Functional and molecular characterization of host cell microRNAs controlling Salmonella infection
Final Report Abstract
Increasing evidence suggests an important role for miRNAs in the molecular interplay between bacterial pathogens and host cells. By performing fluorescence microscopy-based screenings using a genome-wide library of miRNA mimics we identified miRNAs that strongly modulate infection by the bacterial pathogen Salmonella Typhimurium. During this project we characterized in detail the mechanisms by which several of these miRNAs, specifically the miR-15 family and let-7i-3p, inhibit Salmonella infection. Regarding the miR-15 family inhibition of Salmonella infection, we discovered that the effect of these miRNAs is related to the host cell cycle. Specifically, these miRNAs inhibit cell cycle progression, delaying the host cell cycle on the G1/S phase, which we uncovered as inhibitory to Salmonella replication. Importantly, we discovered that these miRNAs are downregulated during Salmonella infection, through the inhibition of the transcription factor E2F1. Thus the derepression of cell cycle related genes (e.g. cyclin D1) during infection and the consequent promotion of G1/S transition are crucial for Salmonella replication in epithelial cells. In addition, we discovered that Salmonella induces G2/M cell cycle arrest in infected cells, further promoting its replication. Overall, these findings uncover a mechanism whereby Salmonella renders host cells more susceptible to infection by controlling cell cycle progression through the active modulation of host cell miRNAs. Regarding let-7i-3p, we characterized the dual regulatory role of this miRNA in the context of Salmonella infection, showing that let-7i-3p inhibits both early and late steps of infection. We demonstrate that let-7i-3p inhibits Salmonella binding to host cells through the induction of actin stress fibers. Moreover, we identify RGS2 protein as a let-7i-3p target and demonstrate that its previously unknown function as regulator of endolysosomal trafficking modulates the late steps of infection, namely bacterial replication within Salmonella containing vacuoles. Importantly, Salmonella decreases let-7i-3p expression through the inhibition of the transcription factor E2F1. By down-regulating let-7i-3p, Salmonella relieves the inhibition imposed by this miRNA on two independent steps of infection, ultimately promoting both bacterial intracellular replication and dissemination. Overall, we characterized the effect of the miR-15 family and let-7i-3p on infection, further contributing to demonstrate the relevance of miRNAs in the infection by bacterial pathogens, and that the characterization of miRNAs and their targets is a relevant strategy to reveal novel host pathways important in the interaction between bacterial pathogens and the host.
Publications
- (2014) Functional high-throughput screening identifies the miR-15 microRNA family as cellular restriction factors for Salmonella infection. Nature Communications 5:4718
Maudet C, Mano M, Sunkavalli U, Sharan M, Giacca M, Forstner K, Eulalio A
(See online at https://doi.org/10.1038/ncomms5718) - (2014) MicroRNAs in the interaction between host and bacterial pathogens. FEBS Letters 588(22):4140-4147
Maudet C, Mano M, Eulalio A
(See online at https://doi.org/10.1016/j.febslet.2014.08.002) - (2015) MicroRNA Screening and the Quest for Biologically Relevant Targets. Journal of Biomolecular Screening 20(8):1003-1017
Eulalio A, Mano M
(See online at https://doi.org/10.1177/1087057115578837) - (2017) Analysis of host microRNA function uncovers a role for miR-29b-2-5p in Shigella capture by filopodia. PLoS Pathogens 13(4):e1006327
Sunkavalli U, Aguilar C, Silva RJ, Sharan M, Cruz AR, Tawk C, Maudet C, Mano M, Eulalio A
(See online at https://doi.org/10.1371/journal.ppat.1006327) - (2020) Functional screenings reveal different requirements for host microRNAs in Salmonella and Shigella infection. Nature Microbiology, 5(1):192-205
Aguilar C, Cruz AR, Maudet C, Rodrigues Lopes I, Sunkavalli U, Silva RJ, Sharan M, Lisowski C, Zaldívar-López S, Garrido JJ, Giacca M, Mano M, Eulalio A
(See online at https://doi.org/10.1038/s41564-019-0614-3)
