Investigating the extracellular RNA content of Staphylococcus aureus

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PROJECT SUMMARY/ABSTRACT Extracellular vesicles (EVs) are nano sized, membrane-enclosed vesicles produced and released by cells in all domains of life. EVs contain a variety of biomolecules, such as lipids, proteins, and nucleic acids, and are involved in a broad range of functions depending on the progenitor cell. The study of EVs in Gram-positive bacteria is a relatively new field. Due to the architecture of the Gram-positive cell wall it was long thought to be impossible for Gram-positive bacteria to produce EVs. However recent studies have shown EV production in a variety of Gram-positive bacteria and implicated EVs in important functions like pathogenesis and immune evasion. Staphylococcus aureus is both a commensal of humans and a highly dangerous bacterial pathogen. EV production by S. aureus has been demonstrated and EVs shown to be cytolytic and play roles in immune activation. Studies of S. aureus EV composition have focused on their protein content and to date there are no reports describing the nucleic acid content of S. aureus EVs. S. aureus DNA and RNA are known to elicit an immune response however the mechanism(s) through which these nucleic acids gain access to the host cell is unknown. We hypothesize that EVs represent one mechanism through which S. aureus DNA/RNA is introduced into host cells. In this exploratory proposal we present preliminary data showing that RNA is located inside and associated with S. aureus EVs. We propose to identify the EV-associated RNA in EVs from five different S. aureus strains by RNA-sequencing, and to use that information to define the S. aureus EV-RNome – i.e. the RNA molecules found in EVs from all S. aureus strains. Defining the EV RNome will allow us to identify candidate RNA molecules for future analysis to determine their contribution to the immune stimulating properties of EVs.