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Shogo Ozaki Last modified date:2024.06.03



Undergraduate School


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Homepage
https://kyushu-u.elsevierpure.com/en/persons/shogo-ozaki
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https://researchmap.jp/read0142745
http://bunsei.phar.kyushu-u.ac.jp/BUNSEI-ENG.html
Phone
092-642-6643
Academic Degree
Ph.D
Country of degree conferring institution (Overseas)
No
Field of Specialization
Molecular biology, Microbiology, Genetics
Total Priod of education and research career in the foreign country
06years00months
Outline Activities
Growing bacteria require careful tuning of cell division processes with dynamic organization of replicating chromosomes. The objective of our project is to understand molecular mechanisms for coordination of chromosome replication with division. To achieve this we make use of different model bacteria including Escherichia coli, Caulobacter crescentus, and Thermotoga maritima.
Research
Research Interests
  • Mechanism and regulation for initiation complex that promotes local duplex DNA unwinding at replication origin during the initiation of chromosomal replication
    keyword : DNA replication, cell cycle, DnaA, AAA+, conformational change
    2005.10.
Academic Activities
Reports
1. Katayama, T, Ozaki, S, Keyamura, K, Fujimitsu, K., Regulation of the replication cycle: conserved and diverse regulatory systems for DnaA and oriC, Nat Rev Microbiol, 2010, 2010.03.
2. Ozaki S., Katayama T., DnaA structure, function, and dynamics in the initiation at the chromosomal origin., Plasmid, 2009.07.
Papers
1. Chuyuan Lu, Ryusei Yoshida, Tsutomu Katayama, Shogo Ozaki, Thermotoga maritima oriC involves a DNA unwinding element with distinct modules and a DnaA-oligomerizing region with a novel directional binding mode, Journal of Biological Chemistry, 10.1016/j.jbc.2023.104888, 104888-104888, 2023.07, Initiation of chromosomal replication requires dynamic nucleoprotein complexes. In most eubacteria, the origin oriC contains multiple DnaA box sequences to which the ubiquitous DnaA initiators bind. In Escherichia coli oriC, DnaA boxes sustain construction of higher-order complexes via DnaA-DnaA interactions, promoting the unwinding of the DNA unwinding element (DUE) within oriC and concomitantly binding the single-stranded DUE to install replication machinery. Despite the significant sequence homologies among DnaA proteins, bacterial oriC sequences are highly diverse. The present study investigated the design of oriC (tma-oriC) from Thermotoga maritima, an evolutionarily ancient eubacterium. The minimal tma-oriC sequence includes a DUE and a flanking region containing five DnaA boxes recognized by the cognate DnaA initiator (tmaDnaA). This DUE was comprised of two distinct functional modules, an unwinding module and a tmaDnaA-binding module. Three direct repeats of the trinucleotide TAG within DUE were essential for both unwinding and single-stranded DUE binding by tmaDnaA complexes constructed on the DnaA boxes. Its surrounding AT-rich sequences stimulated only duplex unwinding. Moreover, head-to-tail oligomers of ATP-bound tmaDnaA were constructed within tma-oriC, irrespective of the directions of the DnaA boxes. This binding mode was considered to be induced by flexible swiveling of DnaA domains III and IV, which were responsible for DnaA-DnaA interactions and DnaA box binding, respectively. Phasing of specific tmaDnaA boxes in tma-oriC DNA was also responsible for unwinding. These findings indicate that a single-stranded DUE recruitment mechanism was responsible for unwinding, and would enhance understanding of the fundamental molecular nature of the origin sequences present in evolutionarily divergent bacteria..
2. Shogo Ozaki, Dengyu Wang, Yasutaka Wakasugi, Naoto Itani, Tsutomu Katayama, The Caulobacter crescentus DciA promotes chromosome replication through topological loading of the DnaB replicative helicase at replication forks, Nucleic Acids Research, 10.1093/nar/gkac1146, 2022.12, Abstract The replicative DNA helicase translocates on single-stranded DNA to drive replication forks during chromosome replication. In most bacteria the ubiquitous replicative helicase, DnaB, co-evolved with the accessory subunit DciA, but how they function remains incompletely understood. Here, using the model bacterium Caulobacter crescentus, we demonstrate that DciA plays a prominent role in DNA replication fork maintenance. Cell cycle analyses using a synchronized Caulobacter cell population showed that cells devoid of DciA exhibit a severe delay in fork progression. Biochemical characterization revealed that the DnaB helicase in its default state forms a hexamer that inhibits self-loading onto single-stranded DNA. We found that upon binding to DciA, the DnaB hexamer undergoes conformational changes required for encircling single-stranded DNA, thereby establishing the replication fork. Further investigation of the functional structure of DciA revealed that the C-terminus of DciA includes conserved leucine residues responsible for DnaB binding and is essential for DciA in vivo functions. We propose that DciA stimulates loading of DnaB onto single strands through topological isomerization of the DnaB structure, thereby ensuring fork progression. Given that the DnaB-DciA modules are widespread among eubacterial species, our findings suggest that a common mechanism underlies chromosome replication..
