Extracellular vesicles and their RNA cargo facilitate bidirectional cross-kingdom communication between human and bacterial cells

Groeger, L.; Rishik, S.; Ludwig, N.; Beganovic, A.; Koch, M.; Rheinheimer, S.; Hart, M.; Koenig, P.; Trampert, T.; Paul, P.; Boese, A.; Lehr, C.-M.; Becker, S. L.; Fuhrmann, G.; Keller, A.; Meese, E.

doi:10.1101/2025.07.10.664124

bioRxiv · 10.1101/2025.07.10.664124

Extracellular vesicles and their RNA cargo facilitate bidirectional cross-kingdom communication between human and bacterial cells

Groeger, L.·Rishik, S.·Ludwig, N.·Beganovic, A.·Koch, M.·Rheinheimer, S.·Hart, M.·Koenig, P.·Trampert, T.·Paul, P.·Boese, A.·Lehr, C.-M.·Becker, S. L.·Fuhrmann, G.·Keller, A.·Meese, E.

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Abstract

While extracellular vesicles (EVs) are established mediators of intra-species signaling, their role as active participants in cross-kingdom communication remains incompletely understood. Here, we reveal that human colon cells and both Gram-positive and Gram-negative gut bacteria engage in species-specific, EV-mediated molecular dialogue, driven in part by RNA cargo. We show that bacterial EVs (BEVs) induce distinct transcriptomic responses in human cells, and that BEV-RNA independently causes similar effects. Conversely, we demonstrate that human EVs and highly abundant miR-192-5p are differentially internalized by bacteria, affecting their physiology. Our findings support a conceptual model in which EVs function as directional messengers that shape host-microbiome interactions. This study introduces a framework for understanding EVs as cross-kingdom regulators and underscores the importance of tailored, context-specific analyses for understanding the scope of EV-mediated interactions in microbiome-host homeostasis and disease. Highlights[1] L. casei, E. faecalis and P. mirabilis produce BEVs that are internalized by Caco-2 cells at different rates. BEVs produced by L. casei have a positive influence on the viability of Caco-2 cells. Incubation of Caco-2 cells with BEVs leads to changes in the gene expression of immune-response-related genes. [2] BEVs carry RNAs and the type of RNA cargo varies significantly between the BEVs from the different bacteria. Comparison of Caco-2 gene deregulation between BEVs and transfection of RNA isolated from BEV highlights component-specific effects. [3] Caco-2 EVs are taken up by E. faecalis and influence their growth. MiRNA-192-5p can be frequently detected in EVs from Caco-2 cells. Synthetic miR-192-5p is internalized by P. mirabilis and the ability to take up human miRNAs by L. casei and E. faecalis can be increased by packaging of the miRNA in artificial liposomes.

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Groeger, L., Rishik, S., Ludwig, N., Beganovic, A., Koch, M., Rheinheimer, S., Hart, M., Koenig, P., Trampert, T., Paul, P., Boese, A., Lehr, C.-M., Becker, S. L., Fuhrmann, G., Keller, A., Meese, E.. 2025-07-15. Extracellular vesicles and their RNA cargo facilitate bidirectional cross-kingdom communication between human and bacterial cells. https://doi.org/10.1101/2025.07.10.664124

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