ClbR Is the Key Transcriptional Activator of Colibactin Gene Expression in <named-content content-type="genus-species">Escherichia coli</named-content>

ABSTRACT Colibactin is a nonribosomal peptide/polyketide hybrid natural product expressed by different members of the Enterobacteriaceae which can be correlated with induction of DNA double-strand breaks and interference with cell cycle progression in eukaryotes. Regulatory features of colibactin ex...

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Autores principales:	Alexander Wallenstein, Nadine Rehm, Marina Brinkmann, Martina Selle, Nadège Bossuet-Greif, Daniel Sauer, Boyke Bunk, Cathrin Spröer, Haleluya Tesfaye Wami, Stefan Homburg, Rudolf von Bünau, Simone König, Jean-Philippe Nougayrède, Jörg Overmann, Eric Oswald, Rolf Müller, Ulrich Dobrindt
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2020
Materias:	secondary metabolite polyketide cytopathic effect RNA-seq VNTR Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/ac9db74a37844ece934f88a379e771a3
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spelling	oai:doaj.org-article:ac9db74a37844ece934f88a379e771a32021-11-15T15:30:51ZClbR Is the Key Transcriptional Activator of Colibactin Gene Expression in <named-content content-type="genus-species">Escherichia coli</named-content>10.1128/mSphere.00591-202379-5042https://doaj.org/article/ac9db74a37844ece934f88a379e771a32020-08-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mSphere.00591-20https://doaj.org/toc/2379-5042ABSTRACT Colibactin is a nonribosomal peptide/polyketide hybrid natural product expressed by different members of the Enterobacteriaceae which can be correlated with induction of DNA double-strand breaks and interference with cell cycle progression in eukaryotes. Regulatory features of colibactin expression are only incompletely understood. We used Escherichia coli strain M1/5 as a model to investigate regulation of expression of the colibactin determinant at the transcriptional level and to characterize regulatory elements located within the colibactin pathogenicity island itself. We measured clbR transcription in vitro and observed that cultivation in defined minimal media led to increased colibactin expression relative to rich media. Transcription of clbR directly responds to iron availability. We also characterized structural DNA elements inside the colibactin determinant involved in ClbR-dependent regulation, i.e., ClbR binding sites and a variable number of tandem repeats located upstream of clbR. We investigated the impact of clbR overexpression or deletion at the transcriptome and proteome levels. Moreover, we compared global gene regulation under these conditions with that occurring upon overexpression or deletion of clbQ, which affects the flux of colibactin production. Combining the results of the transcriptome and proteome analyses with indirect measurements of colibactin levels by cell culture assays and an approximate quantification of colibactin via the second product of colibactin cleavage from precolibactin, N-myristoyl-d-asparagine, we demonstrate that the variable number of tandem repeats plays a significant regulatory role in colibactin expression. We identify ClbR as the only transcriptional activator known so far that is specific and essential for efficient regulation of colibactin production. IMPORTANCE The nonribosomal peptide/polyketide hybrid colibactin can be considered a bacterial virulence factor involved in extraintestinal infection and also a procarcinogen. Nevertheless, and despite its genotoxic effect, colibactin expression can also inhibit bacterial or tumor growth and correlates with probiotic anti-inflammatory and analgesic properties. Although the biological function of this natural compound has been studied extensively, our understanding of the regulation of colibactin expression is still far from complete. We investigated in detail the role of regulatory elements involved in colibactin expression and in the growth conditions that promote colibactin expression. In this way, our data shed light on the regulatory mechanisms involved in colibactin expression and may support the expression and purification of this interesting nonribosomal peptide/polyketide hybrid for further molecular characterization.Alexander WallensteinNadine RehmMarina BrinkmannMartina SelleNadège Bossuet-GreifDaniel SauerBoyke BunkCathrin SpröerHaleluya Tesfaye WamiStefan HomburgRudolf von BünauSimone KönigJean-Philippe NougayrèdeJörg OvermannEric OswaldRolf MüllerUlrich DobrindtAmerican Society for Microbiologyarticlesecondary metabolitepolyketidecytopathic effectRNA-seqVNTRMicrobiologyQR1-502ENmSphere, Vol 5, Iss 4 (2020)
institution	DOAJ
collection	DOAJ
language	EN
topic	secondary metabolite polyketide cytopathic effect RNA-seq VNTR Microbiology QR1-502
