Abstract
So far, bacterial regulatory sRNAs of length less than 50 nucleotides have been poorly understood, and a low number of such molecules has been identified. The first microRNA-size functional ribonucleic acid occurring in a bacterial cell has been described only recently, and it was found to be encoded by a bacteriophage. One of the reasons for such a scarcity in this field is the lack of procedures intended for the isolation and selection of molecules of this size from bacterial cells. To meet these difficulties, we describe here the few-step procedure of isolation, purification, selection, and sequencing library preparation that is dedicated to the fraction of very small, bacterial RNA molecules.
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References
Gottesman S, McCullen CA, Guillier M, Vanderpool CK, Majdalani N, Benhammou J, Thompson KM, FitzGerald PC, Sowa NA, FitzGerald DJ (2006) Small RNA regulators and the bacterial response to stress. Cold Spring Harb Symp Quant Biol 71:1–11. https://doi.org/10.1101/sqb.2006.71.016
Djapgne L, Oglesby AG (2021) Impacts of small RNAs and their chaperones on bacterial pathogenicity. Front Cell Infect Microbiol 11:604511. https://doi.org/10.3389/fcimb.2021.604511
Michaux C, Verneuil N, Hartke A, Giard JC (2014) Physiological roles of small RNA molecules. Microbiology 160:1007–1019. https://doi.org/10.1099/mic.0.076208-0
Richards GR, Vanderpool CK (2011) Molecular call and response: the physiology of bacterial small RNAs. Biochim Biophys Acta 1809:525–531. https://doi.org/10.1016/j.bbagrm.2011.07.013
Vanderpool CK, Balasubramanian D, Lloyd CR (2011) Dual-function RNA regulators in bacteria. Biochimie 93:1943–1949. https://doi.org/10.1016/j.biochi.2011.07.016
Bak G, Lee J, Suk S, Kim D, Young Lee J, Kim KS, Choi BS, Lee Y (2015) Identification of novel sRNAs involved in biofilm formation, motility, and fimbriae formation in Escherichia coli. Sci Rep 5:15287. https://doi.org/10.1038/srep15287
Gottesman S, Storz G (2011) Bacterial small RNA regulators: versatile roles and rapidly evolving variations. Cold Spring Harb Perspect Biol 3:a003798. https://doi.org/10.1101/cshperspect.a003798
Tree JJ, Granneman S, McAteer SP, Tollervey D, Gally DL (2014) Identification of bacteriophage-encoded anti-sRNAs in pathogenic Escherichia coli. Mol Cell 55:199–213. https://doi.org/10.1016/j.molcel.2014.05.006
Bloch S, Lewandowska N, Węgrzyn G, Nejman-Faleńczyk B (2021) Bacteriophages as sources of small non-coding RNA molecules. Plasmid 113:102527. https://doi.org/10.1016/j.plasmid.2020.102527
Lee HJ, Hong SH (2012) Analysis of microRNA-size, small RNAs in Streptococcus mutans by deep sequencing. FEMS Microbiol Lett 326:131–136. https://doi.org/10.1111/j.1574-6968.2011.02441.x
Kang SM, Choi JW, Lee Y, Hong SH, Lee HJ (2013) Identification of microRNA-size, small RNAs in Escherichia coli. Curr Microbiol 67:609–613. https://doi.org/10.1007/s00284-013-0411-9
Furuse Y, Finethy R, Saka HA, Xet-Mull AM, Sisk DM, Smith KL, Lee S, Coers J, Valdivia RH, Tobin DM, Cullen BR (2014) Search for microRNAs expressed by intracellular bacterial pathogens in infected mammalian cells. PLoS One 9:e106434. https://doi.org/10.1371/journal.pone.0106434
Choi JW, Kim SC, Hong SH, Lee HJ (2017) Secretable small RNAs via outer membrane vesicles in periodontal pathogens. J Dent Res 96:458–466. https://doi.org/10.1177/0022034516685071
Choi JW, Kwon T, Hong SH, Lee HJ (2016) Isolation and characterization of a microRNA-size secretable small RNA in Streptococcus sanguinis. Cell Biochem Biophys 76:293–301. https://doi.org/10.1007/s12013-016-0770-5
Nejman-Faleńczyk B, Bloch S, Licznerska K, Dydecka A, Felczykowska A, Topka G, Węgrzyn A, Węgrzyn G (2015) A small, microRNA-size, ribonucleic acid regulating gene expression and development of Shiga toxin-converting bacteriophage Φ24Β. Sci Rep 5:10080. https://doi.org/10.1038/srep10080
Desgranges E, Caldelari I, Marzi S, Lalaouna D (2020) Navigation through the twists and turns of RNA sequencing technologies: application to bacterial regulatory RNAs. Biochim Biophys Acta Gene Regul Mech 1863:194506. https://doi.org/10.1016/j.bbagrm.2020
Diallo I, Provost P (2020) RNA-sequencing analyses of small bacterial RNAs and their emergence as virulence factors in host-pathogen interactions. Int J Mol Sci 21:1627. https://doi.org/10.3390/ijms21051627
Acknowledgments
This work was supported by the National Science Centre (Poland) grants No. 2018/29/B/NZ1/00549 to G.W. and No. 2018/30/E/NZ1/00400 to B.N-F.
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Bloch, S. et al. (2024). Analysis of Phage Regulatory RNAs: Sequencing Library Construction from the Fraction of Small Prokaryotic RNAs Less Than 50 Nucleotides in Length. In: Arluison, V., Valverde, C. (eds) Bacterial Regulatory RNA. Methods in Molecular Biology, vol 2741. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-3565-0_3
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DOI: https://doi.org/10.1007/978-1-0716-3565-0_3
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