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Bacterial Cytokinesis: from Z Ring to Divisome

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Bacterial Cytokinesis: from Z Ring to Divisome REVIEW ARTICLE Cytoskeleton, October 2012 69:778–790 (doi: 10.1002/cm.21054) VC 2012 Wiley Periodicals, Inc. Bacterial Cytokinesis: From Z Ring to Divisome Joe Lutkenhaus, Sebastien Pichoff, and Shishen Du Department of Microbiology, Molecular Genetics and Immunology, University of Kansas Medical Center, Kansas City, Kansas Received 21 June 2012; Revised 18 July 2012; Accepted 20 July 2012 Monitoring Editor: Joseph Sanger Ancestral homologues of the major eukaryotic cytos- Cytokinesis in bacteria can be split into at least three keletal families, tubulin and actin, play critical roles in steps [de Boer, 2010]. First, is the assembly of the Z ring cytokinesis of bacterial cells. FtsZ is the ancestral on the cytoplasmic membrane with the aid of membrane homologue of tubulin and assembles into the Z ring tethering proteins [Pichoff and Lutkenhaus, 2002]. This that determines the division plane. FtsA, a member of step is under spatial and temporal control to ensure that the actin family, is involved in coordinating cell wall the Z ring is assembled between segregated chromosomes synthesis during cytokinesis. FtsA assists in the forma- [Lutkenhaus, 2007]. In the second step, which usually tion of the Z ring and also has a critical role in occurs after a considerable lag, the remaining cell division recruiting downstream division proteins to the Z ring proteins are added to the Z ring to form the complete to generate the divisome that divides the cell. Spatial divisome [Aarsman et al., 2005; Gamba et al., 2009; regulation of cytokinesis occurs at the stage of Z ring Goley et al., 2011]. Formation of this complex machine assembly and regulation of cell size occurs at this stage involves the addition of many essential proteins and an or during Z ring maturation. VC 2012 Wiley Periodicals, Inc increasing number of nonessential proteins that have par- tially overlapping functions [Goehring and Beckwith, Key Words: cytokinesis, FtsZ/tubulin, FtsA/actin 2005; Vicente and Rico, 2006; de Boer, 2010]. Third, the divisome is activated to synthesize septal peptidoglycan, which has to be split so that the progeny cells can separate Introduction [Gerding et al., 2009]. This third step is under complex topological control so that cell wall degrading enzymes are acterial cytokinesis has been studied primarily in E. only activated at the correct place after septal cell wall Bcoli and B. subtilis, two bacteria with a similar rod synthesis has initiated [Uehara and Bernhardt, 2011]. shape. A comparison between these two organisms, which are separated in evolutionary time longer than yeast and FtsZ and Assembly of the Z Ring humans, has revealed the basic components of the cytoki- netic machinery [Errington et al., 2003]. Despite some FtsZ differences, defining the machinery in these two bacteria has demonstrated a core of essential components that are FtsZ is considered the ancestral homologue of eukaryotic used by many bacteria. Furthermore, these investigations tubulins [Nogales et al., 1998]. Overall the amino acid have facilitated the study of cytokinesis in other bacteria, identity is only on the order of 10% with the highest by allowing investigators to hone in on differences. Chlor- degree of conservation involving residues required for oplasts and many Archaea use FtsZ for division but this GTP binding and hydrolysis [Nogales et al., 1998; Erick- will not be discussed here [Wang and Lutkenhaus, 1996; son, 2007] (Fig. 1). GTP is bound on one end of an Miyagishima, 2011]. In contrast, some Archaea and most FtsZ/tubulin (the þ end) subunit with the aid of the sig- mitochondria use Escrt or dynamin based cell division nature FtsZ/tubulin loop (GGGTG[S/T]G) which binds machineries, respectively [Osteryoung, 2001; Bernander the phosphates. The GTPase catalytic site is formed dur- and Ettema, 2010]. ing filament assembly by the addition of the synergy loop (NxDxx[D/E]) from the incoming subunit. Despite this limited sequence identity, however, the similar structures *Address correspondence to: Joe Lutkenhaus, Department of of the monomers and filaments, as well a similar mecha- Microbiology, Molecular Genetics and Immunology, University of nism of the GTPase, argue that these two proteins are ev- Kansas Medical Center, Kansas City, Kansas 66160. E-mail: [email protected] olutionary related filament forming proteins [Michie and Published online 30 August 2012 in Wiley Online Library L€owe, 2006]. Furthermore, the recent isolation of an in- (wileyonlinelibrary.com). hibitor of FtsZ that stabilizes FtsZ filaments and the n 778 Fig. 1. Structures of FtsZ and tubulin. The residues that are most conserved in FtsZ (PDB 2VAW) and tubulin (PDB 1TUB) are involved in the binding and hydrolyzing of GTP (colored cyan) and include the synergy loop (NxDxx[D/E] (important residues in caps and colored magenta in the structure) and the signature loop (GGGTG[T/S]G, colored blue) that binds the phosphates. The synergy loop in b-tubulin does not induce GTP hydrolysis