Reconstitution of Contractile FtsZ Rings in Liposomes

Department of Cell Biology, Duke University Medical Center, Durham, NC 27710-3709, USA.
Science (Impact Factor: 33.61). 06/2008; 320(5877):792-4. DOI: 10.1126/science.1154520
Source: PubMed


FtsZ is a tubulin homolog and the major cytoskeletal protein in bacterial cell division. It assembles into the Z ring, which
contains FtsZ and a dozen other division proteins, and constricts to divide the cell. We have constructed a membrane-targeted
FtsZ (FtsZ-mts) by splicing an amphipathic helix to its C terminus. When mixed with lipid vesicles, FtsZ-mts was incorporated
into the interior of some tubular vesicles. There it formed multiple Z rings that could move laterally in both directions
along the length of the liposome and coalesce into brighter Z rings. Brighter Z rings produced visible constrictions in the
liposome, suggesting that FtsZ itself can assemble the Z ring and generate a force. No other proteins were needed for assembly
and force generation.

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    • "A YFP tagged FtsZ fusion in which the GHP motif of FtsZ has been replaced with an amphipathic helix to ensure association between FtsZ and the membrane, is sufficient to support assembly of FtsZ rings around the interior circumference of tubular liposomes. In the context of these liposomes, the fluorescently tagged FtsZ (FtsZ-mts) was able to assemble into rings that appeared to constrict slightly in the presence of GTP [25], suggesting a role for nucleotide hydrolysis in FtsZ force generation. More "
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    ABSTRACT: In bacteria, animals, fungi, and many single celled eukaryotes, division is initiated by the formation of a ring of cytoskeletal protein at the nascent division site. In bacteria, the tubulin-like GTPase FtsZ serves as the foundation for the cytokinetic ring. A conserved feature of FtsZ is an intrinsically disordered peptide known as the C-terminal linker. Chimeric experiments suggest the linker acts as a flexible boom allowing FtsZ to associate with the membrane through a conserved C-terminal domain and also modulates interactions both between FtsZ subunits and between FtsZ and modulatory proteins in the cytoplasm.
    Seminars in Cell and Developmental Biology 10/2014; 37. DOI:10.1016/j.semcdb.2014.09.017 · 6.27 Impact Factor
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    • "We furthermore investigated the assembly of FtsZ proto-rings on membranes and their curvature sensing and inducing propensity (Arumugam et al., 2012), however with no indication so far that the contractile force required for cell division could result from FtsZ only. Several other research groups presently attempt the reconstitution and assembly of bacterial divisome, based mainly on FtsZ and its anchors, in vesicles (Osawa et al., 2008; Jimenez et al., 2011; Cabre et al., 2013; Osawa and Erickson, 2013). However, considering that a progressively contractile role of FtsZ could so far not be established, in spite of its membrane-sculpting ability, it seems appropriate to explore alternative membrane-transforming machineries to accomplish compartment division. "
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    ABSTRACT: In the framework of synthetic biology, it has become an intriguing question what would be the minimal representation of cell division machinery. Thus, it seems appropriate to compare how cell division is realized in different microorganisms. In particular, the cell division system of Crenarchaeota lacks certain proteins found in most bacteria and Euryarchaeota, such as FtsZ, MreB or the Min system. The Sulfolobaceae family encodes functional homologs of the eukaryotic proteins vacuolar protein sorting 4 (Vps4) and endosomal sorting complex required for transport-III (ESCRT-III). ESCRT-III is essential for several eukaryotic pathways, e.g., budding of intraluminal vesicles, or cytokinesis, whereas Vps4 dissociates the ESCRT-III complex from the membrane. Cell Division A (CdvA) is required for the recruitment of crenarchaeal ESCRT-III proteins to the membrane at mid-cell. The proteins polymerize and form a smaller structure during constriction. Thus, ESCRT-III mediated cell division in Sulfolobus acidocaldarius shows functional analogies to the Z ring observed in prokaryotes like Escherichia coli, which has recently begun to be reconstituted in vitro. In this short perspective, we discuss the possibility of building such an in vitro cell division system on basis of archaeal ESCRT-III.
    Frontiers in Microbiology 06/2014; 5:257. DOI:10.3389/fmicb.2014.00257 · 3.99 Impact Factor
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    • "Maybe Vps4 together with ESCRT-III constitutes a mechanochemical membrane-remodeling machinery, in some ways similar to FtsZ rings or dynamin. FtsZ and dynamin are large multidomain GTPases that power several nucleotide-driven conformational states required for membrane scission (Osawa et al., 2008; Faelber et al., 2011; Shnyrova et al., 2013). The binding of Vps4 may trigger first conformational changes in ESCRT-III and then disassembly or both, but we strongly believe that they are all mechanistically linked. "
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    ABSTRACT: Five endosomal sorting complexes required for transport (ESCRTs) mediate the degradation of ubiquitinated membrane proteins via multivesicular bodies (MVBs) in lysosomes. ESCRT-0, -I, and -II interact with cargo on endosomes. ESCRT-II also initiates the assembly of a ringlike ESCRT-III filament consisting of Vps20, Snf7, Vps24, and Vps2. The AAA-adenosine triphosphatase Vps4 disassembles and recycles the ESCRT-III complex, thereby terminating the ESCRT pathway. A mechanistic role for Vps4 in intraluminal vesicle (ILV) formation has been unclear. By combining yeast genetics, biochemistry, and electron tomography, we find that ESCRT-III assembly on endosomes is required to induce or stabilize the necks of growing MVB ILVs. Yet, ESCRT-III alone is not sufficient to complete ILV biogenesis. Rather, binding of Vps4 to ESCRT-III, coordinated by interactions with Vps2 and Snf7, is coupled to membrane neck constriction during ILV formation. Thus, Vps4 not only recycles ESCRT-III subunits but also cooperates with ESCRT-III to drive distinct membrane-remodeling steps, which lead to efficient membrane scission at the end of ILV biogenesis in vivo.
    The Journal of Cell Biology 04/2014; 205(1). DOI:10.1083/jcb.201310114 · 9.83 Impact Factor
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