Biology subjects

Nussbaum, P.

Publications and source records attributed to Nussbaum, P..

3 recordsLinked to original sources

bioRxiv2020

Archaeal SepF is essential for cell division in Haloferax volcanii

Bacterial cell division has been studied for decades but reports on the different archaeal cell division systems are rare. In many archaea, cell division depends on the tubulin homolog FtsZ, but further components of the divisome in these archaea are unknown. The halophilic archaeon Haloferax volcanii encodes two FtsZ homologs with different functions in cell division and a putative SepF homolog. In bacteria, SepF is part of the divisome and is recruited early to the FtsZ ring, where it most likely stimulates FtsZ ring formation. H. volcanii SepF co-localized with FtsZ1 and FtsZ2 at midcell. Overexpression of SepF had no effect on cell morphology, but no sepF deletion mutants could be generated. SepF depletion led to a severe cell division defect, resulting in cells with a strongly increased size. Overexpression of FtsZ1- and FtsZ2-GFP in SepF-depleted cells resulted in filamentous cells with an increasing number of FtsZ1 rings depending on the cell length, whereas FtsZ2 rings were not increased. Pull-down assays with HA-tagged SepF identified an interaction with FtsZ2 but not with FtsZ1. Archaeal SepF homologs lack the conserved glycine residue important for polymerization in bacteria and the H. volcanii SepF was purified as a dimer, suggesting that in contrast to the bacterial SepF homologs, polymerization does not seem to be important for its function. A model is proposed where first the FtsZ1 ring is formed and where SepF recruits FtsZ2 to the FtsZ1 ring, resulting in the formation of the FtsZ2 ring. This study provides important novel insights into cell division in archaea and shows that SepF is an important part of the divisome in FtsZ containing archaea.

microbiology↗

bioRxiv2020

Analysis of cell-cell bridges in Haloferax volcanii using Electron cryo-tomography reveal a continuous cytoplasm and S-layer

Halophilic archaea 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 cryotomography and fluorescence microscopy to visualize cells forming these intercellular bridges. Electron cryo-tomography showed that the observed bridges were enveloped by an 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 {micro}m. These results shed light on the process of haloarchaeal mating and highlight further mechanistic questions.

microbiology↗

bioRxiv2020

An oscillating MinD protein determines the cellular positioning of the motility machinery in archaea

MinD proteins are well studied in rod-shaped bacteria such as E. coli, where they display self-organized pole-to-pole oscillations that are important for correct positioning of the Z-ring at mid-cell for cell division. Archaea also encode proteins belonging to the MinD family, but their functions are unknown. MinD homologous proteins were found to be widespread in Euryarchaeota and form a sister group to the bacterial MinD family, distinct from the ParA and other related ATPase families. We aimed to identify the function of four archaeal MinD proteins in the model archaeon Haloferax volcanii. Deletion of the minD genes did not cause cell division or size defects, and the Z-ring was still correctly positioned. Instead, one of the mutations ({Delta}minD4) reduced swimming motility, and hampered the correct formation of motility machinery at the cell poles. In {Delta}minD4 cells, there is reduced formation of the motility structure and chemosensory arrays, which are essential for signal transduction. In bacteria, several members of the ParA family can position the motility structure and chemosensory arrays via binding to a landmark protein, and consequently these proteins do not oscillate along the cell axis. However, GFP-MinD4 displayed pole-to-pole oscillation and formed polar patches or foci in H. volcanii. The MinD4 membrane targeting sequence (MTS), homologous to the bacterial MinD MTS, was essential for the oscillation. Surprisingly, MinD4 ATPase domain point-mutations did not block oscillation, but they failed to form pole-patches. Thus, MinD4 from H. volcanii combines traits of different bacterial ParA/MinD proteins.

microbiology↗