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Characterization of the FtsZ C-Terminal Variable (CTV) Region in Z-Ring Assembly and Interaction with the Z-Ring Stabilizer ZapD in E. coli Cytokinesis.

Huang KH, Mychack A, Tchorzewski L, Janakiraman A - PLoS ONE (2016)

Bottom Line: Multiple Z-ring associated proteins (Zaps), also promote lateral interactions between FtsZ protofilaments to stabilize the FtsZ ring in vivo.Our data suggest a mechanism in which the CTV residues, particularly K380, facilitate a conformation for the conserved carboxy-terminal residues in FtsZ, that lie immediately N-terminal to the CTV, to enable optimal contact with ZapD.Further, phylogenetic analyses suggest a correlation between the nature of FtsZ CTV residues and the presence of ZapD in the β- γ-proteobacterial species.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, City College of CUNY, 160 Convent Avenue, MR 526, New York, NY, United States of America.

ABSTRACT
Polymerization of a ring-like cytoskeletal structure, the Z-ring, at midcell is a highly conserved feature in virtually all bacteria. The Z-ring is composed of short protofilaments of the tubulin homolog FtsZ, randomly arranged and held together through lateral interactions. In vitro, lateral associations between FtsZ protofilaments are stabilized by crowding agents, high concentrations of divalent cations, or in some cases, low pH. In vivo, the last 4-10 amino acid residues at the C-terminus of FtsZ (the C-terminal variable region, CTV) have been implicated in mediating lateral associations between FtsZ protofilaments through charge shielding. Multiple Z-ring associated proteins (Zaps), also promote lateral interactions between FtsZ protofilaments to stabilize the FtsZ ring in vivo. Here we characterize the complementary role/s of the CTV of E. coli FtsZ and the FtsZ-ring stabilizing protein ZapD, in FtsZ assembly. We show that the net charge of the FtsZ CTV not only affects FtsZ protofilament bundling, confirming earlier observations, but likely also the length of the FtsZ protofilaments in vitro. The CTV residues also have important consequences for Z-ring assembly and interaction with ZapD in the cell. ZapD requires the FtsZ CTV region for interaction with FtsZ in vitro and for localization to midcell in vivo. Our data suggest a mechanism in which the CTV residues, particularly K380, facilitate a conformation for the conserved carboxy-terminal residues in FtsZ, that lie immediately N-terminal to the CTV, to enable optimal contact with ZapD. Further, phylogenetic analyses suggest a correlation between the nature of FtsZ CTV residues and the presence of ZapD in the β- γ-proteobacterial species.

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FtsZ domain structure, FtsZ C-terminal tail (CTT) structure and FtsZ C-terminal variable (CTV) mutant constructs.A. Domain organization of E. coli FtsZ: an unstructured 10 residues at the N-terminal end (squiggly line), a conserved globular core domain containing the nucleotide binding and hydrolysis residues, a flexible variable linker about 50 residues long (squiggly line), and a conserved carboxy terminal peptide (CCTP) which contains both a constant region of ~13 residues (CTC) and a variable region of 4 residues (CTV). B. Structural model of the FtsZ C-terminal residues 367–383 (PDB 1F47) [29]. In a X-ray crystal structure complex with the essential division protein ZipA, the 17 residue FtsZ CCTP binds as an extended β-strand followed by an α-helix. The CTV residue side-chains are identified in the α-helix: K380 (blue), Q381 and A382 (gray) and D383 (red). C. Schematic of the FtsZ CTV mutant constructs used in the study, not drawn to scale.
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pone.0153337.g001: FtsZ domain structure, FtsZ C-terminal tail (CTT) structure and FtsZ C-terminal variable (CTV) mutant constructs.A. Domain organization of E. coli FtsZ: an unstructured 10 residues at the N-terminal end (squiggly line), a conserved globular core domain containing the nucleotide binding and hydrolysis residues, a flexible variable linker about 50 residues long (squiggly line), and a conserved carboxy terminal peptide (CCTP) which contains both a constant region of ~13 residues (CTC) and a variable region of 4 residues (CTV). B. Structural model of the FtsZ C-terminal residues 367–383 (PDB 1F47) [29]. In a X-ray crystal structure complex with the essential division protein ZipA, the 17 residue FtsZ CCTP binds as an extended β-strand followed by an α-helix. The CTV residue side-chains are identified in the α-helix: K380 (blue), Q381 and A382 (gray) and D383 (red). C. Schematic of the FtsZ CTV mutant constructs used in the study, not drawn to scale.

