Uncharacterized Bacterial Structures Revealed by Electron Cryotomography

Abstract

Electron cryotomography (ECT) can reveal the native structure and arrangement of macromolecular complexes inside intact cells. This technique has greatly advanced our understanding of the ultrastructure of bacterial cells. We now view bacteria as structurally complex assemblies of macromolecular machines rather than as undifferentiated bags of enzymes. To date, our group has applied ECT to nearly 90 different bacterial species, collecting more than 15,000 cryotomograms. In addition to known structures, we have observed, to our knowledge, several uncharacterized features in these tomograms. Some are completely novel structures; others expand the features or species range of known structure types. Here, we present a survey of these uncharacterized bacterial structures in the hopes of accelerating their identification and study, and furthering our understanding of the structural complexity of bacterial cells.IMPORTANCE Bacteria are more structurally complex than is commonly appreciated. Here we present a survey of previously uncharacterized structures that we observed in bacterial cells by electron cryotomography, structures that will initiate new lines of research investigating their identities and roles.KEYWORDS bacteria, electron cryotomography, bacterial ultrastructure, uncharacterized structures, electron microscopy T he history of cell biology has been punctuated by advances in imaging technology. In particular, the development of electron microscopy (EM) in the 1930s produced a wealth of new information about the ultrastructure of cells (1). For the first time, the structure of cell envelopes, internal organelles, cytoskeletal filaments, and even large macromolecular complexes like ribosomes became visible. A further advance came in the 1980s and 1990s with the development of electron cryotomography (ECT) (2), which allows small cells to be imaged intact in 3D in a near-native, "frozen-hydrated" state to "macromolecular" (ϳ5 nm) resolution, without the limitations and artifacts of more traditional specimen-preparation methods (3).ECT has helped reveal the previously unappreciated complexity of "simple" bacterial cells. Our group has been using ECT to study bacteria for more than a decade, generating more than 15,000 tomograms of 88 different species. These tomograms have revealed new insights into, among other things, the bacterial cytoskeleton (4-7), cell wall architecture (8, 9), morphogenesis (10), metabolism (11), motility (12-15),

Bacterial swarming reducesProteus mirabilisandVibrio parahaemolyticuscell stiffness and increases β-lactam susceptibility

“…These changes are consistent with what would be expected from a reduction in PG thickness in swarming cells. We also observed that the length of polysaccharides was reduced in swarming cells, which would not obviously lead to large changes in cell elongation during osmotic shifts if the organization of PG in these cells follows what is known about the structure of PG in E. coli cells (33): polysaccharides reported to be arranged circumferentially around cells with the peptide cross-links oriented parallel to the long axis of cells. Instead, we expect that a decrease in polysaccharide length makes swarmer cells more prone to changes in diameter.…”

Electron cryotomography of Mycoplasma pneumoniae mutants correlates terminal organelle architectural features and function

“…Such ribbons were absent in wild-type or other mutant tomograms and may represent a terminal organelle substructure that has failed to fold or localize correctly in the absence of the paired rods of the core, which may serve to orient normal core assembly. Perhaps significantly, segmented filamentous structures, sometimes bundled, were also recently described in other bacteria (Dobro et al, 2017).…”

Lipid trafficking across the Gram-negative cell envelope

“…It has long been suggested that the presence of the peptidoglycan layer and the size of the periplasm (15-20 nm) are not compatible with vesicular transport. Although periplasmic vesicular structures have recently been observed in cryo-tomograms, they were largely found in cells showing signs of envelope stress and may have limited physiological relevance (10). It has also been hypothesized that lipid transport can occur in membrane adhesion zones between the IM and the OM (11).…”