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Architecture of the Vibrio cholerae toxin-coregulated pilus machine revealed by electron cryotomography

Nature Microbiology volume 2, Article number: 16269 (2017) Cite this article

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Abstract

Type IV pili (T4P) are filamentous appendages found on many Bacteria and Archaea. They are helical fibres of pilin proteins assembled by a multi-component macromolecular machine we call the basal body. Based on pilin features, T4P are classified into type IVa pili (T4aP) and type IVb pili (T4bP)1,2. T4aP are more widespread and are involved in cell motility3, DNA transfer4, host predation5 and electron transfer6. T4bP are less prevalent and are mainly found in enteropathogenic bacteria, where they play key roles in host colonization7. Following similar work on T4aP machines8,9, here we use electron cryotomography10 to reveal the three-dimensional in situ structure of a T4bP machine in its piliated and non-piliated states. The specific machine we analyse is the Vibrio cholerae toxin-coregulated pilus machine (TCPM). Although only about half of the components of the TCPM show sequence homology to components of the previously analysed Myxococcus xanthus T4aP machine (T4aPM), we find that their structures are nevertheless remarkably similar. Based on homologies with components of the M. xanthus T4aPM and additional reconstructions of TCPM mutants in which the non-homologous proteins are individually deleted, we propose locations for all eight TCPM components within the complex. Non-homologous proteins in the T4aPM and TCPM are found to form similar structures, suggesting new hypotheses for their functions and evolutionary histories.

Intact T4aPMs have already been visualized by electron cryotomography (ECT) in Myxococcus xanthus and Thermus thermophilus and were seen to consist of interconnected rings spanning the cell envelope8,9. In M. xanthus, the locations of the various T4aPM components were determined by imaging mutants with individual components either deleted or fused to protein tags8. Fitting atomic models of components11–24 into the reconstructions produced new insights into the overall structure and function of the T4aPM (ref. 8). Although approximately half (five of nine) of the structural components of the T4bPM exhibit sequence homology to T4aPM components and are therefore expected to have similar structures and functions, the other half are novel and so their positions and roles remain unclear.

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Figure 1: Visualizing the TCPM in intact V. cholerae cells.
Figure 2: Comparison between V. cholerae TCPM and M. xanthus T4aPM structures and the inferred TCPM component locations based on the T4aPM component map.
Figure 3: Structures of TCPM in ΔtcpS, ΔtcpB, ΔtcpD and ΔtcpR cells.
Figure 4: Locations of TCPM components and comparison with M. xanthus T4aPM.

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Acknowledgements

The authors thank C. Oikonomou and C. Shaffer for discussions and editorial assistance. This work was supported by NIH grant R01 AI127401 to G.J.J., the Howard Hughes Medical Institute and the John Templeton Foundation as part of the Boundaries of Life project. The opinions expressed in this publication are those of the authors and do not necessarily reflect the views of the John Templeton Foundation.

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Author notes

  1. Ronald K. Taylor: Deceased.

Authors and Affiliations

  1. California Institute of Technology, Pasadena, 91125, California, USA

    Yi-Wei Chang, Davi R. Ortega & Grant J. Jensen

  2. University of Southern Denmark, Campusvej 55, 5230 Odense M, Denmark

    Andreas Kjær

  3. Geisel School of Medicine at Dartmouth, Hanover, 03755, New Hampshire, USA

    Gabriela Kovacikova, John A. Sutherland & Ronald K. Taylor

  4. Howard Hughes Medical Institute, Pasadena, 91125, California, USA

    Lee A. Rettberg & Grant J. Jensen

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Contributions

Y.-W.C. and A.K. collected, processed and analysed the ECT data. D.R.O. performed the bioinformatics analyses. L.A.R. assisted with ECT data processing. G.K., J.A.S. and R.K.T. provided the V. cholerae strains. Y.-W.C., A.K., D.R.O. and G.J.J. wrote the paper.

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Correspondence to Grant J. Jensen.

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Supplementary information

Supplementary Information

Supplementary Discussion, Supplementary Figures 1–7, Supplementary Tables 1–3, Supplementary References. (PDF 14686 kb)

Supplementary Video 1

Comparison of the TCPM conformation in piliated and non-piliated states. Conformational differences between piliated and non-piliated TCPMs are revealed by morphing from one density to another. (MOV 575 kb)

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Chang, YW., Kjær, A., Ortega, D. et al. Architecture of the Vibrio cholerae toxin-coregulated pilus machine revealed by electron cryotomography. Nat Microbiol 2, 16269 (2017). https://doi.org/10.1038/nmicrobiol.2016.269

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