scholarly journals The Stress-Active Cell Division Protein ZapE Alters FtsZ Filament Architecture to Facilitate Division in Escherichia coli

2021 ◽  
Vol 12 ◽  
Author(s):  
Eric C. DiBiasio ◽  
Rebecca A. Dickinson ◽  
Catherine E. Trebino ◽  
Colby N. Ferreira ◽  
Josiah J. Morrison ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Cell Division ◽  
High Temperature ◽  
Environmental Stress ◽  
Atp Hydrolysis ◽  
Major Protein ◽  
Bacterial Cells ◽  
Z Ring ◽  
Cell Division Protein

During pathogenic infections, bacterial cells experience environmental stress conditions, including low oxygen and thermal stress. Bacterial cells proliferate during infection and divide by a mechanism characterized by the assembly of a large cytoskeletal structure at the division site called the Z-ring. The major protein constituting the Z-ring is FtsZ, a tubulin homolog and GTPase that utilizes the nucleotide to assemble into dynamic polymers. In Escherichia coli, many cell division proteins interact with FtsZ and modulate Z-ring assembly, while others direct cell wall insertion and peptidoglycan remodeling. Here, we show that ZapE, an ATPase that accumulates during late constriction, directly interacts with FtsZ and phospholipids in vitro. In the presence of adenosine triphosphate (ATP), ZapE induces bundling of GTP-induced FtsZ polymers; however, ZapE also binds FtsZ in the absence of GTP. The ZapE mutant protein ZapE(K84A), which is defective for ATP hydrolysis, also interacts with FtsZ and induces FtsZ filament bundling. In vivo, cultures of zapE deletion cells contain a low percentage of filamentous cells, suggesting that they have a modest division defect; however, they are able to grow when exposed to stress, such as high temperature and limited oxygen. When combined with the chromosomal deletion of minC, which encodes an FtsZ disassembly factor, ΔzapE ΔminC cells experience growth delays that slow proliferation at high temperature and prevent recovery. This synthetic slow growth phenotype after exposure to stress suggests that ZapE may function to ensure proliferation during and after stress, and this is exacerbated when cells are also deleted for minC. Expression of either ZapE or ZapE(K84A) complements the aberrant growth phenotypes in vivo suggesting that the division-associated role of ZapE does not require ZapE ATP hydrolysis. These results support that ZapE is a stress-regulated cell division protein that interacts directly with FtsZ and phospholipids, promoting growth and division after exposure to environmental stress.

scholarly journals Murein (Peptidoglycan) Binding Property of the Essential Cell Division Protein FtsN from Escherichia coli

2004 ◽  
Vol 186 (20) ◽  
pp. 6728-6737 ◽  
Author(s):  
Astrid Ursinus ◽  
Fusinita van den Ent ◽  
Sonja Brechtel ◽  
Miguel de Pedro ◽  
Joachim-Volker Höltje ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Cell Division ◽  
Disulfide Bridge ◽  
Binding Property ◽  
Wild Type ◽  
E Coli ◽  
Specific Affinity ◽  
Cell Division Protein ◽  
Truncated Variants

ABSTRACT The binding of the essential cell division protein FtsN of Escherichia coli to the murein (peptidoglycan) sacculus was studied. Soluble truncated variants of FtsN, including the complete periplasmic part of the protein as well as a variant containing only the C-terminal 77 amino acids, did bind to purified murein sacculi isolated from wild-type cells. FtsN variants lacking this C-terminal region showed reduced or no binding to murein. Binding of FtsN was severely reduced when tested against sacculi isolated either from filamentous cells with blocked cell division or from chain-forming cells of a triple amidase mutant. Binding experiments with radioactively labeled murein digestion products revealed that the longer murein glycan strands (>25 disaccharide units) showed a specific affinity to FtsN, but neither muropeptides, peptides, nor short glycan fragments bound to FtsN. In vivo FtsN could be cross-linked to murein with the soluble disulfide bridge containing cross-linker DTSSP. Less FtsN, but similar amounts of OmpA, was cross-linked to murein of filamentous or of chain-forming cells compared to levels in wild-type cells. Expression of truncated FtsN variants in cells depleted in full-length FtsN revealed that the presence of the C-terminal murein-binding domain was not required for cell division under laboratory conditions. FtsN was present in 3,000 to 6,000 copies per cell in exponentially growing wild-type E. coli MC1061. We discuss the possibilities that the binding of FtsN to murein during cell division might either stabilize the septal region or might have a function unrelated to cell division.

scholarly journals Assembly Dynamics of FtsZ Rings in Bacillus subtilis and Escherichia coli and Effects of FtsZ-Regulating Proteins

