Biological Functions and Cross-Kingdom Host Gene Regulation of Small RNAs in Lactobacillus plantarum-Derived Extracellular Vesicles

Abstract

Extracellular vesicle-mediated transfer of microRNAs is a novel mode of cell-to-cell genetic transmission. Extracellular vesicles produced by microbes have been shown to contain significant quantities of physiologically active molecules such as proteins, lipids, and RNA, which could be transported to host cells and play a key role in both inter-kingdom signaling and physiological responses. In this study, we identified sRNAs by sequencing small RNAs (sRNAs) from Lactobacillus plantarum-derived extracellular vesicles (LDEVs) and detected the expression levels of vesicular sRNAs using quantitative reverse transcription-polymerase chain reaction (RT-PCR), which demonstrated the presence of microRNA-sized RNAs (msRNAs) within these vesicles. We chose sRNA71, a highly expressed msRNA, for further investigation, predicted its potential target genes for the human genome, and indicated that it could be translocated into mammalian cells. The biological functions of this sRNA71 were subsequently explored through cellular proteomics, western blot, and luciferase reporter assay. According to the findings, transfection with synthetic sRNA71 mimics substantially reduced Tp53 expression in HEK293T cells and suppressed the gene expression through binding to the 3′ UTR of Tp53 mRNA. In conclusion, it is hypothesized that microbial-derived extracellular vesicles serve as carriers of functional molecules such as sRNAs, which play an essential role in regulating microbial-host communication.

Probiotic Lactobacilli activate Formyl-Peptide Receptor 2

“…The copyright holder for this preprint this version posted May 7, 2024. ; https://doi.org/10.1101/2024.05.07.592932 doi: bioRxiv preprint However, also the release via membrane vesicles could be an explanation which is supported by vesicle releasing L. plantarum (Yu et al 2022). Interestingly, systematic analysis of the signal peptides and culture filtrates of different lactobacilli suggested that further bacterial species from the genera Lactobacillus, Akkermansia, and Enterococcus with described probiotic potential (Li et al 2024;Im et al 2023) release putative FPR2 ligands.…”

Apilactobacillus kunkeei releases RNA-associated membrane vesicles and proteinaceous nanoparticles

“…Extracellular MVs have been described in both Gram-negative and Gram-positive bacteria, archaea and eukaryotes, and numerous reports have suggested that they serve various functional roles in for example cell-cell communication processes, host adaptation and antimicrobial defense by mediating the transfer of RNA, proteins and metabolites across cell boundaries (Brown et al 2015 , Liu et al 2018 , Gill et al 2019 , Yu et al 2022 ). An alternative hypothesis, particularly for eukaryotic MVs, is that their basic function is to secrete material out from the cell for off-site degradation and recycling of membrane molecules (Vidal 2019 ).…”

“…Another hypothesis is that the secreted RNA molecules mediate bacterial interactions with the host, such as for example serving host immunomodulatory functions (Tsatsaronis et al 2018 , Rodriguez and Kuehn 2020 , Kurata et al 2022 ). While small, non-coding RNAs have been directly associated with immunomodulatory responses (Koeppen et al 2016 , Yu et al 2022 , Sahr et al 2022 ), the specificity of such effects by bulk-RNA are not fully resolved. Furthermore, previous studies have shown that RNAs are secreted as fragments, arguing against a biological role for the large majority of secreted RNA molecules derived from long mRNA molecules (Rodriguez and Kuehn 2020 , Luz et al 2021 , Kurata et al 2022 ).…”

“…For example, rRNA and tRNA fragments were shown to be enriched in the MVs of Prochlorococcus (Biller et al 2014 ) and Escherichia coli (Ghosal et al 2015 , Blenkiron et al 2016 ). Small RNAs (sRNA) of 80–250 nucleotides were identified in the OMVs of Pseudomonas aeruginosa (Koeppen et al 2016 ) and microRNA-size RNA of less than 70 nucleotides were detected in the MVs of several other bacteria (Choi et al 2017 , Sahr et al 2022 , Yu et al 2022 ). A broad study of the composition of RNA molecules in the MVs of Staphylococcus aureus identified 28 sRNAs and a wide range of RNA fragments that mapped to 273 genes coding for virulence-associated factors, ribosomal proteins, transcriptional regulators and metabolic enzymes (Luz et al 2021 ).…”

A promising area of research in medicine: recent advances in properties and applications of Lactobacillus-derived exosomes

“…Additionally, a highly expressed sequence (sRNA71) was identified in Exosomes which were isolated from the culture supernatant of L. plantarum WCFS1 using ultracentrifugation. sRNA71 substantially reduced Tp53 expression in HEK293T cells and suppressed the gene expression through binding to the 3′UTR of Tp53 mRNA ( Yu et al, 2022 ). These exosomes contain bioactive substances that regulate the intestinal microbiome, such as extracellular enzymes, antibiotics, antimicrobials, antibiotic enzymes, iron carrier proteins, β-glucan, siRNA and polysaccharides, which can impact the type and quantity of intestinal flora, improve the growth and metabolism of probiotics in the intestine, and maintain intestinal health ( Uddin et al, 2020 ; Díaz-Garrido et al, 2021 ; Yang Z. et al, 2022 ).…”