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Cell-of-origin–specific proteomics of extracellular vesicles

The ability to assign cellular origin to low-abundance secreted factors in extracellular vesicles (EVs) would greatly facilitate the analysis of paracrine-mediated signaling. Here, we report a method, named selective isolation of extracellular vesicles (SIEVE), which uses cell type-specific proteome...

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Autores principales: Kehrloesser, Sebastian, Cast, Oliver, Elliott, Thomas S, Ernst, Russell J, Machel, Anne C, Chen, Jia-Xuan, Chin, Jason W, Miller, Martin L
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Oxford University Press 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10119638/
https://www.ncbi.nlm.nih.gov/pubmed/37091541
http://dx.doi.org/10.1093/pnasnexus/pgad107
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author Kehrloesser, Sebastian
Cast, Oliver
Elliott, Thomas S
Ernst, Russell J
Machel, Anne C
Chen, Jia-Xuan
Chin, Jason W
Miller, Martin L
author_facet Kehrloesser, Sebastian
Cast, Oliver
Elliott, Thomas S
Ernst, Russell J
Machel, Anne C
Chen, Jia-Xuan
Chin, Jason W
Miller, Martin L
author_sort Kehrloesser, Sebastian
collection PubMed
description The ability to assign cellular origin to low-abundance secreted factors in extracellular vesicles (EVs) would greatly facilitate the analysis of paracrine-mediated signaling. Here, we report a method, named selective isolation of extracellular vesicles (SIEVE), which uses cell type-specific proteome labeling via stochastic orthogonal recoding of translation (SORT) to install bioorthogonal reactive groups into the proteins derived from the cells targeted for labeling. We establish the native purification of intact EVs from a target cell, via a bioorthogonal tetrazine ligation, leading to copurification of the largely unlabeled EV proteome from the same cell. SIEVE enables capture of EV proteins at levels comparable with those obtained by antibody-based methods, which capture all EVs regardless of cellular origin, and at levels 20× higher than direct capture of SORT-labeled proteins. Using proteomic analysis, we analyze nonlabeled cargo proteins of EVs and show that the enhanced sensitivity of SIEVE allows for unbiased and comprehensive analysis of EV proteins from subpopulations of cells as well as for cell-specific EV proteomics in complex coculture systems. SIEVE can be applied with high efficiency in a diverse range of existing model systems for cell–cell communication and has direct applications for cell-of-origin EV analysis and for protein biomarker discovery.
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spelling pubmed-101196382023-04-22 Cell-of-origin–specific proteomics of extracellular vesicles Kehrloesser, Sebastian Cast, Oliver Elliott, Thomas S Ernst, Russell J Machel, Anne C Chen, Jia-Xuan Chin, Jason W Miller, Martin L PNAS Nexus Biological, Health, and Medical Sciences The ability to assign cellular origin to low-abundance secreted factors in extracellular vesicles (EVs) would greatly facilitate the analysis of paracrine-mediated signaling. Here, we report a method, named selective isolation of extracellular vesicles (SIEVE), which uses cell type-specific proteome labeling via stochastic orthogonal recoding of translation (SORT) to install bioorthogonal reactive groups into the proteins derived from the cells targeted for labeling. We establish the native purification of intact EVs from a target cell, via a bioorthogonal tetrazine ligation, leading to copurification of the largely unlabeled EV proteome from the same cell. SIEVE enables capture of EV proteins at levels comparable with those obtained by antibody-based methods, which capture all EVs regardless of cellular origin, and at levels 20× higher than direct capture of SORT-labeled proteins. Using proteomic analysis, we analyze nonlabeled cargo proteins of EVs and show that the enhanced sensitivity of SIEVE allows for unbiased and comprehensive analysis of EV proteins from subpopulations of cells as well as for cell-specific EV proteomics in complex coculture systems. SIEVE can be applied with high efficiency in a diverse range of existing model systems for cell–cell communication and has direct applications for cell-of-origin EV analysis and for protein biomarker discovery. Oxford University Press 2023-04-03 /pmc/articles/PMC10119638/ /pubmed/37091541 http://dx.doi.org/10.1093/pnasnexus/pgad107 Text en © The Author(s) 2023. Published by Oxford University Press on behalf of National Academy of Sciences. https://creativecommons.org/licenses/by/4.0/This is an Open Access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.
spellingShingle Biological, Health, and Medical Sciences
Kehrloesser, Sebastian
Cast, Oliver
Elliott, Thomas S
Ernst, Russell J
Machel, Anne C
Chen, Jia-Xuan
Chin, Jason W
Miller, Martin L
Cell-of-origin–specific proteomics of extracellular vesicles
title Cell-of-origin–specific proteomics of extracellular vesicles
title_full Cell-of-origin–specific proteomics of extracellular vesicles
title_fullStr Cell-of-origin–specific proteomics of extracellular vesicles
title_full_unstemmed Cell-of-origin–specific proteomics of extracellular vesicles
title_short Cell-of-origin–specific proteomics of extracellular vesicles
title_sort cell-of-origin–specific proteomics of extracellular vesicles
topic Biological, Health, and Medical Sciences
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10119638/
https://www.ncbi.nlm.nih.gov/pubmed/37091541
http://dx.doi.org/10.1093/pnasnexus/pgad107
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