Bacteria and archaea exhibit tactical behavior and can move up and down chemical gradients. This tactical behavior relies on a motility structure, which is guided by a chemosensory system. Environmental signals are sensed by membrane-inserted chemosensory receptors that are organized in large ordered arrays. While the cellular positioning of the chemotaxis machinery and that of the flagellum have been studied in detail in bacteria, we have little knowledge about the localization of such macromolecular assemblies in archaea. Although the archaeal motility structure, the archaellum, is fundamentally different from the flagellum, archaea have received the chemosensory machinery from bacteria and have connected this system with the archaellum. Here, we applied a combination of time-lapse imaging and fluorescence and electron microscopy using the model euryarchaeon Haloferax volcanii and found that archaella were specifically present at the cell poles of actively dividing rod-shaped cells. The chemosensory arrays also had a polar preference, but in addition, several smaller arrays moved freely in the lateral membranes. In the stationary phase, rod-shaped cells became round and chemosensory arrays were disassembled. The positioning of archaella and that of chemosensory arrays are not interdependent and likely require an independent form of positioning machinery. This work showed that, in the rod-shaped haloarchaeal cells, the positioning of the archaellum and of the chemosensory arrays is regulated in time and in space. These insights into the cellular organization of H. volcanii suggest the presence of an active mechanism responsible for the positioning of macromolecular protein complexes in archaea. IMPORTANCE Archaea are ubiquitous single cellular microorganisms that play important ecological roles in nature. The intracellular organization of archaeal cells is among the unresolved mysteries of archaeal biology. With this work, we show that cells of haloarchaea are polarized. The cellular positioning of proteins involved in chemotaxis and motility is spatially and temporally organized in these cells. This suggests the presence of a specific mechanism responsible for the positioning of macromolecular protein complexes in archaea.

In most bacteria, cell division depends on the tubulin homolog FtsZ and other proteins, such as SepF, that form a complex termed the divisome. Cell division also depends on FtsZ in many archaea, but other components of the divisome are unknown. Here, we demonstrate that a SepF homolog plays important roles in cell division in Haloferax volcanii, a halophilic archaeon that is known to have two FtsZ homologs with slightly different functions (FtsZ1 and FtsZ2). SepF co-localizes with both FtsZ1 and FtsZ2 at midcell. Attempts to generate a sepF deletion mutant were unsuccessful, suggesting an essential role. Indeed, SepF depletion leads to severe cell division defects and formation of large cells. Overexpression of FtsZ1-GFP or FtsZ2-GFP in SepF-depleted cells results in formation of filamentous cells with a high number of FtsZ1 rings, while the number of FtsZ2 rings is not affected. Pull-down assays support that SepF interacts with FtsZ2 but not with FtsZ1, although SepF appears delocalized in the absence of FtsZ1. Archaeal SepF homologs lack a glycine residue known to be important for polymerization and function in bacteria, and purified H. volcanii SepF forms dimers, suggesting that polymerization might not be important for the function of archaeal SepF.

Thermoacidophilic archaea belonging to the order Sulfolobales thrive in extreme biotopes, such as sulfuric hot springs and ore deposits. These microorganisms have been model systems for understanding life in extreme environments, as well as for probing the evolution of both molecular genetic processes and central metabolic pathways. Thermoacidophiles, such as the Sulfolobales, use typical microbial responses to persist in hot acid (e.g. motility, stress response, biofilm formation), albeit with some unusual twists. They also exhibit unique physiological features, including iron and sulfur chemolithoautotrophy, that differentiate them from much of the microbial world. Although first discovered more than 50 years ago, it was not until recently that genome sequence data and facile genetic tools have been developed for species in the Sulfolobales. These advances have not only opened up ways to further the probe novel features of these microbes, but have also paved the way for potential biotechnological applications. Discussed here are the nuances of the thermoacidophilic lifestyle of the Sulfolobales, including their evolutionary placement, cell biology, survival strategies, genetic tools, metabolic processes, and physiological attributes together with how these characteristics make thermoacidophiles ideal platforms for specialized industrial processes.

N-glycosylation is a post-translational modification that occurs across evolution. In the thermoacidophilic archaea Sulfolobus acidocaldarius, glycoproteins are modified by an N-linked tri-branched hexasaccharide reminiscent of the N-glycans assembled in Eukarya. Previously, hexose-bearing dolichol phosphate was detected in a S. acidocaldarius Bligh-Dyer lipid extract. Here, we used a specialized protocol for extracting lipid-linked oligosaccharides to detect a dolichol pyrophosphate bearing the intact hexasaccharide, as well as its biosynthetic intermediates. Furthermore, evidence for N-glycosylation of two S. acidocaldarius proteins by the same hexasaccharide and its derivatives was collected. These findings thus provide novel insight into archaeal N-glycosylation.

Halophilic archaea have been proposed to exchange DNA and proteins using a fusion-based mating mechanism. Scanning electron microscopy previously suggested that mating involves an intermediate state, where cells are connected by an intercellular bridge. To better understand this process, we used electron cryo-tomography (cryoET) and fluorescence microscopy to visualize cells forming these intercellular bridges. CryoET showed that the observed bridges were enveloped by an surface layer (S-layer) and connected mating cells via a continuous cytoplasm. Macromolecular complexes like ribosomes and unknown thin filamentous helical structures were visualized in the cytoplasm inside the bridges, demonstrating that these bridges can facilitate exchange of cellular components. We followed formation of a cell–cell bridge by fluorescence time-lapse microscopy between cells at a distance of 1.5 μm. These results shed light on the process of haloarchaeal mating and highlight further mechanistic questions.