2004 ◽  
Vol 186 (17) ◽  
pp. 5775-5781 ◽  
Author(s):  
David E. Anderson ◽  
Frederico J. Gueiros-Filho ◽  
Harold P. Erickson
Keyword(s):  
Escherichia Coli ◽  
Bacillus Subtilis ◽  
Cell Division ◽  
Fluorescent Protein ◽  
Fluorescence Recovery ◽  
Half Time ◽  
Experimental Conditions ◽  
Z Ring

ABSTRACT FtsZ is the major cytoskeletal component of the bacterial cell division machinery. It forms a ring-shaped structure (the Z ring) that constricts as the bacterium divides. Previous in vivo experiments with green fluorescent protein-labeled FtsZ and fluorescence recovery after photobleaching have shown that the Escherichia coli Z ring is extremely dynamic, continually remodeling itself with a half time of 30 s, similar to microtubules in the mitotic spindle. In the present work, under different experimental conditions, we have found that the half time for fluorescence recovery of E. coli Z rings is even shorter (∼9 s). As before, the turnover appears to be coupled to GTP hydrolysis, since the mutant FtsZ84 protein, with reduced GTPase in vitro, showed an ∼3-fold longer half time. We have also extended the studies to Bacillus subtilis and found that this species exhibits equally rapid dynamics of the Z ring (half time, ∼8 s). Interestingly, null mutations of the FtsZ-regulating proteins ZapA, EzrA, and MinCD had only modest effects on the assembly dynamics. This suggests that these proteins do not directly regulate FtsZ subunit exchange in and out of polymers. In B. subtilis, only 30 to 35% of the FtsZ protein was in the Z ring, from which we conclude that a Z ring only 2 or 3 protofilaments thick can function for cell division.

scholarly journals MinC N- and C-Domain Interactions Modulate FtsZ Assembly, Division Site Selection, and MinD-Dependent Oscillation inEscherichia coli

2018 ◽  
Vol 201 (4) ◽  
Author(s):  
Christopher J. LaBreck ◽  
Joseph Conti ◽  
Marissa G. Viola ◽  
Jodi L. Camberg
Keyword(s):  
Escherichia Coli ◽  
Cell Division ◽  
Site Selection ◽  
Fluorescent Protein ◽  
Site Mutation ◽  
Content Type ◽  
Ftsz Ring ◽  
Division Site ◽  
Z Ring

ABSTRACTThe Min system inEscherichia coli, consisting of MinC, MinD, and MinE proteins, regulates division site selection by preventing assembly of the FtsZ-ring (Z-ring) and exhibits polar oscillationin vivo. MinC antagonizes FtsZ polymerization, andin vivo, the cellular location of MinC is controlled by a direct association with MinD at the membrane. To further understand the interactions of MinC with FtsZ and MinD, we performed a mutagenesis screen to identify substitutions inminCthat are associated with defects in cell division. We identified amino acids in both the N- and C-domains of MinC that are important for direct interactions with FtsZ and MinDin vitro, as well as mutations that modify the observedin vivooscillation of green fluorescent protein (GFP)-MinC. Our results indicate that there are two distinct surface-exposed sites on MinC that are important for direct interactions with FtsZ, one at a cleft on the surface of the N-domain and a second on the C-domain that is adjacent to the MinD interaction site. Mutation of either of these sites leads to slower oscillation of GFP-MinCin vivo, although the MinC mutant proteins are still capable of a direct interaction with MinD in phospholipid recruitment assays. Furthermore, we demonstrate that interactions between FtsZ and both sites of MinC identified here are important for assembly of FtsZ-MinC-MinD complexes and that the conserved C-terminal end of FtsZ is not required for MinC-MinD complex formation with GTP-dependent FtsZ polymers.IMPORTANCEBacterial cell division proceeds through the coordinated assembly of the FtsZ-ring, or Z-ring, at the site of division. Assembly of the Z-ring requires polymerization of FtsZ, which is regulated by several proteins in the cell. InEscherichia coli, the Min system, which contains MinC, MinD, and MinE proteins, exhibits polar oscillation and inhibits the assembly of FtsZ at nonseptal locations. Here, we identify regions on the surface of MinC that are important for contacting FtsZ and destabilizing FtsZ polymers.

scholarly journals Coregulated assembly of actin-like FtsA polymers with FtsZ during Z-ring formation and division in Escherichia coli

2021 ◽  
Author(s):  
Josiah J. Morrison ◽  
Joseph Conti ◽  
Jodi L. Camberg
Keyword(s):  
Escherichia Coli ◽  
Atp Hydrolysis ◽  
Direct Interaction ◽  
Ring Formation ◽  
Phospholipid Vesicle ◽  
E Coli ◽  
C Terminus ◽  
Z Ring

