Abstract
Extracellular vesicles (EVs) mediate the intercellular transfer of RNAs, which alter gene expression in target cells. EV heterogeneity has limited progress towards defining their physiological functions and utility as disease-specific biomarkers. CD63 and MHC1 are widely used as markers to purify EVs. CD47 is also present on EVs and alters their effects on target cells, suggesting that specific surface markers define functionally distinct EVs. This hypothesis was addressed by comparing Jurkat T cell EVs captured using CD47, CD63, and MHC1 antibodies. These EV subsets have similar sizes but divergent RNA contents. Apart from differences in numbers of nonannotated transcripts, CD63+, MHC1+, and CD47+ EVs have similar overall contents of most noncoding RNA classes, but the relative enrichment of specific RNAs differs. The enrichment of micro-RNAs is highly divergent, and some including miR320a are selectively concentrated in CD47+ EVs. Small nucleolar RNAs including SNORD116@ and SNHG10 are also selectively enriched in CD47+ EVs, whereas no small nuclear RNAs are enriched in CD47+ EVs. Conversely, MHC1+ EVs are selectively enriched in a subset of tRNAs including TRE-CTC and TRR-CCG. This heterogeneity in RNA composition suggests multiple sorting mechanisms that direct specific RNAs into subsets of EVs that express specific surface markers.
Highlights
Ongoing investigations of extracellular vesicles (EVs) are revealing diverse and complex functions in cell-cell communication, mediated in part by their role in the intercellular transfer of RNAs1–3
We found that T cell-derived Extracellular vesicles (EVs) alter gene expression and functional signaling in endothelial cells in a CD47-dependent manner[21]
Bulk EVs isolated from Jurkat T cell conditioned medium using a commercial polymeric precipitation method were fractionated using ~15 nm magnetic nanoparticles (MNPs) conjugated with antibodies specific for the exosome markers CD63, MHC class 1 (MHC1), and CD47
Summary
Ongoing investigations of extracellular vesicles (EVs) are revealing diverse and complex functions in cell-cell communication, mediated in part by their role in the intercellular transfer of RNAs1–3. Engineered EVs are promising deliver vehicles for therapeutic uses including the targeted delivery of miRNAs9,10 To achieve these goals, barriers must be overcome to standardize EV nomenclature[11], characterize their heterogeneity[12], and define the molecular mechanisms controlling passive or active sorting of specific RNAs into different types of EVs13. We report here that each marker-defined subset of EVs has a distinct RNA profile and is enriched in different miRNAs and other coding and noncoding RNAs relative to EVs lacking each respective marker. This suggests the existence of multiple sorting pathways that package RNAs into distinct EV populations within the same cell
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