A live cell reporter of exosome secretion and uptake reveals pathfinding behavior of migrating cells.
Journal
Nature communications
ISSN: 2041-1723
Titre abrégé: Nat Commun
Pays: England
ID NLM: 101528555
Informations de publication
Date de publication:
29 04 2020
29 04 2020
Historique:
received:
18
07
2019
accepted:
20
03
2020
entrez:
1
5
2020
pubmed:
1
5
2020
medline:
30
7
2020
Statut:
epublish
Résumé
Small extracellular vesicles called exosomes affect multiple autocrine and paracrine cellular phenotypes. Understanding the function of exosomes requires a variety of tools, including live imaging. Our previous live-cell reporter, pHluorin-CD63, allows dynamic subcellular monitoring of exosome secretion in migrating and spreading cells. However, dim fluorescence and the inability to make stably-expressing cell lines limit its use. We incorporated a stabilizing mutation in the pHluorin moiety, M153R, which now exhibits higher, stable expression in cells and superior monitoring of exosome secretion. Using this improved construct, we visualize secreted exosomes in 3D culture and in vivo and identify a role for exosomes in promoting leader-follower behavior in 2D and 3D migration. Incorporating an additional non-pH-sensitive red fluorescent tag allows visualization of the exosome lifecycle, including multivesicular body (MVB) trafficking, MVB fusion, exosome uptake and endosome acidification. This reporter will be a useful tool for understanding both autocrine and paracrine roles of exosomes.
Identifiants
pubmed: 32350252
doi: 10.1038/s41467-020-15747-2
pii: 10.1038/s41467-020-15747-2
pmc: PMC7190671
doi:
Substances chimiques
PHluorin
0
Tetraspanin 30
0
Green Fluorescent Proteins
147336-22-9
Types de publication
Journal Article
Research Support, N.I.H., Extramural
Langues
eng
Sous-ensembles de citation
IM
Pagination
2092Subventions
Organisme : NCI NIH HHS
ID : R01 CA206458
Pays : United States
Organisme : NIGMS NIH HHS
ID : R01 GM117916
Pays : United States
Organisme : NCI NIH HHS
ID : R01 CA249424
Pays : United States
Organisme : NCI NIH HHS
ID : U19 CA179514
Pays : United States
Organisme : NCI NIH HHS
ID : R01 CA216248
Pays : United States
Organisme : NCATS NIH HHS
ID : UL1 TR002243
Pays : United States
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