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Whole-Genome Analysis Reveals That the Nucleoid Protein IHF Predominantly Binds to the Replication Origin oriC Specifically at the Time of Initiation

The structure and function of bacterial chromosomes are dynamically regulated by a wide variety of nucleoid-associated proteins (NAPs) and DNA superstructures, such as DNA supercoiling. In Escherichia coli, integration host factor (IHF), a NAP, binds to specific transcription promoters and regulator...

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Autores principales: Kasho, Kazutoshi, Oshima, Taku, Chumsakul, Onuma, Nakamura, Kensuke, Fukamachi, Kazuki, Katayama, Tsutomu
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8407004/
https://www.ncbi.nlm.nih.gov/pubmed/34475859
http://dx.doi.org/10.3389/fmicb.2021.697712
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author Kasho, Kazutoshi
Oshima, Taku
Chumsakul, Onuma
Nakamura, Kensuke
Fukamachi, Kazuki
Katayama, Tsutomu
author_facet Kasho, Kazutoshi
Oshima, Taku
Chumsakul, Onuma
Nakamura, Kensuke
Fukamachi, Kazuki
Katayama, Tsutomu
author_sort Kasho, Kazutoshi
collection PubMed
description The structure and function of bacterial chromosomes are dynamically regulated by a wide variety of nucleoid-associated proteins (NAPs) and DNA superstructures, such as DNA supercoiling. In Escherichia coli, integration host factor (IHF), a NAP, binds to specific transcription promoters and regulatory DNA elements of DNA replication such as the replication origin oriC: binding to these elements depends on the cell cycle but underlying mechanisms are unknown. In this study, we combined GeF-seq (genome footprinting with high-throughput sequencing) with synchronization of the E. coli cell cycle to determine the genome-wide, cell cycle-dependent binding of IHF with base-pair resolution. The GeF-seq results in this study were qualified enough to analyze genomic IHF binding sites (e.g., oriC and the transcriptional promoters of ilvG and osmY) except some of the known sites. Unexpectedly, we found that before replication initiation, oriC was a predominant site for stable IHF binding, whereas all other loci exhibited reduced IHF binding. To reveal the specific mechanism of stable oriC–IHF binding, we inserted a truncated oriC sequence in the terC (replication terminus) locus of the genome. Before replication initiation, stable IHF binding was detected even at this additional oriC site, dependent on the specific DnaA-binding sequence DnaA box R1 within the site. DnaA oligomers formed on oriC might protect the oriC–IHF complex from IHF dissociation. After replication initiation, IHF rapidly dissociated from oriC, and IHF binding to other sites was sustained or stimulated. In addition, we identified a novel locus associated with cell cycle-dependent IHF binding. These findings provide mechanistic insight into IHF binding and dissociation in the genome.
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spelling pubmed-84070042021-09-01 Whole-Genome Analysis Reveals That the Nucleoid Protein IHF Predominantly Binds to the Replication Origin oriC Specifically at the Time of Initiation Kasho, Kazutoshi Oshima, Taku Chumsakul, Onuma Nakamura, Kensuke Fukamachi, Kazuki Katayama, Tsutomu Front Microbiol Microbiology The structure and function of bacterial chromosomes are dynamically regulated by a wide variety of nucleoid-associated proteins (NAPs) and DNA superstructures, such as DNA supercoiling. In Escherichia coli, integration host factor (IHF), a NAP, binds to specific transcription promoters and regulatory DNA elements of DNA replication such as the replication origin oriC: binding to these elements depends on the cell cycle but underlying mechanisms are unknown. In this study, we combined GeF-seq (genome footprinting with high-throughput sequencing) with synchronization of the E. coli cell cycle to determine the genome-wide, cell cycle-dependent binding of IHF with base-pair resolution. The GeF-seq results in this study were qualified enough to analyze genomic IHF binding sites (e.g., oriC and the transcriptional promoters of ilvG and osmY) except some of the known sites. Unexpectedly, we found that before replication initiation, oriC was a predominant site for stable IHF binding, whereas all other loci exhibited reduced IHF binding. To reveal the specific mechanism of stable oriC–IHF binding, we inserted a truncated oriC sequence in the terC (replication terminus) locus of the genome. Before replication initiation, stable IHF binding was detected even at this additional oriC site, dependent on the specific DnaA-binding sequence DnaA box R1 within the site. DnaA oligomers formed on oriC might protect the oriC–IHF complex from IHF dissociation. After replication initiation, IHF rapidly dissociated from oriC, and IHF binding to other sites was sustained or stimulated. In addition, we identified a novel locus associated with cell cycle-dependent IHF binding. These findings provide mechanistic insight into IHF binding and dissociation in the genome. Frontiers Media S.A. 2021-08-12 /pmc/articles/PMC8407004/ /pubmed/34475859 http://dx.doi.org/10.3389/fmicb.2021.697712 Text en Copyright © 2021 Kasho, Oshima, Chumsakul, Nakamura, Fukamachi and Katayama. https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Microbiology
Kasho, Kazutoshi
Oshima, Taku
Chumsakul, Onuma
Nakamura, Kensuke
Fukamachi, Kazuki
Katayama, Tsutomu
Whole-Genome Analysis Reveals That the Nucleoid Protein IHF Predominantly Binds to the Replication Origin oriC Specifically at the Time of Initiation
title Whole-Genome Analysis Reveals That the Nucleoid Protein IHF Predominantly Binds to the Replication Origin oriC Specifically at the Time of Initiation
title_full Whole-Genome Analysis Reveals That the Nucleoid Protein IHF Predominantly Binds to the Replication Origin oriC Specifically at the Time of Initiation
title_fullStr Whole-Genome Analysis Reveals That the Nucleoid Protein IHF Predominantly Binds to the Replication Origin oriC Specifically at the Time of Initiation
title_full_unstemmed Whole-Genome Analysis Reveals That the Nucleoid Protein IHF Predominantly Binds to the Replication Origin oriC Specifically at the Time of Initiation
title_short Whole-Genome Analysis Reveals That the Nucleoid Protein IHF Predominantly Binds to the Replication Origin oriC Specifically at the Time of Initiation
title_sort whole-genome analysis reveals that the nucleoid protein ihf predominantly binds to the replication origin oric specifically at the time of initiation
topic Microbiology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8407004/
https://www.ncbi.nlm.nih.gov/pubmed/34475859
http://dx.doi.org/10.3389/fmicb.2021.697712
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