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FtsZ does not initiate membrane constriction at the onset of division.

ABSTRACT: The source of constriction required for division of a bacterial cell remains enigmatic. FtsZ is widely believed to be a key player, because in vitro experiments indicate that it can deform liposomes when membrane tethered. However in vivo evidence for such a role has remained elusive as it has been challenging to distinguish the contribution of FtsZ from that of peptidoglycan-ingrowth. To differentiate between these two possibilities we studied the early stages of division in Escherichia coli, when FtsZ is present at the division site but peptidoglycan synthesizing enzymes such as FtsI and FtsN are not. Our approach was to use correlative cryo-fluorescence and cryo-electron microscopy (cryo-CLEM) to monitor the localization of fluorescently labeled FtsZ, FtsI or FtsN correlated with the septal ultra-structural geometry in the same cell. We noted that the presence of FtsZ at the division septum is not sufficient to deform membranes. This observation suggests that, although FtsZ can provide a constrictive force, the force is not substantial at the onset of division. Conversely, the presence of FtsN always correlated with membrane invagination, indicating that allosteric activation of peptidoglycan ingrowth is the trigger for constriction of the cell envelope during cell division in E. coli.

SUBMITTER: Daley DO

PROVIDER: S-EPMC5016829 | biostudies-other | 2016

REPOSITORIES: biostudies-other

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FtsZ does not initiate membrane constriction at the onset of division.

Daley Daniel O DO Skoglund Ulf U Söderström Bill B

Scientific reports 20160909

The source of constriction required for division of a bacterial cell remains enigmatic. FtsZ is widely believed to be a key player, because in vitro experiments indicate that it can deform liposomes when membrane tethered. However in vivo evidence for such a role has remained elusive as it has been challenging to distinguish the contribution of FtsZ from that of peptidoglycan-ingrowth. To differentiate between these two possibilities we studied the early stages of division in Escherichia coli, w ...[more]

PMID: 27609565

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Project description:To divide, bacteria must constrict their membranes against significant force from turgor pressure. A tubulin homolog, FtsZ, is thought to drive constriction, but how FtsZ filaments might generate constrictive force in the absence of motor proteins is not well understood. There are two predominant models in the field. In one, FtsZ filaments overlap to form complete rings around the circumference of the cell, and attractive forces cause filaments to slide past each other to maximize lateral contact. In the other, filaments exert force on the membrane by a GTP-hydrolysis-induced switch in conformation from straight to bent. Here, we developed software, ZCONSTRICT, for quantitative three-dimensional (3D) simulations of Gram-negative bacterial cell division to test these two models and identify critical conditions required for them to work. We find that the avidity of any kind of lateral interactions quickly halts the sliding of filaments, so a mechanism such as depolymerization or treadmilling is required to sustain constriction by filament sliding. For filament bending, we find that a mechanism such as the presence of a rigid linker is required to constrain bending to within the division plane and maintain the distance observed in vivo between the filaments and the membrane. Of these two models, only the filament bending model is consistent with our lab's recent observation of constriction associated with a single, short FtsZ filament.IMPORTANCE FtsZ is thought to generate constrictive force to divide the cell, possibly via one of two predominant models in the field. In one, FtsZ filaments overlap to form complete rings which constrict as filaments slide past each other to maximize lateral contact. In the other, filaments exert force on the membrane by switching conformation from straight to bent. Here, we developed software, ZCONSTRICT, for three-dimensional (3D) simulations to test these two models. We find that a mechanism such as depolymerization or treadmilling are required to sustain constriction by filament sliding. For filament bending, we find that a mechanism that constrains bending to within the division plane is required to maintain the distance observed in vivo between the filaments and the membrane.

Project description:Bacteria such as Escherichia coli divide by organizing filaments of FtsZ, a tubulin homolog that assembles into dynamic treadmilling membrane-associated protein filaments at the cell midpoint. FtsA and ZipA proteins are required to tether these filaments to the inner face of the cytoplasmic membrane, and loss of either tether is lethal. ZipA from E. coli and other closely related species harbors a long linker region that connects the essential N-terminal transmembrane domain to the C-terminal globular FtsZ-binding domain, and part of this linker includes a P/Q-rich peptide that is predicted to be intrinsically disordered. We found unexpectedly that several large deletions of the ZipA linker region, including the entire P/Q rich peptide, had no effect on cell division under normal conditions. However, we found that the loss of the P/Q region made cells more resistant to excess levels of FtsA and more sensitive to conditions that displaced FtsA from FtsZ. FtsA also harbors a short ∼20-residue peptide linker that connects the main globular domain with the C-terminal amphipathic helix that is important for membrane binding. In analogy with ZipA, deletion of 11 of the central residues in the FtsA linker had little effect on FtsA function in cell division.IMPORTANCEEscherichia coli cells divide using a cytokinetic ring composed of polymers of the tubulin-like FtsZ. To function properly, these polymers must attach to the inner surface of the cytoplasmic membrane via two essential membrane-associated tethers, FtsA and ZipA. Both FtsA and ZipA contain peptide linkers that connect their membrane-binding domains with their FtsZ-binding domains. Although they are presumed to be crucial for cell division activity, the importance of these linkers has not yet been rigorously tested. Here, we show that large segments of these linkers can be removed with few consequences for cell division, although several subtle defects were uncovered. Our results suggest that ZipA, in particular, can function in cell division without an extended linker.

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FtsZ does not initiate membrane constriction at the onset of division.

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FtsZ does not initiate membrane constriction at the onset of division.

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