AbstractIn Escherichia coli, the actin homolog FtsA localizes the cell division machinery, beginning with the Z-ring, to the cytoplasmic membrane through direct interaction with FtsZ. FtsZ polymers are first to assemble at the Z-ring at midcell, where they direct constriction and septation. While FtsZ polymerization is critical for establishing a functional Z-ring that leads to constriction, the assembly state of FtsA and the role of FtsA ATP utilization during division in E. coli remain unclear. Here, we show that ATP hydrolysis, FtsZ interaction, and phospholipid vesicle remodeling by FtsA are impaired by a substitution mutation at the predicted active site for hydrolysis. This mutation, Glu 14 to Arg, also impairs Z-ring assembly and division in vivo. To further investigate the role of phospholipid engagement and ATP utilization in regulating FtsA function, we characterized a truncated E. coli FtsA variant, FtsA(ΔMTS), which lacks the region at the C-terminus important for engaging the membrane and is defective for ATP hydrolysis. We show that E. coli FtsA(ΔMTS) forms ATP-dependent actin-like filaments and assembly is antagonized by FtsZ. Polymerization of FtsZ with GTP, or a non-hydrolyzable analog, blocks inhibition of ATP-dependent FtsA assembly, and instead favors coassembly of stable FtsA/FtsZ polymers. In the cell, FtsA/FtsZ coassembly is favored at midcell, where FtsZ polymerizes, and inhibited at regions where FtsZ polymers are destabilized by regulators, such as MinC at the poles or SlmA at the nucleoid. We show that MinC prevents recruitment of FtsZ, via FtsA, to phospholipids, suggesting that local interactions of MinC with FtsZ block membrane tethering and uncouple the Z-ring from its major membrane contact. During Z-ring formation, the coassembly of FtsZ polymers with FtsA is coordinated and is a critical early step in division. This step also serves as a checkpoint by responding to the suite of FtsZ assembly regulators in the cell that modulate Z-ring position and dynamics prior to initiating cell wall synthesis.

scholarly journals A conserved cell division protein directly regulates FtsZ dynamics in filamentous and unicellular actinobacteria

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Félix Ramos-León ◽  
Matthew J Bush ◽  
Joseph W Sallmen ◽  
Govind Chandra ◽  
Jake Richardson ◽  
...  
Keyword(s):  
Cell Division ◽  
Bacterial Cell ◽  
Mycobacterium Smegmatis ◽  
Bacterial Cell Division ◽  
Z Ring ◽  
Cell Division Protein ◽  
Ring Architecture ◽  
Contractile Structure

Bacterial cell division is driven by the polymerization of the GTPase FtsZ into a contractile structure, the so-called Z-ring. This essential process involves proteins that modulate FtsZ dynamics and hence the overall Z-ring architecture. Actinobacteria like Streptomyces and Mycobacterium lack known key FtsZ-regulators. Here we report the identification of SepH, a conserved actinobacterial protein that directly regulates FtsZ dynamics. We show that SepH is crucially involved in cell division in Streptomyces venezuelae and that it binds FtsZ via a conserved helix-turn-helix motif, stimulating the assembly of FtsZ protofilaments. Comparative in vitro studies using the SepH homolog from Mycobacterium smegmatis further reveal that SepH can also bundle FtsZ protofilaments, indicating an additional Z-ring stabilizing function in vivo. We propose that SepH plays a crucial role at the onset of cytokinesis in actinobacteria by promoting the assembly of FtsZ filaments into division-competent Z-rings that can go on to mediate septum synthesis.

Dynamic FtsZ polymerization is sensitive to the GTP to GDP ratio and can be maintained at steady state using a GTP-regeneration system

Microbiology ◽  
2003 ◽  
Vol 149 (8) ◽  
pp. 2235-2242 ◽  
Author(s):  
Elaine Small ◽  
Stephen G. Addinall
Keyword(s):  
Steady State ◽  
Cell Division ◽  
Dynamic Properties ◽  
Bacterial Cell Division ◽  
Regeneration System ◽  
Z Ring ◽  
Cell Division Protein ◽  
Hydrolysis Of

