Microfluidic trapping of vesicles reveals membrane-tension dependent FtsZ cytoskeletal re-organisation

AbstractThe geometry of reaction compartments can affect the outcome of chemical reactions. Synthetic biology commonly uses giant unilamellar vesicles (GUVs) to generate cell-sized, membrane-bound reaction compartments. However, these liposomes are always spherical due to surface area minimization. Here, we have developed a microfluidic chip to trap and reversibly deform GUVs into rod- or cigar-like shapes, including a constriction site in the trap mimicking the membrane furrow in cell division. When we introduce into these GUVs the bacterial tubulin homologue FtsZ, the primary protein of the bacterial Z ring, we find that FtsZ organization changes from dynamic rings to elongated filaments upon GUV deformation, and that these FtsZ filaments align preferentially with the short GUV axis, in particular at the membrane neck. In contrast, pulsing Min oscillations in GUVs remained largely unaffected. We conclude that microfluidic traps are a useful tool for deforming GUVs into non-spherical membrane shapes, akin to those seen in cell division, and for investigating the effect of confinement geometry on biochemical reactions, such as protein filament self-organization.

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FtsZ assembles the bacterial cell division machinery by a diffusion-and-capture mechanism

10.1101/485656 ◽

2018 ◽

Cited By ~ 1

Author(s):

Natalia Baranova ◽

Philipp Radler ◽

Víctor M. Hernández-Rocamora ◽

Carlos Alfonso ◽

Mar López-Pelegrín ◽

...

Keyword(s):

Cell Division ◽

Bacterial Cell ◽

Bacterial Cell Division ◽

Peptidoglycan Synthesis ◽

Division Site ◽

Spatiotemporal Information ◽

Capture Mechanism ◽

Z Ring ◽

Membrane Bound ◽

Transient Interactions

AbstractThe mechanism of bacterial cell division is largely unknown. The protein machinery performing cell division is organized by FtsZ, a tubulin-homolog that forms treadmilling filaments at the cell division site. Treadmilling is thought to actively move proteins around the cell thereby distributing peptidoglycan synthesis to make two new cell poles. To understand this process, we reconstituted part of the bacterial cell division machinery using the purified components FtsZ, FtsA and truncated transmembrane proteins essential for cell division. We found that membrane-bound cytosolic peptides of FtsN and FtsQ co-migrated with treadmilling FtsZ-FtsA filaments. Remarkably, rather than moving in a directed fashion, individual peptides followed FtsZ filaments by a diffusion-and-capture mechanism. Our work provides a mechanism for how the Z-ring dynamically recruits divisome proteins and highlights the importance of transient interactions for the self-organization of complex biological structures. We propose that this mechanism is used more widely to organize and transmit spatiotemporal information in living cells.One Sentence SummaryFtsZ treadmilling assembles bacterial division machinery by diffusion-and-capture mechanism.

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Cell Division in Bacillus subtilis: FtsZ and FtsA Association Is Z-Ring Independent, and FtsA Is Required for Efficient Midcell Z-Ring Assembly

Journal of Bacteriology ◽

10.1128/jb.187.18.6536-6544.2005 ◽

2005 ◽

Vol 187 (18) ◽

pp. 6536-6544 ◽

Cited By ~ 63

Author(s):

S. O. Jensen ◽

L. S. Thompson ◽

E. J. Harry

Keyword(s):

Bacillus Subtilis ◽

Cell Division ◽

Ring Formation ◽

Division Site ◽

Ring Assembly ◽

Synchronous Cell ◽

Z Ring ◽

Membrane Bound ◽

Chemical Cross Linking

ABSTRACT The earliest stage in cell division in bacteria is the assembly of a Z ring at the division site at midcell. Other division proteins are also recruited to this site to orchestrate the septation process. FtsA is a cytosolic division protein that interacts directly with FtsZ. Its function remains unknown. It is generally believed that FtsA localization to the division site occurs immediately after Z-ring formation or concomitantly with it and that FtsA is responsible for recruiting the later-assembling membrane-bound division proteins to the division site. Here, we report the development of an in vivo chemical cross-linking assay to examine the association between FtsZ and FtsA in Bacillus subtilis cells. We subsequently use this assay in a synchronous cell cycle to show that these two proteins can interact prior to Z-ring formation. We further show that in a B. subtilis strain containing an ftsA deletion, FtsZ localized at regular intervals along the filament but the majority of Z rings were abnormal. FtsA in this organism is therefore critical for the efficient formation of functional Z rings. This is the first report of abnormal Z-ring formation resulting from the loss of a single septation protein. These results suggest that in this organism, and perhaps others, FtsA ensures recruitment of the membrane-bound division proteins by ensuring correct formation of the Z ring.