3. Shogo Ozaki, Yasutaka Wakasugi, Tsutomu Katayama, Z-Ring-Associated Proteins Regulate Clustering of the Replication Terminus-Binding Protein ZapT in Caulobacter crescentus, mBio, 10.1128/mBio.02196-20, 2021.01.
4. Shogo Ozaki, Urs Jenal, Tsutomu Katayama, Novel divisome-associated protein spatially coupling the z-ring with the chromosomal replication terminus in caulobacter crescentus, mBio, 10.1128/mBio.00487-20, 11, 2, 2020.03, Cell division requires proper spatial coordination with the chromosome, which undergoes dynamic changes during chromosome replication and segregation. FtsZ is a bacterial cytoskeletal protein that assembles into the Z-ring, providing a platform to build the cell division apparatus. In the model bacterium Caulobacter crescentus, the cellular localization of the Z-ring is controlled during the cell cycle in a chromosome replication-coupled manner. Although dynamic localization of the Z-ring at midcell is driven primarily by the replication origin-associated FtsZ inhibitor MipZ, the mechanism ensuring accurate positioning of the Z-ring remains unclear. In this study, we showed that the Z-ring colocalizes with the replication terminus region, located opposite the origin, throughout most of the C. crescentus cell cycle. Spatial organization of the two is mediated by ZapT, a previously uncharacterized protein that inter-acts with the terminus region and associates with ZapA and ZauP, both of which are part of the incipient division apparatus. While the Z-ring and the terminus region coin-cided with the presence of ZapT, colocalization of the two was perturbed in cells lacking zapT, which is accompanied by delayed midcellular positioning of the Z-ring. Moreover, cells overexpressing ZapT showed compromised positioning of the Z-ring and MipZ. These findings underscore the important role of ZapT in controlling cell division pro-cesses. We propose that ZapT acts as a molecular bridge that physically links the terminus region to the Z-ring, thereby ensuring accurate site selection for the Z-ring. Because ZapT is conserved in proteobacteria, these findings may define a general mechanism coordinating cell division with chromosome organization. IMPORTANCE Growing bacteria require careful tuning of cell division processes with dynamic organization of replicating chromosomes. In enteric bacteria, ZapA associates with the cytoskeletal Z-ring and establishes a physical linkage to the chromosomal replication terminus through its interaction with ZapB-MatP-DNA complexes. However, because ZapB and MatP are found only in enteric bacteria, it remains unclear how the Z-ring and the terminus are coordinated in the vast majority of bacteria. Here, we provide evidence that a novel conserved protein, termed ZapT, mediates colocalization of the Z-ring with the terminus in Caulobacter crescentus, a model organism that is phylo-genetically distant from enteric bacteria. Given that ZapT facilitates cell division processes in C. crescentus, this study highlights the universal importance of the physical linkage between the Z-ring and the terminus in maintaining cell integrity..
Presentations
1. Shogo Ozaki, Chuyuan Lu, Ryusei Yoshida, Yasutaka Wakasugi, Shohei Sato, Tsutomu Katayama, A common mechanism and architecture of the dynamic nucleoprotein complex formed at the origin DNA of eubacterial chromosome replication, 第46回日本分子生物学会年会, 2023.12, Initiation of chromosome replication requires dynamic nucleoprotein complexes to establish replication forks. In Escherichia coli, the origin, oriC, comprises an AT-rich DNA unwinding element (DUE) and asymmetric DnaA box sequences guiding the DnaA initiator to form a highly ordered complex. This initiation complex promotes DUE unwinding and concomitantly binds the resultant single-stranded DUE in a sequence-specific manner, stabilizing the unwound state and facilitating assembly of the replisome components. While DnaA homologs are ubiquitous in eubacteria, the origin sequences differ significantly in respect with the number, direction, and spatial arrangement of DnaA boxes and DUE sequences, limiting our understanding of the general principles governing the process of DUE unwinding. Here, we investigate molecular mechanisms of a prototypic DnaA-oriC complex in the evolutionarily ancient hyperthermophile Thermotoga maritima. We reveal that the DnaA protein from this bacterium retains the ability to form an initiation complex on the cognate oriC, irrespective of DnaA box directions. This flexibility in DnaA assembly is likely driven by substantial swiveling of the DnaA box-binding domain relative to the DnaA AAA+ domain supporting DnaA-DnaA interaction. The resultant DnaA-oriC complexes bind to the three direct repeats of the trinucleotide TAG within single-stranded DUE, stabilizing DUE unwinding (Lu et al. JBC 2023). These findings provide insights into the conservation of fundamental mechanisms for DUE unwinding and single-stranded DUE binding by DnaA proteins across eubacteria, supporting the tunable nature of origin sequences in constructing the initiation complex.
