease ubiquitously expressed in mammalian cells where it cuts several membrane proteins implicated in multiple physiological processes. The dysregulation of ADAM10 expression and function has been implicated in pathological conditions, including Alzheimer’s disease (AD). Although it has been suggested that ADAM10 is expressed as a zymogen and the removal of the prodomain results in its activation, other potential mechanisms for the ADAM10 proteolytic function and activation remain unclear. Another suggested mechanism is post-translational modification of the cytoplasmic domain, which regulates ADAM10-dependent protein ectodomain shedding. Therefore, the precise and temporal activation of ADAM10 is highly desirable to reveal the fine details of ADAM10-mediated cleavage mechanisms and protease-dependent therapeutic applications. Here, we present a strategy to control prodomain and cytosolic tail cleavage to regulate ADAM10 shedding activity without the intervention of small endogenous molecule signaling pathways. We generated a series of engineered ADAM10 analogs containing Tobacco Etch Virus protease (TEV) cleavage site (TEVcs), rendering ADAM10 cleavable by TEV. This strategy revealed that, in the absence of other stimuli, the TEV-mediated removal of the prodomain could not activate ADAM10. However, the TEV-mediated cleavage of the cytosolic domain significantly increased ADAM10 activity. Then, we generated ADAM10 with a minimal constitutively catalytic activity that increased significantly in the presence of TEV or after activating a chemically activatable TEV. Our results revealed a bioengineering strategy for controlling the ADAM10 activity in living cells, paving the way to obtain spatiotemporal control of ADAM10. Finally, we proved that our approach of controlling ADAM10 promoted α-secretase activity and the non-amyloidogenic cleavage of amyloid-β precursor protein (APP), thereby increasing the production of the neuroprotective soluble ectodomain (sAPPα). Our bioengineering strategy has the potential to be exploited as a next-generation gene therapy for AD. MDPI 2023-01-04 /pmc/articles/PMC9863580/ /pubmed/36674432 http://dx.doi.org/10.3390/ijms24020917 Text en © 2023 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Pastore, Francesco
Battistoni, Martina
Sollazzo, Raimondo
Renna, Pietro
Paciello, Fabiola
Li Puma, Domenica Donatella
Barone, Eugenio
Dagliyan, Onur
Ripoli, Cristian
Grassi, Claudio
A Bioengineering Strategy to Control ADAM10 Activity in Living Cells
title A Bioengineering Strategy to Control ADAM10 Activity in Living Cells
title_full A Bioengineering Strategy to Control ADAM10 Activity in Living Cells
title_fullStr A Bioengineering Strategy to Control ADAM10 Activity in Living Cells
title_full_unstemmed A Bioengineering Strategy to Control ADAM10 Activity in Living Cells
title_short A Bioengineering Strategy to Control ADAM10 Activity in Living Cells
title_sort bioengineering strategy to control adam10 activity in living cells
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9863580/
https://www.ncbi.nlm.nih.gov/pubmed/36674432
http://dx.doi.org/10.3390/ijms24020917
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