Novel Chromosome Organization Pattern in
            <i>Actinomycetales</i>
            —Overlapping Replication Cycles Combined with Diploidy

Böhm, Kati; Meyer, Fabian; Rhomberg, Agata; Kalinowski, Joern; Donovan, Catriona; Bramkamp, Marc

doi:10.1128/mbio.00511-17

Cited by 21 publications

(33 citation statements)

References 93 publications

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“…Recent work showed that M. smegmatis compensates for its elongated replication time (more than 2 h) by initiating chromosome replication before cytokinesis (26) and having a relatively large fraction of cells (up to 15%) undergoing multifork chromosome replication (28). In this, M. smegmatis resembles other actinobacterial species, such as Corynebacterium glutamicum (54). Our experiments revealed that, in mycobacteria, the initiation of the next replication round and the start of septum formation precede sister chromosome separation by ϳ20 min and 40 min, respectively (Fig.…”

Section: Discussionmentioning

confidence: 99%

The Origin of Chromosomal Replication Is Asymmetrically Positioned on the Mycobacterial Nucleoid, and the Timing of Its Firing Depends on HupB

et al. 2018

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The bacterial chromosome undergoes dynamic changes in response to ongoing cellular processes and adaptation to environmental conditions. Among the many proteins involved in maintaining this dynamism, the most abundant is the nucleoid-associated protein (NAP) HU. In mycobacteria, the HU homolog, HupB, possesses an additional C-terminal domain that resembles that of eukaryotic histones H1/H5. Recently, we demonstrated that the highly abundant HupB protein occupies the entirety of the chromosome and that the HupB-binding sites exhibit a bias from the origin () to the terminus (). In this study, we used HupB fused with enhanced green fluorescent protein (EGFP) to perform the first analysis of chromosome dynamics and to track the and replication machinery directly on the chromosome during the mycobacterial cell cycle. We show that the chromosome is located in an off-center position that reflects the unequal division and growth of mycobacterial cells. Moreover, unlike the situation in, the sister regions of move asymmetrically along the mycobacterial nucleoid. Interestingly, in this slow-growing organism, the initiation of the next round of replication precedes the physical separation of sister chromosomes. Finally, we show that HupB is involved in the precise timing of replication initiation. Although our view of mycobacterial nucleoid organization has evolved considerably over time, we still know little about the dynamics of the mycobacterial nucleoid during the cell cycle. HupB is a highly abundant mycobacterial nucleoid-associated protein (NAP) with an indispensable histone-like tail. It was previously suggested as a potential target for antibiotic therapy against tuberculosis. Here, we fused HupB with enhanced green fluorescent protein (EGFP) to study the dynamics of the mycobacterial chromosome in real time and to monitor the replication process directly on the chromosome. Our results reveal that, unlike the situation in , the nucleoid of an apically growing mycobacterium is positioned asymmetrically within the cell throughout the cell cycle. We show that HupB is involved in controlling the timing of replication initiation. Since tuberculosis remains a serious health problem, studies concerning mycobacterial cell biology are of great importance.

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Section: Discussionmentioning

confidence: 99%

The Origin of Chromosomal Replication Is Asymmetrically Positioned on the Mycobacterial Nucleoid, and the Timing of Its Firing Depends on HupB

et al. 2018

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“…Firstly, cellular localization of fluorescent ParB-eYFP foci is similar to wild type, positioning at cell poles and migrating to the newly formed septa (Fig. 2B, S3) 40 . Interestingly, the combination of a single parS site with ParB-eYFP resulted in 7% anucleoid mini-cells (Fig.…”

Section: Resultsmentioning

confidence: 65%

“…Previous studies on C. glutamicum chromosome partitioning have revealed two stable ParB- oriC clusters at each cell pole, while newly replicated origins are segregated towards a division septa formed at midcell 40 . In B. subtilis , C. crescentus and P. aeruginosa ParAB-mediated chromosome segregation and folding depends on parS sites 18,19,21 .…”

Section: Resultsmentioning

confidence: 93%

“…For more accurate cluster estimations, PALM analysis was performed using slow-growing cells resulting in significantly fewer ParB macro-clusters per cell (Fig. S6B) 40 . Since segregation of oriC complexes might alter their DNA compaction, we focused on the two largest macroclusters per cell, stably tethered at cell poles.…”

