Vesicles From Vibrio cholerae Contain AT-Rich DNA and Shorter mRNAs That Do Not Correlate With Their Protein Products

Abstract

Extracellular vesicles secreted by Gram-negative bacteria have proven to be important in bacterial defense, communication and host–pathogen relationships. They resemble smaller versions of the bacterial mother cell, with similar contents of proteins, LPS, DNA, and RNA. Vesicles can elicit a protective immune response in a range of hosts, and as vaccine candidates, it is of interest to properly characterize their cargo. Genetic sequencing data is already available for vesicles from several bacterial strains, but it is not yet clear how the genetic makeup of vesicles differ from that of their parent cells, and which properties may characterize enriched genetic material. The present study provides evidence for DNA inside vesicles from Vibrio cholerae O395, and key characteristics of their genetic and proteomic content are compared to that of whole cells. DNA analysis reveals enrichment of fragments containing ToxR binding sites, as well as a positive correlation between AT-content and enrichment. Some mRNAs were highly enriched in the vesicle fraction, such as membrane protein genes ompV, ompK, and ompU, DNA-binding protein genes hupA, hupB, ihfB, fis, and ssb, and a negative correlation was found between mRNA enrichment and transcript length, suggesting mRNA inclusion in vesicles may be a size-dependent process. Certain non-coding and functional RNAs were found to be enriched, such as VrrA, GcvB, tmRNA, RNase P, CsrB2, and CsrB3. Mass spectrometry revealed enrichment of outer membrane proteins, known virulence factors, phage components, flagella and extracellular proteins in the vesicle fraction, and a low, negative correlation was found between transcript-, and protein enrichment. This result opposes the hypothesis that a significant degree of protein translation occurs in vesicles after budding. The abundance of viral-, and flagellar proteins in the vesicle fraction underlines the importance of purification during vesicle isolation.

Environmental Plasticity of the RNA Content of Staphylococcus aureus Extracellular Vesicles

“…Second, the relative abundance of several RNAs between two environmental conditions was different in the EVs and the EV-producing cells. Other studies also found that some RNA populations were enriched in EVs from Gram-negative and Gram-positive pathogenic bacteria ( Ghosal et al, 2015 ; Koeppen et al, 2016 ; Resch et al, 2016 ; Malabirade et al, 2018 ; Malge et al, 2018 ; Han et al, 2019 ; Langlete et al, 2019 ; Zhang et al, 2020 ). This notably includes sRNAs, which can play regulatory activity in the host ( Koeppen et al, 2016 ; Malabirade et al, 2018 ; Langlete et al, 2019 ; Zhang et al, 2020 ).…”

“…Other studies also found that some RNA populations were enriched in EVs from Gramnegative and Gram-positive pathogenic bacteria (Ghosal et al, 2015;Koeppen et al, 2016;Resch et al, 2016;Malabirade et al, 2018;Malge et al, 2018;Han et al, 2019;Langlete et al, 2019;Zhang et al, 2020). This notably includes sRNAs, which can play regulatory activity in the host (Koeppen et al, 2016;Malabirade et al, 2018;Langlete et al, 2019;Zhang et al, 2020). The enrichment of RNAs associated with bacterial diseases in EVs derived from many pathogenic bacteria reinforces the physiopathological role of these structures in host-pathogen interaction and host cell invasion, which could be borne by their RNA cargo as well as by their protein cargo.…”

Transcriptome Profiling of Staphylococcus aureus Associated Extracellular Vesicles Reveals Presence of Small RNA-Cargo

“…Sometimes fusion of vesicles have resulted in formation of filamentous structures, also known as nanopods or nanotubes (Dongre et al, 2011 ; Dubey and Ben-Yehuda, 2011 ; Gill et al, 2018 ). EVs may contain virulence factors (Devos et al, 2015 ; Askarian et al, 2018 ; Wagner et al, 2018 ; Nadeem et al, 2020 ), such as toxins (Rivera et al, 2010 ; Coelho et al, 2019 ) as well as other enzymes (Smalley and Birss, 1987 ; Elhenawy et al, 2014 ), quorum sensing molecules (Mashburn and Whiteley, 2005 ; Brameyer et al, 2018 ; Morinaga et al, 2018 ) and nucleic acids such as DNA (Hagemann et al, 2014 ; Bitto et al, 2017 ; Langlete et al, 2019 ) and RNA (Sjöström et al, 2015 ; Koeppen et al, 2016 ; Choi J.-W. et al, 2017 ). Their content may vary depending on species and growth conditions (Bager et al, 2013 ; Ghosal et al, 2015 ; Koeppen et al, 2016 ).…”

Outer Membrane Vesicles Derived from Klebsiella pneumoniae Influence the miRNA Expression Profile in Human Bronchial Epithelial BEAS-2B Cells

“…The surface of these vesicles is composed of lipopolysaccharide (LPS), phospholipids and outer membrane proteins [ 19 , 20 ]. The vesicular lumen, however, contains periplasmic and cytoplasmic components, including genetic material and virulence factors, such as invasion associated factors, toxins, and immune response modulators [ 21 , 22 , 23 ]. Thermolabile toxins and cytolysin have been identified in OMVs produced by Escherichia coli [ 24 , 25 , 26 ].…”