The GTPase Activity of <i>Escherichia coli</i> FtsZ Determines the Magnitude of the FtsZ Polymer Bundling by ZapA <i>in Vitro</i>

Mohammadi, Tamimount; Ploeger, Ginette E. J.; Verheul, Jolanda; Comvalius, Anouskha D.; Martos, Ariadna; Alfonso, Carlos; Marle, J. van; Rivas, Germán; Blaauwen, Tanneke den

doi:10.1021/bi901461p

Cited by 86 publications

(173 citation statements)

References 54 publications

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“…Further, Smith et al did not observe protofilament bundling when FtsZ1 and FtsZ2 were mixed, whereas we observed extensive bundling under a variety of conditions. Finally, Smith et al concluded that polymerization of FtsZ1 and FtsZ2 requires GTP hydrolysis based on a lack of assembly in the presence a nonhydrolozyable GTP analog, but we found that FtsZ1 and FtsZ2 mutants lacking GTPase activity still exhibit GTP-dependent assembly, consistent with data showing that assembly of bacterial FtsZ, although GTP-dependent, does not require GTP hydrolysis (9,33,43,56). These differences may be due in part to differences in assay conditions or C-terminal tags, but another explanation could be dissimilarities in the lengths of the recombinant proteins used in the two studies, which were truncated to remove the predicted chloroplast transit peptides.…”

Section: Discussionsupporting

confidence: 90%

GTP-dependent Heteropolymer Formation and Bundling of Chloroplast FtsZ1 and FtsZ2

Olson

Wang

Osteryoung

2010

Journal of Biological Chemistry

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Bacteria and chloroplasts require the ring-forming cytoskeletal protein FtsZ for division. Although bacteria accomplish division with a single FtsZ, plant chloroplasts require two FtsZ types for division, FtsZ1 and FtsZ2. These proteins colocalize to a mid-plastid Z ring, but their biochemical relationship is poorly understood. We investigated the in vitro behavior of recombinant FtsZ1 and FtsZ2 separately and together. Both proteins bind and hydrolyze GTP, although GTPase activities are low compared with the activity of Escherichia coli FtsZ. Each protein undergoes GTP-dependent assembly into thin protofilaments in the presence of calcium as a stabilizing agent, similar to bacterial FtsZ. In contrast, when mixed without calcium, FtsZ1 and FtsZ2 exhibit slightly elevated GTPase activity and coassembly into extensively bundled protofilaments. Coassembly is enhanced by FtsZ1, suggesting that it promotes lateral interactions between protofilaments. Experiments with GTPase-deficient mutants reveal that FtsZ1 and FtsZ2 form heteropolymers. Maximum coassembly occurs in reactions containing equimolar FtsZ1 and FtsZ2, but significant coassembly occurs at other stoichiometries. The FtsZ1:FtsZ2 ratio in coassembled structures mirrors their input ratio, suggesting plasticity in protofilament and/or bundle composition. This behavior contrasts with that of ␣-and ␤-tubulin and the bacterial tubulin-like proteins BtubA and BtubB, which coassemble in a strict 1:1 stoichiometry. Our findings raise the possibility that plasticity in FtsZ filament composition and heteropolymerization-induced bundling could have been a driving force for the coevolution of FtsZ1 and FtsZ2 in the green lineage, perhaps arising from an enhanced capacity for the regulation of Z ring composition and activity in vivo.Cell division in bacteria and chloroplast division in plants both require FtsZ, a tubulin-like GTPase that functions as a contractile ring, the Z ring, inside the cell or chloroplast. Mutations in bacterial FtsZ genes disrupt cell division, resulting in long filamented cells (reviewed in Refs. 1-3). Purified bacterial FtsZ undergoes GTP-dependent, hydrolysis-independent polymerization primarily into single protofilaments (4, 5), but protofilament sheets and bundles form in the presence of stabilizing agents that promote lateral interactions (6 -8). Polymerization stimulates FtsZ GTPase activity because the catalytic site for GTP hydrolysis lies in the longitudinal interface between adjacent monomers within the polymer (9). GTP hydrolysis destabilizes protofilaments, leading to disassembly (10). The in vivo structure of the bacterial Z ring is not yet clear, but recent models suggest it may be built from short overlapping protofilaments that are stabilized at the division site by accessory factors (11, 12). The Z ring functions partly as a scaffold for other cell division proteins and recently has also been shown to provide contractile force for membrane constriction (13,14).In most bacteria, including the cyanobacterial relatives of chloropl...

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Section: Discussionsupporting

confidence: 90%

GTP-dependent Heteropolymer Formation and Bundling of Chloroplast FtsZ1 and FtsZ2

Olson

Wang

Osteryoung

2010

Journal of Biological Chemistry

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“…Subsequently, polymerization and staining were carried out as published (Mohammadi et al 2009) in the presence of 5 mM MgCl 2 and 0.2 mM GTP. After 10 min of incubation at 30°C, samples were applied to a microscope grid, stained with uranyl acetate, and examined with transmission electron microscopy.…”

Section: In Vitro Polymerization Of Ftsz and Pelleting Assaymentioning

confidence: 99%

Positive control of cell division: FtsZ is recruited by SsgB during sporulation ofStreptomyces

Willemse¹,

Borst²,

Waal³

et al. 2011

Genes Dev.