Mentions: The FtsZ monomer comprises four domains: an unstructured poorly conserved region of ~10 residues at the extreme N-terminus; a highly conserved globular core containing the GTP binding and hydrolytic functions; a disordered flexible linker that is poorly conserved in length and sequence among species; and the C-terminal conserved peptide (CCTP) which contains two sub-regions: a conserved C-terminal constant region (CTC) and a C-terminal variable region (CTV) (Fig 1) [17–19]. In vitro, the globular core alone is sufficient for formation of FtsZ protofilaments [20,21]. But, recent work has implicated the flexible linker and FtsZ CTV sequences to be key determinants of the end-to-end and lateral interactions of FtsZ in vitro implying a role for these domains in the architecture of FtsZ assemblies in the cell [17,18,22]. Most known stabilizers and destabilizers of FtsZ interact with the CCTP, which serves as a dock for proteins that regulate FtsZ-ring assembly dynamics. Such proteins include the essential FtsZ membrane tethers FtsA and ZipA, positional regulators MinC and SlmA, the conserved protease ClpX, and the Z-ring stabilizer ZapD [23–28]. The structure of CCTP bound to ZipA in E. coli and FtsA in T. maritima have been solved [29,30]. Although the CCTP in each case contains a helical segment starting at a conserved proline, the extended structures are not identical, suggesting that the CCTP is capable of acquiring a variety of structures possibly to enable interactions with varied binding partners [30,31].


Characterization of the FtsZ C-Terminal Variable (CTV) Region in Z-Ring Assembly and Interaction with the Z-Ring Stabilizer ZapD in E. coli Cytokinesis.

Huang KH, Mychack A, Tchorzewski L, Janakiraman A - PLoS ONE (2016)

FtsZ domain structure, FtsZ C-terminal tail (CTT) structure and FtsZ C-terminal variable (CTV) mutant constructs.A. Domain organization of E. coli FtsZ: an unstructured 10 residues at the N-terminal end (squiggly line), a conserved globular core domain containing the nucleotide binding and hydrolysis residues, a flexible variable linker about 50 residues long (squiggly line), and a conserved carboxy terminal peptide (CCTP) which contains both a constant region of ~13 residues (CTC) and a variable region of 4 residues (CTV). B. Structural model of the FtsZ C-terminal residues 367–383 (PDB 1F47) [29]. In a X-ray crystal structure complex with the essential division protein ZipA, the 17 residue FtsZ CCTP binds as an extended β-strand followed by an α-helix. The CTV residue side-chains are identified in the α-helix: K380 (blue), Q381 and A382 (gray) and D383 (red). C. Schematic of the FtsZ CTV mutant constructs used in the study, not drawn to scale.
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pone.0153337.g001: FtsZ domain structure, FtsZ C-terminal tail (CTT) structure and FtsZ C-terminal variable (CTV) mutant constructs.A. Domain organization of E. coli FtsZ: an unstructured 10 residues at the N-terminal end (squiggly line), a conserved globular core domain containing the nucleotide binding and hydrolysis residues, a flexible variable linker about 50 residues long (squiggly line), and a conserved carboxy terminal peptide (CCTP) which contains both a constant region of ~13 residues (CTC) and a variable region of 4 residues (CTV). B. Structural model of the FtsZ C-terminal residues 367–383 (PDB 1F47) [29]. In a X-ray crystal structure complex with the essential division protein ZipA, the 17 residue FtsZ CCTP binds as an extended β-strand followed by an α-helix. The CTV residue side-chains are identified in the α-helix: K380 (blue), Q381 and A382 (gray) and D383 (red). C. Schematic of the FtsZ CTV mutant constructs used in the study, not drawn to scale.
Mentions: The FtsZ monomer comprises four domains: an unstructured poorly conserved region of ~10 residues at the extreme N-terminus; a highly conserved globular core containing the GTP binding and hydrolytic functions; a disordered flexible linker that is poorly conserved in length and sequence among species; and the C-terminal conserved peptide (CCTP) which contains two sub-regions: a conserved C-terminal constant region (CTC) and a C-terminal variable region (CTV) (Fig 1) [17–19]. In vitro, the globular core alone is sufficient for formation of FtsZ protofilaments [20,21]. But, recent work has implicated the flexible linker and FtsZ CTV sequences to be key determinants of the end-to-end and lateral interactions of FtsZ in vitro implying a role for these domains in the architecture of FtsZ assemblies in the cell [17,18,22]. Most known stabilizers and destabilizers of FtsZ interact with the CCTP, which serves as a dock for proteins that regulate FtsZ-ring assembly dynamics. Such proteins include the essential FtsZ membrane tethers FtsA and ZipA, positional regulators MinC and SlmA, the conserved protease ClpX, and the Z-ring stabilizer ZapD [23–28]. The structure of CCTP bound to ZipA in E. coli and FtsA in T. maritima have been solved [29,30]. Although the CCTP in each case contains a helical segment starting at a conserved proline, the extended structures are not identical, suggesting that the CCTP is capable of acquiring a variety of structures possibly to enable interactions with varied binding partners [30,31].

