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A new view into prokaryotic cell biology from electron cryotomography

A Corrigendum to this article was published on 28 December 2016

This article has been updated

Key Points

  • Electron cryotomography (ECT) can reveal the structure of intact cells in a native state in 3D to macromolecular (4 nm) resolution.

  • ECT bridges the resolution gap between atomic structures of individual proteins (solved by X-ray crystallography, NMR spectroscopy and cryo-electron microscopy single-particle reconstruction) and overall cell morphology as visualized by light microscopy.

  • ECT has provided new insights into many aspects of prokaryotic cell biology, including basic membrane structure, cell wall architecture, morphogenesis, basic metabolism, motility, chemotaxis, sporulation, inter-species cooperation and competition, and viral infection.

  • ECT has enabled the direct visualization, for the first time, of the arrangement of polymers in the bacterial cell wall. This revealed unexpected structural conservation between Gram-positive and Gram-negative envelopes, eroding the distinction between them, as well as conversion of an inner to an outer membrane during sporulation, which suggests a new hypothesis for the evolution of the diderm cell plan.

  • ECT has confirmed the existence of a bacterial cytoskeleton, and also uncovered its complexity, including diverse filaments mediating processes that range from shape maintenance to motility to division.

  • ECT has revealed the structures of large nanomachines (macromolecular complexes that comprise many unique components), such as flagellar motors, chemoreceptor arrays and secretion systems intact in their native state within cells. Because these nanomachines typically cannot be purified or reconstituted completely for traditional structural analysis, ECT has provided a necessary new source of information on their overall architecture. Placing atomic models of components into their relative positions within ECT reconstructions sheds light on the mechanisms these machines use to carry out their functions; for example, in motility, interactions with other organisms and pathogenicity.

  • Continuing technological developments are increasing the quality of ECT data. Combined with other structure determination methods and fluorescence microscopy, ECT is moving us towards the ultimate goal: a complete understanding of the structure and location of every protein in a prokaryotic cell.


Electron cryotomography (ECT) enables intact cells to be visualized in 3D in an essentially native state to 'macromolecular' (4 nm) resolution, revealing the basic architectures of complete nanomachines and their arrangements in situ. Since its inception, ECT has advanced our understanding of many aspects of prokaryotic cell biology, from morphogenesis to subcellular compartmentalization and from metabolism to complex interspecies interactions. In this Review, we highlight how ECT has provided structural and mechanistic insights into the physiology of bacteria and archaea and discuss prospects for the future.

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Figure 1: Example of electron cryotomography.
Figure 2: Cellular organization.
Figure 3: Motility and navigation.
Figure 4: Division and differentiation.
Figure 5: Interaction with other cells.

Change history

  • 30 November 2016

    It has recently come to the authors' attention that this Review is the first place in which Figure 1 was published. Yi-Wei Chang, who collected the data for Figure 1 and helped to revise the text, should have been included as an author. Therefore, on the title page, the corrected author list should read Catherine M. Oikonomou, Yi-Wei Chang and Grant J. Jensen


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The authors apologize that they could not discuss all of the work in this burgeoning field. The authors thank members of the Jensen laboratory for helpful comments on the manuscript, and J. Ding and Y.-W. Chang for producing the accompanying movie. The authors also thank L. Sockett (University of Nottingham) for the gift of the Bdellovibrio bacteriovorus strain imaged in figure 1 and shown in the accompanying movie. Microbial electron cryotomography (ECT) in the Jensen laboatory is supported, in part, by the Howard Hughes Medical Institute, the US National Institutes of Health (grants RO1 GM101425 and RO1 GM094800), the Beckman Institute at Caltech, Caltech's Center for Environmental Microbial Interactions, and gifts to Caltech from the Gordon and Betty Moore Foundation and the Agouron Institute.

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Correspondence to Grant J. Jensen.

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Supplementary information

Supplementary information S1 (movie)

An intact Bdellovibrio bacteriovorus cell in standard media was plunge-frozen and imaged by ECT. The resulting tilt-series of images was reconstructed into a 3D tomogram. The movie shows the full reconstruction and segmentation, as well as fitting of crystal structures into EM densities. (MOV 190651 kb)

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Phase plates

Electron microscope components that shift the phases of the scattered beam with respect to the unscattered beam to boost contrast.

Vitreous cryosectioning

A sample preparation technique in which thick samples are frozen rapidly to prevent ice formation and then cut with a diamond blade into thin (50–400 nm) slices that can be imaged by electron cryotomography.


A polymer crosslinked into a mesh-like network that forms the cell walls of most bacteria.


Sac-like exoskeletons (peptidoglycan cell walls) of bacterial cells.

Stalked cell

In the context of Caulobacter crescentus, one of the two cells produced by asymmetric division. Unlike the motile swarmer cell, the stalked cell is attached to a surface and is capable of replication and division. Swarmer cells later differentiate into stalked cells.


Compartments containing ferrous microcrystals that are used by magnetotactic bacteria to orient the cell in a magnetic field.

Template matching

A digital image processing technique to search a 3D tomogram for an object of interest (the template). The template is typically a single-particle reconstruction or X-ray crystallographical structure of a macromolecule or complex of interest.

Archaeal Richmond Mine acidophilic nanoorganisms

(ARMAN). A highly divergent group of archaea isolated from an extremely acidic environment in California.

Reaction centres

Complexes of enzymes, pigments and cofactors that convert solar energy that is captured by light-harvesting antennae into chemical energy during photosynthesis.

Correlated light microscopy and ECT

A specialization of correlated light microscopy and electron microscopy (CLEM) that is used to locate structures of interest within visually crowded tomograms. Cells containing a fluorescently labelled target protein are rapidly frozen on electron microscope grids that contain landmarks for correlation, and first imaged by light microscopy to identify the location of fluorescent signals. Grids are then transferred to the electron microscope and the same locations are imaged at high resolution by electron cryotomography (ECT).

Electron microscopy docking

A method for modelling the structure of macromolecular complexes by fitting high-resolution models (like X-ray crystallographical structures) of components into electron microscopy density maps, often informed by biochemical studies detailing component interactions.

Subtomogram averaging

An image processing technique to increase the clarity of structures that are present in multiple copies and/or in multiple tomograms. Averaging particles increases the signal-to-noise ratio and can yield reliable high-resolution (<1 nm) detail, including secondary protein structure.

Endosomal-sorting complexes required for transport

(ESCRT). Protein complexes that remodel cellular membranes to carry out various processes, including cell division and viral budding.

Diderm cell plan

Under a classification system not based on the Gram stain, diderm bacteria are surrounded by two lipid bilayer membranes, as opposed to monoderms, which have only one.

Correlated cryo-PALM and ECT

A specialized application of correlated light microscopy and electron cryotomography (ECT) using a super-resolution microscopy technique — photoactivated localization microscopy (PALM) — to increase the localization precision of a structure of interest in fluorescence images for correlation with high-resolution ECT images.


Type VI secretion system (T6SS)-like contractile nanomachines, related to phage tails, that bacteria use to kill other bacteria.

Elementary bodies

The non-replicating forms of Chlamydia spp., which are responsible for cellular infection.

Reticulate bodies

The metabolically active forms of Chlamydia spp., which are found in the cytoplasm of infected cells.

Single-particle reconstruction

A transmission electron microscopy technique in which many identical copies of a purified macromolecule or complex are imaged, providing different projection views of the particle that can then be computationally combined into a 3D reconstruction.

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Oikonomou, C., Chang, YW. & Jensen, G. A new view into prokaryotic cell biology from electron cryotomography. Nat Rev Microbiol 14, 205–220 (2016).

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