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Thai Students’ Decision Making in Societal Issue of Surface Area and Concentrated Solutions as a Factor in the Rate of Chemical Reactions

Mediterranean Journal of Social Sciences ◽

10.5901/mjss.2015.v6n6s1p56 ◽

2015 ◽

Cited By ~ 1

Author(s):

Jeeruwan Chattabud ◽

Paisan Suwannoi ◽

Tawee Sranamkam ◽

Chokchai Yuenyong

Keyword(s):

Decision Making ◽

Surface Area ◽

Chemical Reactions ◽

Concentrated Solutions ◽

Societal Issue ◽

Thai Students

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Improving cell-free glycoprotein synthesis by characterizing and enriching native membrane vesicles

Nature Communications ◽

10.1038/s41467-021-22329-3 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Jasmine M. Hershewe ◽

Katherine F. Warfel ◽

Shaelyn M. Iyer ◽

Justin A. Peruzzi ◽

Claretta J. Sullivan ◽

...

Keyword(s):

Gene Expression ◽

Escherichia Coli ◽

Synthetic Biology ◽

Membrane Protein ◽

Membrane Vesicles ◽

Processing Methods ◽

Glycoprotein Synthesis ◽

On Demand ◽

Free Gene ◽

Membrane Bound

AbstractCell-free gene expression (CFE) systems from crude cellular extracts have attracted much attention for biomanufacturing and synthetic biology. However, activating membrane-dependent functionality of cell-derived vesicles in bacterial CFE systems has been limited. Here, we address this limitation by characterizing native membrane vesicles in Escherichia coli-based CFE extracts and describing methods to enrich vesicles with heterologous, membrane-bound machinery. As a model, we focus on bacterial glycoengineering. We first use multiple, orthogonal techniques to characterize vesicles and show how extract processing methods can be used to increase concentrations of membrane vesicles in CFE systems. Then, we show that extracts enriched in vesicle number also display enhanced concentrations of heterologous membrane protein cargo. Finally, we apply our methods to enrich membrane-bound oligosaccharyltransferases and lipid-linked oligosaccharides for improving cell-free N-linked and O-linked glycoprotein synthesis. We anticipate that these methods will facilitate on-demand glycoprotein production and enable new CFE systems with membrane-associated activities.

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MinD directly interacting with FtsZ at the H10 helix suggests a model for robust activation of MinC to destabilize FtsZ polymers

Biochemical Journal ◽

10.1042/bcj20170357 ◽

2017 ◽

Vol 474 (18) ◽

pp. 3189-3205 ◽

Cited By ~ 6

Author(s):

Ashoka Chary Taviti ◽

Tushar Kant Beuria

Keyword(s):

Cell Division ◽

Single Point ◽

Point Mutations ◽

Direct Interaction ◽

Ring Formation ◽

Z Ring ◽

Single Point Mutations ◽

Biochemical Analyses ◽

Ftsz Assembly ◽

The Mind

Cell division in bacteria is a highly controlled and regulated process. FtsZ, a bacterial cytoskeletal protein, forms a ring-like structure known as the Z-ring and recruits more than a dozen other cell division proteins. The Min system oscillates between the poles and inhibits the Z-ring formation at the poles by perturbing FtsZ assembly. This leads to an increase in the FtsZ concentration at the mid-cell and helps in Z-ring positioning. MinC, the effector protein, interferes with Z-ring formation through two different mechanisms mediated by its two domains with the help of MinD. However, the mechanism by which MinD triggers MinC activity is not yet known. We showed that MinD directly interacts with FtsZ with an affinity stronger than the reported MinC–FtsZ interaction. We determined the MinD-binding site of FtsZ using computational, mutational and biochemical analyses. Our study showed that MinD binds to the H10 helix of FtsZ. Single-point mutations at the charged residues in the H10 helix resulted in a decrease in the FtsZ affinity towards MinD. Based on our findings, we propose a novel model for MinCD–FtsZ interaction, where MinD through its direct interaction with FtsZ would trigger MinC activity to inhibit FtsZ functions.

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Biochemistry: A Very Short Introduction

10.1093/actrade/9780198833871.001.0001 ◽

2021 ◽

Author(s):

Mark Lorch

Keyword(s):

Synthetic Biology ◽

Chemical Reactions ◽

Biochemical Reactions ◽

Short Introduction ◽

Chemical Structures ◽

Living Things ◽

Historical Figures ◽

Key Concepts ◽

Current Science

Biochemistry: A Very Short Introduction discusses the key concepts of biochemistry, as well as the historical figures in the field and the molecules they studied. From bacteria to humans, all living things are composed of cells of one type or another, all of which have fundamentally the same chemistry. Biochemistry is the study of the chemical reactions within these cells; the molecules that are created, manipulated, and destroyed as a result of them; and the chemical structures such as DNA on which these biochemical reactions take place. This VSI considers the current science and innovations in the field. It also looks at the interaction between biochemistry, biotechnology, and synthetic biology.