Section: Resultsmentioning

confidence: 99%

“…C. glutamicum is a polar growing actinobacterium, whose genome encodes both SMC/ScpAB and MksBEFG. In this species, the two oriC s are continuously associated with the polar scaffold protein DivIVA, while newly replicated sister oriC s segregate towards division septa via the ParABS system 40–42 . In contrast to B. subtilis , C. glutamicum ParAB are by themselves crucially important drivers of reliable nucleoid separation prior to cell division, where ParAB deletions yield in 20 % of anucleoid cells 43–45 .…”

Section: Introductionmentioning

confidence: 99%

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Chromosome organization by a conserved condensin-ParB system in the actinobacteriumCorynebacterium glutamicum

Boehm

Giacomelli

Schmidt

et al. 2019

Preprint

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20Higher-order chromosome folding and segregation is tightly regulated in all domains 21 of life. In bacteria, details on nucleoid organization regulatory mechanisms and function 22 remains poorly characterized, especially in non-model species. Here, we investigate 23 the role of DNA partitioning protein ParB and condensin complexes, two key players 24 in bacterial chromosome structuring, in the actinobacterium Corynebacterium 25 glutamicum. Chromosome conformation capture reveals SMC-mediated long-range 26 interactions around ten centromere-like parS sites clustered at the replication origin 27 (oriC). At least one oriC-proximal parS site is necessary for a reliable chromosome 28 segregation. Using a combination of chromatin immunoprecipitation and 29 photoactivated single molecule localization microscopy evidences the formation of 30 distinct ParB-nucleoprotein subclusters in dependence of parS numbers. We further 31 identified and functionally characterized two condensin paralogs. Whereas 32 SMC/ScpAB complexes are loaded via ParB at parS sites mediating chromosomal 33 inter-arm contacts like in Bacillus subtilis, the MukBEF-like SMC complex MksBEFG 34 does not contribute to chromosomal DNA-folding. Rather, the MksBEFG complex is 35 involved in plasmid maintenance and interacts with the polar oriC-tethering factor 36 DivIVA. These data complement current models of ParB-SMC/ScpAB crosstalk, while 37 showing that some condensin complexes evolved functions uncoupled from 38 chromosome folding. 39 Keywords 40 ParB, parS, SMC, MksB, condensin, actinobacteria, chromosome conformation 41 capture, chromatin immunoprecipitation, super-resolution microscopy, DNA 42 segregation 43 44 can cause severe DNA segregation phenotypes 18,19 . To date, only few studies 71 investigated the impact of chromosomal parS localization on DNA-segregation and 72 folding 18-22 . 73 In addition to ParABS systems, most bacteria harbor condensin complexes, members 74 of the structural maintenance of chromosomes (SMC) family of proteins found in all 75 kingdoms of life 23 . In standard model organisms, condensins are equally essential for 76 faithful chromosome segregation by compacting DNA into separate nucleoids 24-26 . The 77 SMC/ScpAB (structural maintenance of chromosomes) complex is well-studied in B. 78 subtilis, where it consists of two large SMC subunits and the kleisin ScpA associated 79 with dimeric accessory protein ScpB that assemble into a ring-like structure 27 . A recent 80 study suggests progressive extrusion of condensin-encircled DNA loops upon 81 conformational changes in the SMC subunit, which leads to a gradual size increase of 82 trapped DNA molecules 28 . The active process(es) driving DNA extrusion 29,30 allow(s) 83 for translocation along the chromosome with velocities of around 50 Kb/ min 31 , and 84 depend(s) on the ATPase activity of SMC 32 . To be loaded on parS sites, SMC/ScpAB 85 complexes necessitate ParB 20,22,33,34 . They redistribute to distant chromosomal 86 regions, promoting topological changes, and notab...

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Chromosome Organization and Cell Growth of Corynebacterium glutamicum

Böhm

Giacomelli

Bramkamp

2020

Corynebacterium Glutamicum

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Novel Chromosome Organization Pattern in Actinomycetales —Overlapping Replication Cycles Combined with Diploidy

Cited by 21 publications

References 93 publications

The Origin of Chromosomal Replication Is Asymmetrically Positioned on the Mycobacterial Nucleoid, and the Timing of Its Firing Depends on HupB

The Origin of Chromosomal Replication Is Asymmetrically Positioned on the Mycobacterial Nucleoid, and the Timing of Its Firing Depends on HupB

Chromosome organization by a conserved condensin-ParB system in the actinobacteriumCorynebacterium glutamicum

Chromosome Organization and Cell Growth of Corynebacterium glutamicum

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