Proteomic analysis of necroptotic extracellular vesicles

Abstract Necroptosis is a regulated and inflammatory form of cell death. We, and others, have previously reported that necroptotic cells release extracellular vesicles (EVs). We have found that necroptotic EVs are loaded with proteins, including the phosphorylated form of the key necroptosis-executi...

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Autores principales:	Inbar Shlomovitz, Ziv Erlich, Gali Arad, Liat Edry-Botzer, Sefi Zargarian, Hadar Cohen, Tal Manko, Yifat Ofir-Birin, Tomer Cooks, Neta Regev-Rudzki, Motti Gerlic
Formato:	article
Lenguaje:	EN
Publicado:	Nature Publishing Group 2021
Materias:	Cytology QH573-671
Acceso en línea:	https://doaj.org/article/7b5dc9c19a2c4f41a176d2a1c9cf881e
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spelling	oai:doaj.org-article:7b5dc9c19a2c4f41a176d2a1c9cf881e2021-11-14T12:06:56ZProteomic analysis of necroptotic extracellular vesicles10.1038/s41419-021-04317-z2041-4889https://doaj.org/article/7b5dc9c19a2c4f41a176d2a1c9cf881e2021-11-01T00:00:00Zhttps://doi.org/10.1038/s41419-021-04317-zhttps://doaj.org/toc/2041-4889Abstract Necroptosis is a regulated and inflammatory form of cell death. We, and others, have previously reported that necroptotic cells release extracellular vesicles (EVs). We have found that necroptotic EVs are loaded with proteins, including the phosphorylated form of the key necroptosis-executing factor, mixed lineage kinase domain-like kinase (MLKL). However, neither the exact protein composition, nor the impact, of necroptotic EVs have been delineated. To characterize their content, EVs from necroptotic and untreated U937 cells were isolated and analyzed by mass spectrometry-based proteomics. A total of 3337 proteins were identified, sharing a high degree of similarity with exosome proteome databases, and clearly distinguishing necroptotic and control EVs. A total of 352 proteins were significantly upregulated in the necroptotic EVs. Among these were MLKL and caspase-8, as validated by immunoblot. Components of the ESCRTIII machinery and inflammatory signaling were also upregulated in the necroptotic EVs, as well as currently unreported components of vesicle formation and transport, and necroptotic signaling pathways. Moreover, we found that necroptotic EVs can be phagocytosed by macrophages to modulate cytokine and chemokine secretion. Finally, we uncovered that necroptotic EVs contain tumor neoantigens, and are enriched with components of antigen processing and presentation. In summary, our study reveals a new layer of regulation during the early stage of necroptosis, mediated by the secretion of specific EVs that influences the microenvironment and may instigate innate and adaptive immune responses. This study sheds light on new potential players in necroptotic signaling and its related EVs, and uncovers the functional tasks accomplished by the cargo of these necroptotic EVs.Inbar ShlomovitzZiv ErlichGali AradLiat Edry-BotzerSefi ZargarianHadar CohenTal MankoYifat Ofir-BirinTomer CooksNeta Regev-RudzkiMotti GerlicNature Publishing GrouparticleCytologyQH573-671ENCell Death and Disease, Vol 12, Iss 11, Pp 1-13 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Cytology QH573-671
spellingShingle	Cytology QH573-671 Inbar Shlomovitz Ziv Erlich Gali Arad Liat Edry-Botzer Sefi Zargarian Hadar Cohen Tal Manko Yifat Ofir-Birin Tomer Cooks Neta Regev-Rudzki Motti Gerlic Proteomic analysis of necroptotic extracellular vesicles
description	Abstract Necroptosis is a regulated and inflammatory form of cell death. We, and others, have previously reported that necroptotic cells release extracellular vesicles (EVs). We have found that necroptotic EVs are loaded with proteins, including the phosphorylated form of the key necroptosis-executing factor, mixed lineage kinase domain-like kinase (MLKL). However, neither the exact protein composition, nor the impact, of necroptotic EVs have been delineated. To characterize their content, EVs from necroptotic and untreated U937 cells were isolated and analyzed by mass spectrometry-based proteomics. A total of 3337 proteins were identified, sharing a high degree of similarity with exosome proteome databases, and clearly distinguishing necroptotic and control EVs. A total of 352 proteins were significantly upregulated in the necroptotic EVs. Among these were MLKL and caspase-8, as validated by immunoblot. Components of the ESCRTIII machinery and inflammatory signaling were also upregulated in the necroptotic EVs, as well as currently unreported components of vesicle formation and transport, and necroptotic signaling pathways. Moreover, we found that necroptotic EVs can be phagocytosed by macrophages to modulate cytokine and chemokine secretion. Finally, we uncovered that necroptotic EVs contain tumor neoantigens, and are enriched with components of antigen processing and presentation. In summary, our study reveals a new layer of regulation during the early stage of necroptosis, mediated by the secretion of specific EVs that influences the microenvironment and may instigate innate and adaptive immune responses. This study sheds light on new potential players in necroptotic signaling and its related EVs, and uncovers the functional tasks accomplished by the cargo of these necroptotic EVs.
format	article
author	Inbar Shlomovitz Ziv Erlich Gali Arad Liat Edry-Botzer Sefi Zargarian Hadar Cohen Tal Manko Yifat Ofir-Birin Tomer Cooks Neta Regev-Rudzki Motti Gerlic
author_facet	Inbar Shlomovitz Ziv Erlich Gali Arad Liat Edry-Botzer Sefi Zargarian Hadar Cohen Tal Manko Yifat Ofir-Birin Tomer Cooks Neta Regev-Rudzki Motti Gerlic
author_sort	Inbar Shlomovitz
title	Proteomic analysis of necroptotic extracellular vesicles
title_short	Proteomic analysis of necroptotic extracellular vesicles
title_full	Proteomic analysis of necroptotic extracellular vesicles
title_fullStr	Proteomic analysis of necroptotic extracellular vesicles
title_full_unstemmed	Proteomic analysis of necroptotic extracellular vesicles
title_sort	proteomic analysis of necroptotic extracellular vesicles
publisher	Nature Publishing Group
publishDate	2021
url	https://doaj.org/article/7b5dc9c19a2c4f41a176d2a1c9cf881e
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Proteomic analysis of necroptotic extracellular vesicles

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