In vitro polymerization of the essential bacterial cell division protein FtsZ, in the presence of GTP, is rapid and transient due to its efficient binding and hydrolysis of GTP. In contrast, the in vivo polymeric FtsZ structure which drives cell division – the Z-ring – is present in cells for extended periods of time whilst undergoing constant turnover of FtsZ. It is demonstrated that dynamic polymerization of Escherichia coli FtsZ in vitro is sensitive to the ratio of GTP to GDP concentration. Increase of GDP concentration in the presence of a constant GTP concentration reduces both the duration of FtsZ polymerization and the initial light-scattering maximum which occurs upon addition of GTP. It is also demonstrated that by use of a GTP-regeneration system, polymers of FtsZ can be maintained in a steady state for up to 85 min, while preserving their dynamic properties. The authors therefore present the use of a GTP-regeneration system for FtsZ polymerization as an assay more representative of the in vivo situation, where FtsZ polymers are subject to a constant, relatively high GTP to GDP ratio.

scholarly journals FtsW Is a Dispensable Cell Division Protein Required for Z-Ring Stabilization during Sporulation Septation in Streptomyces coelicolor

2008 ◽  
Vol 190 (16) ◽  
pp. 5555-5566 ◽  
Author(s):  
Bhavesh V. Mistry ◽  
Ricardo Del Sol ◽  
Chris Wright ◽  
Kim Findlay ◽  
Paul Dyson
Keyword(s):  
Cell Division ◽  
Streptomyces Coelicolor ◽  
Cross Wall ◽  
Penicillin Binding Protein ◽  
Aerial Hyphae ◽  
Class B ◽  
Z Ring ◽  
Cell Division Protein

ABSTRACT The conserved rodA and ftsW genes encode polytopic membrane proteins that are essential for bacterial cell elongation and division, respectively, and each gene is invariably linked with a cognate class B high-molecular-weight penicillin-binding protein (HMW PBP) gene. Filamentous differentiating Streptomyces coelicolor possesses four such gene pairs. Whereas rodA, although not its cognate HMW PBP gene, is essential in these bacteria, mutation of SCO5302 or SCO2607 (sfr) caused no gross changes to growth and septation. In contrast, disruption of either ftsW or the cognate ftsI gene blocked the formation of sporulation septa in aerial hyphae. The inability of spiral polymers of FtsZ to reorganize into rings in aerial hyphae of these mutants indicates an early pivotal role of an FtsW-FtsI complex in cell division. Concerted assembly of the complete divisome was unnecessary for Z-ring stabilization in aerial hyphae as ftsQ mutants were found to be blocked at a later stage in cell division, during septum closure. Complete cross wall formation occurred in vegetative hyphae in all three fts mutants, indicating that the typical bacterial divisome functions specifically during nonessential sporulation septation, providing a unique opportunity to interrogate the function and dependencies of individual components of the divisome in vivo.

scholarly journals A conserved cell division protein directly regulates FtsZ dynamics in filamentous and unicellular actinobacteria

2020 ◽  
Author(s):  
Felix Ramos-Léon ◽  
Matthew J. Bush ◽  
Joseph W. Sallmen ◽  
Govind Chandra ◽  
Jake Richardson ◽  
...  
Keyword(s):  
Cell Division ◽  
Bacterial Cell ◽  
Crucial Role ◽  
Bacterial Cell Division ◽  
Z Ring ◽  
Cell Division Protein ◽  
Ring Architecture ◽  
Contractile Structure

AbstractBacterial cell division is driven by the polymerization of the GTPase FtsZ into a contractile structure, the so-called Z-ring. This essential process involves proteins that modulate FtsZ dynamics and hence the overall Z-ring architecture. Actinobacteria, like Streptomyces and Mycobacterium lack known key FtsZ-regulators. Here we report the identification of SepH, a conserved actinobacterial protein that directly regulates FtsZ dynamics. We show that SepH is crucially involved in cell division in Streptomyces and that it binds FtsZ via a conserved helix-turn-helix motif, stimulating the assembly of FtsZ protofilaments. Comparative in vitro studies using the SepH homolog from Mycobacterium further reveal that SepH can also bundle FtsZ protofilaments, indicating an additional Z-ring stabilizing function in vivo. We propose that SepH plays a crucial role at the onset of cytokinesis in actinobacteria by promoting the rapid assembly of FtsZ filaments into division-competent Z-rings that can go on to mediate septum synthesis.

scholarly journals Crystal structure of the Z-ring associated cell division protein ZapC from Escherichia coli

FEBS Letters ◽  
2015 ◽  
Vol 589 (24PartB) ◽  
pp. 3822-3828 ◽  
Author(s):  
Cristina Ortiz ◽  
Danguole Kureisaite-Ciziene ◽  
Florian Schmitz ◽  
Stephen H. McLaughlin ◽  
Miguel Vicente ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Escherichia Coli ◽  
Cell Division ◽  
Z Ring ◽  
Cell Division Protein
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