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Live-Cell Imaging of the Assembly and Ejection Processes of the Bacterial Flagella by Fluorescence Microscopy

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Bacterial and Archaeal Motility

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2646))

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Abstract

Bacterial flagella are molecular machines used for motility and chemotaxis. The flagellum consists of a thin extracellular helical filament as a propeller, a short hook as a universal joint, and a basal body as a rotary motor. The filament is made up of more than 20,000 flagellin molecules and can grow to several micrometers long but only 20 nanometers thick. The regulation of flagellar assembly and ejection is important for bacterial environmental adaptation. However, due to the technical difficulty to observe these nanostructures in live cells, our understanding of the flagellar growth and loss is limited. In the last three decades, the development of fluorescence microscopy and fluorescence labeling of specific cellular structure has made it possible to perform the real-time observation of bacterial flagellar assembly and ejection processes. Furthermore, flagella are not only critical for bacterial motility but also important antigens stimulating host immune responses. The complete understanding of bacterial flagellar production and ejection is valuable for understanding macromolecular self-assembly, cell adaptation, and pathogen-host interactions.

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References

  1. Macnab RM (2003) How bacteria assemble flagella. Annu Rev Microbiol 57:77–100

    Article  CAS  PubMed  Google Scholar 

  2. Honko AN, Mizel SB (2005) Effects of flagellin on innate and adaptive immunity. Immunol Res 33:83–101

    Article  CAS  PubMed  Google Scholar 

  3. Salazar-Gonzalez RM, McSorley SJ (2005) Salmonella flagellin, a microbial target of the innate and adaptive immune system. Immunol Lett 101:117–122

    Article  CAS  PubMed  Google Scholar 

  4. Miao EA, Andersen-Nissen E, Warren SE et al (2007) TLR5 and Ipaf: dual sensors of bacterial flagellin in the innate immune system. Semin Immunopathol 29:275–288

    Article  CAS  PubMed  Google Scholar 

  5. Armitage JP, Berry RM (2020) Assembly and dynamics of the bacterial flagellum. Annu Rev Microbiol 74:181–200

    Article  CAS  PubMed  Google Scholar 

  6. Nirody JA, Sun Y, Lo C (2017) The biophysicist’s guide to the bacterial flagellar motor. Adv Phys X 2(2):324–343

    CAS  Google Scholar 

  7. Iino T (1974) Assembly of Salmonella flagellin in vitro and in vivo. J Supramol Struct 2:372–384

    Article  CAS  PubMed  Google Scholar 

  8. Turner L, Stern AS, Berg HC (2012) Growth of flagellar filaments of Escherichia coli is independent of filament length. J Bacteriol 194:2437

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Renault TT, Abraham AO, Bergmiller T et al (2017) Bacterial flagella grow through an injection-diffusion mechanism. elife 6:e23136

    Article  PubMed  PubMed Central  Google Scholar 

  10. Chen M, Zhao Z, Yang J et al (2017) Length-dependent flagellar growth of Vibrio alginolyticus revealed by real time fluorescent imaging. elife 6:e22140

    Article  PubMed  PubMed Central  Google Scholar 

  11. Glauert AM, Kerridge D, Horne RW (1963) The fine structure and mode of attachment of the sheathed flagellum of Vibrio metchnikovii. J Cell Biol 18:327–336

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Allen RD, Baumann P (1971) Structure and arrangement of flagella in species of the genus Beneckea and Photobacterium fischeri. J Bacteriol 107:295–302

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. McCarter LL (2001) Polar flagellar motility of the Vibrionaceae. Microbiol Mol Biol Rev 65:445–462

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Grossart HP, Steward GF, Martinez J et al (2000) A simple, rapid method for demonstrating bacterial flagella. Appl Environ Microbiol 66:3632

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Wu Y, Yeh FL, Mao F et al (2009) Biophysical characterization of styryl dye-membrane interactions. Biophys J 97:101–109

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Ferreira JL, Gao FZ, Rossmann FM et al (2019) γ-proteobacteria eject their polar flagella under nutrient depletion, retaining flagellar motor relic structures. PLoS Biol 17:e3000165

    Article  PubMed  PubMed Central  Google Scholar 

  17. Kaplan M, Subramanian P, Ghosal D et al (2019) In situ imaging of the bacterial flagellar motor disassembly and assembly processes. EMBO J 38:e100957

    Article  PubMed  PubMed Central  Google Scholar 

  18. Zhu S, Schniederberend M, Zhitnitsky D et al (2019) In situ structures of polar and lateral flagella revealed by cryo-electron tomography. J Bacteriol 201:e00117–e00119

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Zhuang X, Guo S, Li Z et al (2020) Live-cell fluorescence imaging reveals dynamic production and loss of bacterial flagella. Mol Microbiol 114:279–291

    Article  CAS  PubMed  Google Scholar 

  20. Zhuang X, Lo C (2020) Construction and loss of bacterial flagellar filaments. Biomol Ther 10:1528

    CAS  Google Scholar 

  21. Furuno M, Atsumi T, Yamada T et al (1997) Characterization of polar-flagellar-length mutants in Vibrio alginolyticus. Microbiology 143:1615–1621

    Article  CAS  PubMed  Google Scholar 

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Acknowledgments

The work was supported by the Ministry of Science and Technology of the Republic of China under contract No. MOST-109-2628-M-008-001-MY4.

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Correspondence to Chien-Jung Lo .

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Zhuang, XY., Tseng, CK., Lo, CJ. (2023). Live-Cell Imaging of the Assembly and Ejection Processes of the Bacterial Flagella by Fluorescence Microscopy. In: Minamino, T., Miyata, M., Namba, K. (eds) Bacterial and Archaeal Motility. Methods in Molecular Biology, vol 2646. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-3060-0_4

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  • DOI: https://doi.org/10.1007/978-1-0716-3060-0_4

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-3059-4

  • Online ISBN: 978-1-0716-3060-0

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