spellingShingle	secondary metabolite polyketide cytopathic effect RNA-seq VNTR Microbiology QR1-502 Alexander Wallenstein Nadine Rehm Marina Brinkmann Martina Selle Nadège Bossuet-Greif Daniel Sauer Boyke Bunk Cathrin Spröer Haleluya Tesfaye Wami Stefan Homburg Rudolf von Bünau Simone König Jean-Philippe Nougayrède Jörg Overmann Eric Oswald Rolf Müller Ulrich Dobrindt ClbR Is the Key Transcriptional Activator of Colibactin Gene Expression in <named-content content-type="genus-species">Escherichia coli</named-content>
description	ABSTRACT Colibactin is a nonribosomal peptide/polyketide hybrid natural product expressed by different members of the Enterobacteriaceae which can be correlated with induction of DNA double-strand breaks and interference with cell cycle progression in eukaryotes. Regulatory features of colibactin expression are only incompletely understood. We used Escherichia coli strain M1/5 as a model to investigate regulation of expression of the colibactin determinant at the transcriptional level and to characterize regulatory elements located within the colibactin pathogenicity island itself. We measured clbR transcription in vitro and observed that cultivation in defined minimal media led to increased colibactin expression relative to rich media. Transcription of clbR directly responds to iron availability. We also characterized structural DNA elements inside the colibactin determinant involved in ClbR-dependent regulation, i.e., ClbR binding sites and a variable number of tandem repeats located upstream of clbR. We investigated the impact of clbR overexpression or deletion at the transcriptome and proteome levels. Moreover, we compared global gene regulation under these conditions with that occurring upon overexpression or deletion of clbQ, which affects the flux of colibactin production. Combining the results of the transcriptome and proteome analyses with indirect measurements of colibactin levels by cell culture assays and an approximate quantification of colibactin via the second product of colibactin cleavage from precolibactin, N-myristoyl-d-asparagine, we demonstrate that the variable number of tandem repeats plays a significant regulatory role in colibactin expression. We identify ClbR as the only transcriptional activator known so far that is specific and essential for efficient regulation of colibactin production. IMPORTANCE The nonribosomal peptide/polyketide hybrid colibactin can be considered a bacterial virulence factor involved in extraintestinal infection and also a procarcinogen. Nevertheless, and despite its genotoxic effect, colibactin expression can also inhibit bacterial or tumor growth and correlates with probiotic anti-inflammatory and analgesic properties. Although the biological function of this natural compound has been studied extensively, our understanding of the regulation of colibactin expression is still far from complete. We investigated in detail the role of regulatory elements involved in colibactin expression and in the growth conditions that promote colibactin expression. In this way, our data shed light on the regulatory mechanisms involved in colibactin expression and may support the expression and purification of this interesting nonribosomal peptide/polyketide hybrid for further molecular characterization.
format	article
author	Alexander Wallenstein Nadine Rehm Marina Brinkmann Martina Selle Nadège Bossuet-Greif Daniel Sauer Boyke Bunk Cathrin Spröer Haleluya Tesfaye Wami Stefan Homburg Rudolf von Bünau Simone König Jean-Philippe Nougayrède Jörg Overmann Eric Oswald Rolf Müller Ulrich Dobrindt
author_facet	Alexander Wallenstein Nadine Rehm Marina Brinkmann Martina Selle Nadège Bossuet-Greif Daniel Sauer Boyke Bunk Cathrin Spröer Haleluya Tesfaye Wami Stefan Homburg Rudolf von Bünau Simone König Jean-Philippe Nougayrède Jörg Overmann Eric Oswald Rolf Müller Ulrich Dobrindt
author_sort	Alexander Wallenstein
title	ClbR Is the Key Transcriptional Activator of Colibactin Gene Expression in <named-content content-type="genus-species">Escherichia coli</named-content>
title_short	ClbR Is the Key Transcriptional Activator of Colibactin Gene Expression in <named-content content-type="genus-species">Escherichia coli</named-content>
title_full	ClbR Is the Key Transcriptional Activator of Colibactin Gene Expression in <named-content content-type="genus-species">Escherichia coli</named-content>
title_fullStr	ClbR Is the Key Transcriptional Activator of Colibactin Gene Expression in <named-content content-type="genus-species">Escherichia coli</named-content>
title_full_unstemmed	ClbR Is the Key Transcriptional Activator of Colibactin Gene Expression in <named-content content-type="genus-species">Escherichia coli</named-content>
title_sort	clbr is the key transcriptional activator of colibactin gene expression in <named-content content-type="genus-species">escherichia coli</named-content>
publisher	American Society for Microbiology
publishDate	2020
url	https://doaj.org/article/ac9db74a37844ece934f88a379e771a3
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ClbR Is the Key Transcriptional Activator of Colibactin Gene Expression in <named-content content-type="genus-species">Escherichia coli</named-content>

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