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- W4384697949 abstract "Abstract Cells communicate with neighboring cells through shedding and uptake of extracellular vesicles (EVs). EVs are small packages of nucleic acid and protein enclosed in a lipid bilayer. Despite being broadly studied since 2010, the biogenesis of EVs remains elusive. Currently, cells in interphase are thought to shed the bulk of EVs. Given that cancer cells shed more EVs than non-cancerous cells, we hypothesized that the primary origin of large EVs may be mitosis. Here, we demonstrate that the biogenesis of large EVs occurs in mitosis. We found that the canonical EV markers, CD9, CD63, CD81, flotillin-1, and dsRNA, localize to the spindle midzone microtubules, midbody, and midbody remnant (MBR) during mitosis. These EV markers co-localized with the master regulator of cytokinesis, MKLP1, throughout mitosis, suggesting that large EV assembly and packaging occur during mitosis. Next, we showed that standard, commercial, and our newly developed EV isolation methods all isolate MBRs, evidenced by the presence of CD9 and MKLP1 double-positive vesicles. We developed two new methods of large EV isolation utilizing 1.5% PEG 6000 and PEGylated gold nanoparticles. Regardless of the isolation method, EVs are characterized using markers for transmembrane proteins called tetraspanins (CD9, CD63, and CD81), and found that ~99% of all isolated MKLP1-labeled vesicles were also CD9 positive, suggesting that large EVs are MBRs, not a separate class of EVs. Additionally, all isolation methods tested suggest that large EV have translation capacity, distinct from other EV classes. Our results demonstrate that mitotic cells give rise to the bulk of large EVs commonly researched in the field. Importantly, we show that all commercial isolation methods for EVs actually isolate MBRs, suggesting that experiments performed in the EV field are of mitotic origin. This work has broad implications for the assembly and packaging of large EVs, specifically for engineering drug therapeutics and isolation of large EVs, or MBRs, for cancer diagnosis." @default.
- W4384697949 created "2023-07-20" @default.
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- W4384697949 date "2023-07-18" @default.
- W4384697949 modified "2023-09-27" @default.
- W4384697949 title "The biogenesis of large extracellular vesicles occurs in mitosis" @default.
- W4384697949 doi "https://doi.org/10.21203/rs.3.rs-3112385/v1" @default.
- W4384697949 hasPublicationYear "2023" @default.
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