Investigation of RNA in Extracellular Vesicles
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CitationTer-Ovanesyan, Dmitry. 2019. Investigation of RNA in Extracellular Vesicles. Doctoral dissertation, Harvard University, Graduate School of Arts & Sciences.
AbstractExtracellular vesicles (EVs) are small membrane packages that are released by all cells. About a decade ago, it was found that EVs contain RNA, and it was proposed that EVs may transfer RNA between cells as a mechanism of intercellular communication. In this thesis, I used human cell culture systems to develop new methods assessing whether EVs and RNA are transferred between cells, and also extensively characterized the RNA in EVs. Using EVs isolated from human cells, I developed single EV imaging methods and used high throughput RNA sequencing (RNA-Seq) to profile EV RNA. Additionally, I developed a new technique based on enzymatic treatments with proteinase and then RNase to differentiate what extracellular RNA is inside vesicles from that which is outside. Applying this technique, I performed RNA-Seq on RNA isolated from EVs in cell culture media and I found that the mRNA profile in EVs is highly correlated to that mRNA profile of the donor cells. This finding suggests that mRNA is generally packaged into EVs in a non-specific manner, and has significant implications for using EVs to non-invasively read out the transcriptome of human cells from biological fluids. Towards that end, I describe methods for isolating cell-type specific EV, using EVs from neurons as a proof of principle. My work highlights the potential of EVs for diagnostics, but also casts doubt on the notion that EVs are used to physiologically transfer RNA between cells.
Citable link to this pagehttp://nrs.harvard.edu/urn-3:HUL.InstRepos:41121318
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