CD63, MHC class 1, and CD47 identify subsets of extracellular vesicles containing distinct populations of noncoding RNAs

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.