Bottom Line: Multiple Z-ring associated proteins (Zaps), also promote lateral interactions between FtsZ protofilaments to stabilize the FtsZ ring in vivo.Our data suggest a mechanism in which the CTV residues, particularly K380, facilitate a conformation for the conserved carboxy-terminal residues in FtsZ, that lie immediately N-terminal to the CTV, to enable optimal contact with ZapD.Further, phylogenetic analyses suggest a correlation between the nature of FtsZ CTV residues and the presence of ZapD in the β- γ-proteobacterial species.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, City College of CUNY, 160 Convent Avenue, MR 526, New York, NY, United States of America.

ABSTRACT
Polymerization of a ring-like cytoskeletal structure, the Z-ring, at midcell is a highly conserved feature in virtually all bacteria. The Z-ring is composed of short protofilaments of the tubulin homolog FtsZ, randomly arranged and held together through lateral interactions. In vitro, lateral associations between FtsZ protofilaments are stabilized by crowding agents, high concentrations of divalent cations, or in some cases, low pH. In vivo, the last 4-10 amino acid residues at the C-terminus of FtsZ (the C-terminal variable region, CTV) have been implicated in mediating lateral associations between FtsZ protofilaments through charge shielding. Multiple Z-ring associated proteins (Zaps), also promote lateral interactions between FtsZ protofilaments to stabilize the FtsZ ring in vivo. Here we characterize the complementary role/s of the CTV of E. coli FtsZ and the FtsZ-ring stabilizing protein ZapD, in FtsZ assembly. We show that the net charge of the FtsZ CTV not only affects FtsZ protofilament bundling, confirming earlier observations, but likely also the length of the FtsZ protofilaments in vitro. The CTV residues also have important consequences for Z-ring assembly and interaction with ZapD in the cell. ZapD requires the FtsZ CTV region for interaction with FtsZ in vitro and for localization to midcell in vivo. Our data suggest a mechanism in which the CTV residues, particularly K380, facilitate a conformation for the conserved carboxy-terminal residues in FtsZ, that lie immediately N-terminal to the CTV, to enable optimal contact with ZapD. Further, phylogenetic analyses suggest a correlation between the nature of FtsZ CTV residues and the presence of ZapD in the β- γ-proteobacterial species.

Show MeSH
Related in: MedlinePlus