Replication initiator DnaA binds at the
            <i>Caulobacter</i>
            centromere and enables chromosome segregation

Mera, Paola E.; Kalogeraki, Virginia S.; Shapiro, Lucy

doi:10.1073/pnas.1418989111

Cited by 23 publications

(46 citation statements)

References 38 publications

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“…There is some precedence for DnaA affecting oriC positioning. In Caulobacter crescentus , which requires a functioning ParABS system for oriC segregation (Mohl and Gober, ; Lim et al ., ), DnaA has been shown to promote oriC segregation independent of its role in initiating DNA replication (Mera et al ., ). This finding raises the interesting possibility that other bacteria might also utilize initiator proteins to facilitate chromosome segregation.…”

Section: Discussionmentioning

confidence: 99%

The DnaA inhibitor SirA acts in the same pathway as Soj (ParA) to facilitate oriC segregation during Bacillus subtilis sporulation

Duan

Huey

Herman

2016

Molecular Microbiology

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DNA replication and chromosome segregation must be carefully regulated to ensure reproductive success. During Bacillus subtilis sporulation, chromosome copy number is reduced to two, and cells divide asymmetrically to produce the future spore (forespore) compartment. For successful sporulation, oriC must be captured in the forespore. New rounds of DNA replication are prevented in part by SirA, a protein that utilizes residues in its N-terminus to directly target Domain I of the bacterial initiator, DnaA. Using a quantitative forespore chromosome organization assay, we show that SirA also acts in the same pathway as another DnaA regulator, Soj, to promote oriC capture in the forespore. By analyzing loss-of-function variants of both SirA and DnaA, we observe that SirA's ability to inhibit DNA replication can be genetically separated from its role in oriC capture. In addition, we identify substitutions near the C-terminus of SirA and in DnaA Domain III that enhance interaction between the two proteins. One such variant, SirA , remained functional in regard to inhibiting replication, but was unable to support oriC capture. Collectively, our results support a model in which SirA targets DnaA Domain I to inhibit DNA replication, and DnaA Domain III to facilitate Soj-dependent oriC capture in the forespore.

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Section: Discussionmentioning

confidence: 99%

The DnaA inhibitor SirA acts in the same pathway as Soj (ParA) to facilitate oriC segregation during Bacillus subtilis sporulation

Duan

Huey

Herman

2016

Molecular Microbiology

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“…MipZ inhibits FtsZ polymerization; hence, by associating with ParB, MipZ ends up localized primarily to the polar regions of the Caulobacter , leaving the mid-cell region free for FtsZ polymerization. A recent study suggested that the parS site in Caulobacter interacts not only with ParB, but potentially also with DnaA, providing a link between DNA replication initiation and origin segregation (Mera et al 2014). The model proposed posits that DnaA binds parS sites, somehow altering the structure of the DNA around it in a manner that promotes ParB binding and, consequently, proper segregation.…”

Section: Bacterial Chromosome Segregationmentioning

confidence: 99%

Bacterial Chromosome Organization and Segregation

Badrinarayanan¹,

Le²,

Laub

2015

Annu. Rev. Cell Dev. Biol.

263

260

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If fully stretched out, a typical bacterial chromosome would be nearly one millimeter long, or approximately 1000 times the length of a cell. Not only must cells massively compact their genetic material, but they must also organize their DNA in a manner that is compatible with a range of cellular processes, including DNA replication, DNA repair, homologous recombination, and horizontal gene transfer. Recent work, driven in part by technological advances, has begun to reveal the general principles of chromosome organization in bacteria. Here, drawing on studies of many different organisms, we review the emerging picture of how bacterial chromosomes are structured at multiple length-scales, highlighting the functions of various DNA-binding proteins and impact of physical forces. Additionally, we discuss the spatial dynamics of chromosomes, particularly during their segregation to daughter cells. Although there has been tremendous progress, we also highlight gaps that remain in understanding chromosome organization and segregation.

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“…Sub-physiological levels of DnaA results in translocation of ori away from stalked pole In C. crescentus, the centromere is the first chromosomal locus to segregate away from the stalked pole [24]. Previous analyses of cells expressing sub-physiological levels of DnaA (not sufficient to initiate replication) revealed a DnaA-dependent and replication-independent segregation of the centromere [31]. Under these sub-physiological levels of DnaA, cells move the un-replicated centromere from the stalked pole to the new pole.…”

Section: Construction Of Indicator Strain With Fluorescently Labeled mentioning

confidence: 99%

“…Based on those results, we asked if the same subphysiological levels of DnaA could also trigger the movement of ori, independently of replication. To reach sub-physiological levels of DnaA in the cell, we used the same vanillate promoter to regulate dnaA expression [31]. Using our indicator strain PM500, we tracked the localization of ori using highresolution microscopy.…”

Section: Construction Of Indicator Strain With Fluorescently Labeled mentioning

confidence: 99%

“…With the goal to further understand what regulates the onset of these key cell cycle regulators (DnaA and ParA), we examined whether the onset of replication and segregation are restricted to ori-centromere's intrinsic localization, which in C. crescentus is near the stalked pole. To resolve this question, we genetically engineered a C. crescentus strain where movement of ori/centromere can be triggered in the absence of replication initiation [31]. Once ori was translocated to different sites over the cell, we tested for competency of replication initiation and chromosome segregation.…”

Section: Introductionmentioning

confidence: 99%

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Chromosome dynamics in bacteria: triggering replication at opposite location and segregation in opposite direction

Meléndez

Menikpurage

Mera

2019

Preprint

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The accurate onset of chromosome replication and segregation are fundamental for the survival of the cell. In bacteria, regulation of chromosome replication lies primarily at the initiation step. The bacterial replication initiator DnaA recognizes the origin of replication (ori) and opens this double stranded site allowing for the assembly of the DNA replication machinery. Following the onset of replication initiation, the partitioning protein ParA triggers the onset of chromosome segregation by direct interactions with ParB-bound to the centromere. The subcellular organization of ori and centromere are maintained after the completion of each cell cycle. It remains unclear what triggers the onset of these key chromosome regulators DnaA and ParA. One potential scenario is that the microenvironment of where the onset of replication and segregation take place hosts the regulators that trigger the activity of DnaA and ParA. In order to address this, we analyzed whether the activity of DnaA and ParA are restricted to only one site within the cell. In non-dividing cells of the alpha proteobacterium Caulobacter crescentus, ori and centromere are found near the stalked pole. To test DnaA's ability to initiate replication away from the stalked pole, we engineered a strain where movement of ori was induced in the absence of chromosome replication. Our data show that DnaA can initiate replication of the chromosome independently of the subcellular localization of ori. Furthermore, we discovered that the partitioning protein ParA was functional and could segregate the replicated centromere in the opposite direction from the new pole toward the stalked pole. We showed that the organization of the ParA gradient can be completely reconstructed in the opposite orientation by rearranging the location of the centromere. Our data reveal the high flexibility of the machineries that trigger the onset of chromosome replication and segregation in bacteria. Our work also provides insights into the coordination between replication and segregation with the cellular organization of specific chromosomal loci.

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Replication initiator DnaA binds at the Caulobacter centromere and enables chromosome segregation

Cited by 23 publications

References 38 publications

The DnaA inhibitor SirA acts in the same pathway as Soj (ParA) to facilitate oriC segregation during Bacillus subtilis sporulation

The DnaA inhibitor SirA acts in the same pathway as Soj (ParA) to facilitate oriC segregation during Bacillus subtilis sporulation

Bacterial Chromosome Organization and Segregation

Chromosome dynamics in bacteria: triggering replication at opposite location and segregation in opposite direction

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