171

201

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In bacteria that divide by binary fission, cell division starts with the polymerization of the tubulin homolog FtsZ at mid-cell to form a cell division scaffold (the Z ring), followed by recruitment of the other divisome components. The current view of bacterial cell division control starts from the principle of negative checkpoints that prevent incorrect Z-ring positioning. Here we provide evidence of positive control of cell division during sporulation of Streptomyces, via the direct recruitment of FtsZ by the membrane-associated divisome component SsgB. In vitro studies demonstrated that SsgB promotes the polymerization of FtsZ. The interactions are shown in vivo by time-lapse imaging and Fö rster resonance energy transfer and fluorescence lifetime imaging microscopy (FRET-FLIM), and are corroborated independently via two-hybrid studies. As determined by fluorescence recovery after photobleaching (FRAP), the turnover of FtsZ protofilaments increased strongly at the time of Z-ring formation. The surprising positive control of Z-ring formation by SsgB implies the evolution of an entirely new way of Z-ring control, which may be explained by the absence of a mid-cell reference point in the long multinucleoid hyphae. In turn, the localization of SsgB is mediated through the orthologous SsgA, and premature expression of the latter is sufficient to directly activate multiple Z-ring formation and hyperdivision at early stages of the Streptomyces cell cycle.

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“…These proteins act at midcell, promoting the transition of FtsZ polymers from a helical band into a compact ring by cooperatively stimulating the lateral association of protofilaments (47,48). The Zap proteins bind to a conserved motif of 10 -16 residues found within the C-terminal end of FtsZ (49 -52).…”

Section: The Proto-ring: the Scaffold Of The Divisomementioning

confidence: 99%

In the Beginning, Escherichia coli Assembled the Proto-ring: An Initial Phase of Division

Rico

Krupka²,

Vicente

2013

Journal of Biological Chemistry

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Cell division in Escherichia coli begins by assembling three proteins, FtsZ, FtsA, and ZipA, to form a proto-ring at midcell. These proteins nucleate an assembly of at least 35 components, the divisome. The structuring of FtsZ to form a ring and the processes that effect constriction have been explained by alternative but not mutually exclusive mechanisms. We discuss how FtsA and ZipA provide anchoring of the cytoplasmic FtsZ to the membrane and how a temporal sequence of alternative protein interactions may operate in the maturation and stability of the proto-ring. How the force needed for constriction is generated and how the proto-ring proteins relate to peptidoglycan synthesis remain as the main challenges for future research. Overview of SeptationCell division is the last event in the bacterial cell cycle. It takes place by producing a rigid transversal septum after the growth processes fully duplicate the contents of the cell. Septation is always preceded by segregation of the duplicated nucleoids, ensuring that each of the daughter cells contains a fully replicated chromosome. Specialized molecules control the positioning of the division machinery at midcell, preventing any harmful fragmentation that septum progression could cause on unsegregated nucleoids.The division machinery, the divisome, establishes a complex interacting network together with DNA, lipids, and peptidoglycan components. It comprises a variable number of proteins, depending on the bacterial species. These proteins have redundant multiple functions and connections serving as safeguards to complete division and to guarantee the efficiency and versatility of the process. Altogether, an efficient division process allows the proliferation of bacteria under a variety of conditions, and it is one of the reasons for their success in the environment. Free-living bacteria, such as Escherichia coli, have more elaborated divisomes, whereas those that need to grow as intracellular parasites, as such Mycoplasma genitalium, have streamlined ones (1, 2).The structure of the E. coli divisome as revealed by immunofluorescence images of dividing cells is called the division ring (3). For its assembly, three essential proteins, FtsZ, FtsA, and ZipA, are first gathered at midcell, forming a proto-ring attached to the inner membrane ( Fig. 1) (4). In the proto-ring structure, the cytoplasmic GTPase FtsZ is anchored to the membrane by its interaction with ZipA and FtsA. ZipA is a bitopic protein containing a transmembrane domain (5). FtsA is a protein of the actin family that associates with the membrane by a short amphipathic helix (6, 7). The assembly of the FtsZ ring is dependent on a continuous energy supply. The FtsZ ring requires either ZipA or FtsA (8). Although it can be formed in the presence of either of them, division is affected unless both are present (9). A maturation period takes place after the protoring is assembled and before the rest of the divisome proteins become incorporated (10). During maturation, the proto-ring assembly is not stable...

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The GTPase Activity of Escherichia coli FtsZ Determines the Magnitude of the FtsZ Polymer Bundling by ZapA in Vitro

Cited by 86 publications

References 54 publications

GTP-dependent Heteropolymer Formation and Bundling of Chloroplast FtsZ1 and FtsZ2

GTP-dependent Heteropolymer Formation and Bundling of Chloroplast FtsZ1 and FtsZ2

Positive control of cell division: FtsZ is recruited by SsgB during sporulation ofStreptomyces

In the Beginning, Escherichia coli Assembled the Proto-ring: An Initial Phase of Division

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