Rli51 Attenuates Transcription of the Listeria Pathogenicity Island 1 Gene mpl and Functions as a Trans-Acting sRNA in Intracellular Bacteria
Abstract
1. Introduction
2. Results
2.1. Rli51 Does Not Play a Relevant Role in the Upregulation of mpl during Infection
2.2. Condition-Specific Premature Termination of Rli51-mpl Transcription
2.3. Trans-Acting Regulation of Rli51-Mediated Transcriptional Termination
2.4. Rli51 Is a Trans-Acting sRNA in Bacteria Infecting Epithelial Cells
3. Discussion
4. Materials and Methods
4.1. Bacterial Strains and Growth Conditions
4.2. Cloning Procedures
4.3. Determination of L. monocytogenes Intracellular Proliferation (Gentamicin-Resistance Assay)
4.4. RNA Isolation from Cultured and Intracellular Bacteria
4.5. Reverse Transcription and Real-Time Quantitative PCR (qPCR)
4.6. Production of Recombinant His-MBP-CP
4.7. MS2-Affinity Purification of Rli51-Binding RNAs (RNA Pull Down)
4.8. PASIFIC Structural Analysis of Rli51
4.9. sRNA-sRNA Interaction Prediction
4.10. RNA Sequencing: Library Construction and Next-Generation DNA Sequencing
4.11. RNA Sequencing Data Analysis
5. Conclusions
- Rli51 is an sRNA located in the 5′UTR of the LIPI-1 virulence factor mpl that functions both as a cis- and trans-acting sRNA.
- Under nutrient-rich laboratory conditions, the transcription of rli51-mpl is prematurely terminated, producing a 121-nucleotide-long sRNA at very low levels.
- The sRNA Rli21/RliI binds to a single-stranded RNA loop in Rli51, which is crucial for premature transcription termination.
- Under intracellular infection conditions, rli51 is induced, leading to a higher abundance of the short Rli51 sRNA and allowing for transcriptional read-through into mpl.
- Transcriptional read-through allows Mpl virulence factor expression in intracellular bacteria and the short Rli51 sRNA regulates in trans transcripts encoding iron scavenging and cell surface proteins.
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Morón, Á.; Ortiz-Miravalles, L.; Peñalver, M.; García-del Portillo, F.; Pucciarelli, M.G.; Ortega, A.D. Rli51 Attenuates Transcription of the Listeria Pathogenicity Island 1 Gene mpl and Functions as a Trans-Acting sRNA in Intracellular Bacteria. Int. J. Mol. Sci. 2024, 25, 9380. https://doi.org/10.3390/ijms25179380
Morón Á, Ortiz-Miravalles L, Peñalver M, García-del Portillo F, Pucciarelli MG, Ortega AD. Rli51 Attenuates Transcription of the Listeria Pathogenicity Island 1 Gene mpl and Functions as a Trans-Acting sRNA in Intracellular Bacteria. International Journal of Molecular Sciences. 2024; 25(17):9380. https://doi.org/10.3390/ijms25179380
Chicago/Turabian StyleMorón, Álvaro, Laura Ortiz-Miravalles, Marcos Peñalver, Francisco García-del Portillo, M. Graciela Pucciarelli, and Alvaro Darío Ortega. 2024. "Rli51 Attenuates Transcription of the Listeria Pathogenicity Island 1 Gene mpl and Functions as a Trans-Acting sRNA in Intracellular Bacteria" International Journal of Molecular Sciences 25, no. 17: 9380. https://doi.org/10.3390/ijms25179380
APA StyleMorón, Á., Ortiz-Miravalles, L., Peñalver, M., García-del Portillo, F., Pucciarelli, M. G., & Ortega, A. D. (2024). Rli51 Attenuates Transcription of the Listeria Pathogenicity Island 1 Gene mpl and Functions as a Trans-Acting sRNA in Intracellular Bacteria. International Journal of Molecular Sciences, 25(17), 9380. https://doi.org/10.3390/ijms25179380