Regular cellular distribution of plasmids by oscillating and filament‐forming ParA ATPase of plasmid pB171

All cells require the ability to process spatial information to properly position intracellular molecules. Many protein complexes and DNA molecules are actively positioned either at the cell midpoint or cell poles, but the processes which drive intracellular positioning are still poorly understood. Using computational modeling we propose a bimodal centering/segregation mechanism in bacteria which is driven by the dynamic instability of polymerizing filaments, which grow and shrink with regularity. Modeled cell centering via dynamically unstable filaments is confirmed experimentally via in vivo time-lapse, colocalization measurements of a model system of clustered plasmid-DNA centered by the dynamically unstable actin-like protein filaments Alp7A in Bacillus subtilis. Generalizing to any cylindrical cell, we find strong cell-length dependence in the centering ability of dynamically unstable filaments, culminating in pole positioning when cell length decreases significantly below the theoretically predicted average filament length. Modeling dynamic instability-driven positioning mechanisms from multiple anisotropic in vivo systems demonstrates that dynamically unstable filaments are a general mechanism for both midcell and cell-pole (segregation) positioning, and that desired positioning is preferentially selected in vivo by intrinsic filament polymerization rates and number.cytoskeleton | bacterial actin polymerization | mathematical modeling

Section: Resultsmentioning

confidence: 99%

Dynamic instability-driven centering/segregating mechanism in bacteria

Drew

Pogliano

2011

“…Unlike ParM, which forms a filamentous structure in vivo extending between two segregating plasmids, many ParA proteins oscillate from one end of the cell to the other with periods that do not correspond to the dynamics of plasmid or chromosome segregation (11,13,16,(23)(24)(25). Furthermore, the structures of ParA filaments differ from their type II counterparts, suggesting that type I par systems rely on distinct mechanisms to mediate segregation.…”

mentioning

confidence: 95%

“…Several of these proteins have been shown to form dynamic filamentous structures in vivo and/or ATPdependent filaments in vitro (9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20). The best example of how a Par ATPase can drive the partitioning process is provided by studies of ParM, the type II ATPase of plasmid R1.…”

mentioning

confidence: 99%

“…Furthermore, the structures of ParA filaments differ from their type II counterparts, suggesting that type I par systems rely on distinct mechanisms to mediate segregation. Several ParA homologs-including SopA of F (Ia), ParF of pTP228 (Ib), and ParA of pB171 (Ib)-form bundled filaments with one frayed end when incubated with ATP, leading to speculation that filament polarity is important for function (12,16,17).…”

mentioning

confidence: 99%

See 1 more Smart Citation

ParA2, aVibrio choleraechromosome partitioning protein, forms left-handed helical filaments on DNA

Hui

Galkin²,

Xiong³

et al. 2010

Most bacterial chromosomes contain homologs of plasmid partitioning (par) loci. These loci encode ATPases called ParA that are thought to contribute to the mechanical force required for chromosome and plasmid segregation. In Vibrio cholerae, the chromosome II (chrII) par locus is essential for chrII segregation. Here, we found that purified ParA2 had ATPase activities comparable to other ParA homologs, but, unlike many other ParA homologs, did not form high molecular weight complexes in the presence of ATP alone. Instead, formation of high molecular weight ParA2 polymers required DNA. Electron microscopy and three-dimensional reconstruction revealed that ParA2 formed bipolar helical filaments on double-stranded DNA in a sequence-independent manner. These filaments had a distinct change in pitch when ParA2 was polymerized in the presence of ATP versus in the absence of a nucleotide cofactor. Fitting a crystal structure of a ParA protein into our filament reconstruction showed how a dimer of ParA2 binds the DNA. The filaments formed with ATP are left-handed, but surprisingly these filaments exert no topological changes on the right-handed B-DNA to which they are bound. The stoichiometry of binding is one dimer for every eight base pairs, and this determines the geometry of the ParA2 filaments with 4.4 dimers per 120 Å pitch left-handed turn. Our findings will be critical for understanding how ParA proteins function in plasmid and chromosome segregation.nucleoprotein filament | DNA segregation

“…ParA proteins interact with the ParB-parS complex, and their ATPase activity is essential for plasmid partitioning (5,6). Several ParA homologues have been shown to form ATPdependent polymers in vitro and to oscillate in vivo (7)(8)(9)(10). Still, the molecular mechanisms by which the interactions of ParA with the ParB-parS complex mediate plasmid localization and partitioning are not well understood, although several models have been proposed to account for Par-mediated plasmid segregation (9-13).…”

mentioning

confidence: 99%

par genes and the pathology of chromosome loss in Vibrio cholerae

Yamaichi¹,

Fogel

Waldor

2007

107

The causes and consequences of chromosome loss in bacteria with multiple chromosomes are unknown. Vibrio cholerae, the causative agent of the severe diarrheal disease cholera, has two circular chromosomes. Like many other bacterial chromosomes, both V. cholerae chromosomes contain homologues of plasmid partitioning (par) genes. In plasmids, par genes act to segregate plasmid molecules to daughter cells and thereby ensure plasmid maintenance; however, the contribution of par genes to chromosome segregation is not clear. Here, we show that the chromosome II parAB2 genes are essential for the segregation of chromosome II but not chromosome I. In a parAB2 deletion mutant, chromosome II is mislocalized and frequently fails to segregate, yielding cells with only chromosome I. These cells divide once; their progeny are not viable. Instead, chromosome II-deficient cells undergo dramatic cell enlargement, nucleoid condensation and degradation, and loss of membrane integrity. The highly consistent nature of these cytologic changes suggests that prokaryotes, like eukaryotes, may possess characteristic death pathways.chromosome segregation ͉ parA ͉ parB ͉ apoptosis