Fragmentation of extracellular ribosomes and tRNAs shapes the extracellular RNAome

Tosar Rovira, Juan Pablo - Segovia, Mercedes - Castellano, Mauricio - Gámbaro, Fabiana - Akiyama, Y. - Olivera, Álvaro - Costa Camacho, Bruno Alejo - Possi, Tania - Hill, Marcelo - Ivanov, Pavel - Cayota, Alfonso

Resumen:

A major proportion of extracellular RNAs (exRNAs) do not copurify with extracellular vesicles (EVs) and remain in ultracentrifugation supernatants of cell-conditioned medium or mammalian blood serum. However, little is known about exRNAs beyond EVs. We have previously shown that the composition of the nonvesicular exRNA fraction is highly biased toward specific tRNA-derived fragments capable of forming RNase-protecting dimers. To solve the problem of stability in exRNA analysis, we developed a method based on sequencing the size exclusion chromatography (SEC) fractions of nonvesicular extracellular samples treated with RNase inhibitors (RI). This method revealed dramatic compositional changes in exRNA population when enzymatic RNA degradation was inhibited. We demonstrated the presence of ribosomes and full-length tRNAs in cell-conditioned medium of a variety of mammalian cell lines. Their fragmentation generates some small RNAs that are highly resistant to degradation. The extracellular biogenesis of some of the most abundant exRNAs demonstrates that extracellular abundance is not a reliable input to estimate RNA secretion rates. Finally, we showed that chromatographic fractions containing extracellular ribosomes are probably not silent from an immunological perspective and could possibly be decoded as damage-associated molecular patterns.


Detalles Bibliográficos
2020
Inglés
Universidad de la República
COLIBRI
https://hdl.handle.net/20.500.12008/31657
Acceso abierto
Licencia Creative Commons Atribución (CC - By 4.0)

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