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2. Shogo Ozaki,Chuyuan Lu, Ryusei Yoshida, Yasutaka Wakasugi, Tsutomu Katayama, A common insight into mechanisms of duplex unwinding at the origin DNA during eubacterial chromosome replication , 第96回日本生化学会大会ワークショップ「染色体DNA複製開始複合体と開始制御メカニズムの新たな展望」, 2023.11.
3. Shogo Ozaki, The mechanism of chromosome replication in the alpha-proteobacterium Caulobacter crescentus., Research seminar, 2023.06.
4. Shogo Ozaki, Dengyu Wang, Yasutaka Wakasugi, Naoto Itani, and Tsutomu Katayama, Analysis on the loading mechanism of the bacterial replicative DnaB helicase in the alpha-proteobacterium Caulobacter crescentus., NIG International Symposium 2022, 2022.10.
5. Shogo Ozaki,Tsutomu Katayama, Analysis of the chromosomal replication mechanism in the eubacterium Caulobacter crescentus, 日本分子生物学会第44回大会, 2021.12.
6. Shogo Ozaki、Yasutaka Wakasugi、Urs Jenal, Tsutomu Katayama, A novel divisome-associated protein spatially couples the Z-ring with the chromosomal replication terminus in Caulobacter crescentus, CAULOCONFERENCE 2020, 2020.04.
7. 尾崎 省吾, 若杉泰敬, 片山 勉, Identification and characterization of a novel DNA binding protein that colocalizes the chromosome replication terminus with cell division apparatus in Caulobacter crescentus, 第42回日本分子生物学会年会, 2019.12, The bacterial chromosomes are spatially and timely organized to execute chromoso
me replication and segregation in a manner coupled to cell division. In the gram
-negative, alphaproteobacterium Caulobacter crescentus, subcellular posit
ioning of the individual chromosomal loci alters dynamically during chromosome r
eplication. In a pre-replication stage, the origin of chromosome replication is
sequestered to one pole of the cell while the terminus region is localized at th
e other cell pole. Upon initiation of chromosome replication, one of the newly-r
eplicated origin DNAs is driven toward the other cell pole to establish bipolar
localization of the origin DNAs. Concomitantly, the terminus region is recruited
to the midcell position where the bacterial tubulin homolog FtsZ colocalizes to
form cytokinetic Z-rings. Although this colocalization likely coordinates cell
division with dynamic chromosome architecture, the molecular basis for a physica
l link between the Z-rings and the terminus region remains unknown. In this stud
y, we identify the terminus-interactive DNA binding protein ZapT and show that i
t directly binds to a component associated with the Z-rings. Moreover, the za
pT
gene is crucial to maintain normal cell division control in C. crescentus
. Together, we propose that ZapT acts as a molecular bridge that physically link
s the terminus region to the Z-rings, thereby ensuring precise processes in chro
mosome segregation and cell division. Because ZapT orthologs are conserved among
diverse proteobacterial species, our findings may represent a general mechanism
to coordinate cell division with chromosome organization in time and space..
8. Shogo Ozaki, Lori Christian, Urs Jenal, The second messenger signaling drives chromosome replication in the asymmetrically dividing bacterium Caulobacter crescentus, 第56回日本生物物理学会年会, 2018.09.
9. Analysis on the minimal functional structure of the DnaA complex for the regulation of duplex DNA unwinding.
10. Analysis on the minimal functional elements of the replication origin in duplex DNA unwinding by DnaA.
Membership in Academic Society
  • American Society for Microbiology
  • The Japanese Biochemical Society
  • The Genetics Society of Japan
  • The Molecular Biology Society of Japan
Awards
  • Best Papers Awards in the 80th Annual Meeting of the Genetic Society of Japan
  • Best Papers Awards in the 79th Annual Meeting of the Genetic Society of Japan