due to a positive charged residue (K) substituted for an acidic residue (E) in the loop. A substitution of the acidic residue in FtsZ with G (FtsZ2) results in loss of GTPase activity. realization that its binding site is analogous to the taxol is accounted for by having an inactive monomer undergo binding site in tubulin, which stabilizes microtubules, fur- an izomerization reaction to an active form before associa- ther highlights the similarity [Haydon et al., 2008; tion with the next monomer [Dajkovic et al., 2008; Hue- Andreu et al., 2010; Elsen et al., 2012]. cas et al., 2008; Miraldi et al., 2008]. The existence of FtsZ forms dynamic filaments in the presence of GTP two forms of FtsZ is supported by the recent structure of that are structurally similar to a protofilament present in a FtsZ in the presence of an inhibitor [Elsen et al., 2012]. microtubule [Mukherjee and Lutkenhaus, 1998; L€owe and FtsZ filaments display different degrees of curvature Amos, 1999]. The filaments readily bundle depending depending upon the nucleotide. Initially, straight filaments upon the in vitro conditions, however, the basic unit of were associated with bound GTP and highly curved fila- assembly is a filament that is a single subunit thick ments with GDP [Lu et al., 2000]. However, bound GTP [Mukherjee and Lutkenhaus, 1994; Chen et al., 2005]. has also been associated with a gently curved form by Under conditions that favor maximal GTPase activity the both atomic force microscopy (AFM) and electron micros- average filament contains about 30 subunits, however, copy [Mingorance et al., 2005]. The possibility that GTP many in vitro conditions lead to lateral bundling which containing filaments adopt a fixed curvature, which can slows down the GTPase and results in longer filaments deform membranes in vitro, has been used as a basis for a [Chen and Erickson, 2009]. However, close inspection of proposal that FtsZ provides the force for constriction bundled filaments did not reveal an orderly arrangement [Erickson et al., 2010]. The reader is referred to a com- and it was argued that lateral bonds do not exist [Erickson prehensive review on the in vitro properties and behavior et al., 2010]. of FtsZ [Erickson et al., 2010]. Although there was some concern whether assembly of a filament a single molecule thick would undergo nucleated assembly and display a critical concentration Z Ring [Romberg et al., 2001], it is now clear that FtsZ displays The Z ring was first visualized as an entity by immunoe- a critical concentration around 1 lM, remarkably similar lectron microscopy [Bi and Lutkenhaus, 1991], then by to tubulin [Mukherjee and Lutkenhaus, 1998; Chen immunofluorescence microscopy [Addinall et al., 1996; et al., 2005]. The nucleated assembly of a linear filament Levin and Losick, 1996], but it is now readily visualized CYTOSKELETON Bacterial Cytokinesis: From Z Ring to Divisome 779 n Fig. 2. FtsZ filaments are tethered to the membrane by FtsA and ZipA. ZipA and FtsA bind to the highly conserved C-tail of FtsZ, which is connected to main body of FtsZ by a flexible linker. Both ZipA and FtsA are tethered to the membrane by a flexible linker that connects the membrane binding domains of these proteins to the main body of the protein. in live cells by tagging FtsZ or any of a number of com- ments completely encircling the septum [Anderson et al., ponents that are recruited to the Z ring with green fluo- 2004; Geissler et al., 2007]. This suggests not all FtsZ fil- rescent protein (GFP) [Ma et al., 1996]. So far all studies aments are captured by cryoelectron microscopy. FRAP have been done with GFP tagged FtsZ, which can not (fluorescent recovery after photobleaching) studies have substitute for FtsZ and is toxic. However, the expression shown that the subunits in the Z ring are rapidly turning of GFP-FtsZ does not appear to interfere with division if over (T1/2 of 8–10 s) [Stricker et al., 2002; Erickson the expression level is less than 25% of the native FtsZ. et al., 2010]. To account for the amount of FtsZ in the Z The use of GFP-FtsZ revealed that the Z ring is ring and the rapid turnover, it was proposed that the Z assembled gradually with FtsZ being initially loosely ring consists of short overlapping filaments. One of the organized at midcell before eventually coalescing into a major questions in the field is the substructure of the Z ring. This transition correlates with the segregation of the ring. Overproduction of FtsZ or removal of spatial regula- nucleoids [Inoue et al., 2009] and is promoted by various tors leads to Z rings at midcell and the poles of the cell, nonessential Z associated proteins (Zap). Since some of or doublets between nucleoids in long cells, rather than a these Zap proteins have the ability to bundle FtsZ fila- thickening of the existing ring suggesting that Z rings ments in vitro [Gueiros-Filho and Losick, 2002; Monahan have a defined structure [Bi and Lutkenhaus, 1990; Yu et al., 2009; Dajkovic et al., 2010; Hale et al., 2011; and Margolin, 1999; Quardokus et al., 2001; Weart and Durand-Heredia et al., 2012], it suggests that some degree Levin, 2003].
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