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Faculty Opinions recommendation of Single-molecule imaging reveals that Z-ring condensation is essential for cell division in Bacillus subtilis.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.739766772.793584640 ◽

2021 ◽

Author(s):

Erin Goley

Keyword(s):

Bacillus Subtilis ◽

Cell Division ◽

Single Molecule ◽

Single Molecule Imaging ◽

Z Ring ◽

Ring Condensation

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Loss of PodJ in Agrobacterium tumefaciens Leads to Ectopic Polar Growth, Branching, and Reduced Cell Division

Journal of Bacteriology ◽

10.1128/jb.00198-16 ◽

2016 ◽

Vol 198 (13) ◽

pp. 1883-1891 ◽

Cited By ~ 13

Author(s):

James C. Anderson-Furgeson ◽

John R. Zupan ◽

Romain Grangeon ◽

Patricia C. Zambryski

Keyword(s):

Cell Cycle ◽

Cell Division ◽

Agrobacterium Tumefaciens ◽

Cell Envelope ◽

Critical Factor ◽

Polar Growth ◽

Content Type ◽

Growth Pole ◽

Envelope Material ◽

Z Ring

ABSTRACTAgrobacterium tumefaciensis a rod-shaped Gram-negative bacterium that elongates by unipolar addition of new cell envelope material. Approaching cell division, the growth pole transitions to a nongrowing old pole, and the division site creates new growth poles in sibling cells. TheA. tumefacienshomolog of theCaulobacter crescentuspolar organizing protein PopZ localizes specifically to growth poles. In contrast, theA. tumefacienshomolog of theC. crescentuspolar organelle development protein PodJ localizes to the old pole early in the cell cycle and accumulates at the growth pole as the cell cycle proceeds. FtsA and FtsZ also localize to the growth pole for most of the cell cycle prior to Z-ring formation. To further characterize the function of polar localizing proteins, we created a deletion ofA. tumefacienspodJ(podJAt). ΔpodJAtcells display ectopic growth poles (branching), growth poles that fail to transition to an old pole, and elongated cells that fail to divide. In ΔpodJAtcells,A. tumefaciensPopZ-green fluorescent protein (PopZAt-GFP) persists at nontransitioning growth poles postdivision and also localizes to ectopic growth poles, as expected for a growth-pole-specific factor. Even though GFP-PodJAtdoes not localize to the midcell in the wild type, deletion ofpodJAtimpacts localization, stability, and function of Z-rings as assayed by localization of FtsA-GFP and FtsZ-GFP. Z-ring defects are further evidenced by minicell production. Together, these data indicate that PodJAtis a critical factor for polar growth and that ΔpodJAtcells display a cell division phenotype, likely because the growth pole cannot transition to an old pole.IMPORTANCEHow rod-shaped prokaryotes develop and maintain shape is complicated by the fact that at least two distinct species-specific growth modes exist: uniform sidewall insertion of cell envelope material, characterized in model organisms such asEscherichia coli, and unipolar growth, which occurs in several alphaproteobacteria, includingAgrobacterium tumefaciens. Essential components for unipolar growth are largely uncharacterized, and the mechanism constraining growth to one pole of a wild-type cell is unknown. Here, we report that the deletion of a polar development gene,podJAt, results in cells exhibiting ectopic polar growth, including multiple growth poles and aberrant localization of cell division and polar growth-associated proteins. These data suggest that PodJAtis a critical factor in normal polar growth and impacts cell division inA. tumefaciens.

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Study of the pozzolanic activity and hydration products of cement pastes with addition of natural zeolites

Clay Minerals ◽

10.1180/claymin.2011.046.2.241 ◽

2011 ◽

Vol 46 (2) ◽

pp. 241-250 ◽

Cited By ~ 8

Author(s):

V. Lilkov ◽

O. Petrov ◽

V. Petkova ◽

N. Petrova ◽

Y. Tzvetanova

Keyword(s):

Surface Area ◽

Cement Paste ◽

Chemical Reactions ◽

Natural Zeolite ◽

Early Stage ◽

Pozzolanic Activity ◽

Large Surface Area ◽

Natural Zeolites ◽

Hydration Products ◽

Cement Pastes

AbstractThis paper presents results from comparative thermogravimetric, calorimetric and pozzolanic activity analyses of five natural zeolite samples from Bulgaria, Slovakia, Philippines, USA and North Korea. The zeolites actively participate in the hydration processes of cement. Their activity in the early stage of hydration is based mainly on the large surface area of the particles while, in the later stages of activation, chemical reactions occur between the products of the hydration of cement and the soluble SiO2 that is present in the bulk of the zeolites. It has been shown that in all cement pastes which contain zeolite additives, the quantity of portlandite is lower than that in pure cement paste or is even totally absent. The amounts of hydration products are greater when 30% zeolite is used than when 10% zeolite is added (excluding the sample with chabazite). The lowest pozzolanic activity is shown by chabazite, which possessed the lowest SiO2/Al2O2 ratio.

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