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Identification of σ(E)-Dependent Promoter Upstream of clpB from the Pathogenic Spirochaete Leptospira interrogans by Applying an E. coli Two-Plasmid System

There is limited information on gene expression in the pathogenic spirochaete Leptospira interrogans and genetic mechanisms controlling its virulence. Transcription is the first step in gene expression that is often determined by environmental effects, including infection-induced stresses. Alteratio...

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Autores principales: Kędzierska-Mieszkowska, Sabina, Potrykus, Katarzyna, Arent, Zbigniew, Krajewska, Joanna
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2019
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6941012/
https://www.ncbi.nlm.nih.gov/pubmed/31847479
http://dx.doi.org/10.3390/ijms20246325
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author Kędzierska-Mieszkowska, Sabina
Potrykus, Katarzyna
Arent, Zbigniew
Krajewska, Joanna
author_facet Kędzierska-Mieszkowska, Sabina
Potrykus, Katarzyna
Arent, Zbigniew
Krajewska, Joanna
author_sort Kędzierska-Mieszkowska, Sabina
collection PubMed
description There is limited information on gene expression in the pathogenic spirochaete Leptospira interrogans and genetic mechanisms controlling its virulence. Transcription is the first step in gene expression that is often determined by environmental effects, including infection-induced stresses. Alterations in the environment result in significant changes in the transcription of many genes, allowing effective adaptation of Leptospira to mammalian hosts. Thus, promoter and transcriptional start site identification are crucial for determining gene expression regulation and for the understanding of genetic regulatory mechanisms existing in Leptospira. Here, we characterized the promoter region of the L. interrogans clpB gene (clpB(Li)) encoding an AAA(+) molecular chaperone ClpB essential for the survival of this spirochaete under thermal and oxidative stresses, and also during infection of the host. Primer extension analysis demonstrated that transcription of clpB in L. interrogans initiates at a cytidine located 41 bp upstream of the ATG initiation codon, and, to a lesser extent, at an adenine located 2 bp downstream of the identified site. Transcription of both transcripts was heat-inducible. Determination of clpB(Li) transcription start site, combined with promoter transcriptional activity assays using a modified two-plasmid system in E. coli, revealed that clpB(Li) transcription is controlled by the ECF σ(E) factor. Of the ten L. interrogans ECF σ factors, the factor encoded by LIC_12757 (LA0876) is most likely to be the key regulator of clpB gene expression in Leptospira cells, especially under thermal stress. Furthermore, clpB expression may be mediated by ppGpp in Leptospira.
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spelling pubmed-69410122020-01-09 Identification of σ(E)-Dependent Promoter Upstream of clpB from the Pathogenic Spirochaete Leptospira interrogans by Applying an E. coli Two-Plasmid System Kędzierska-Mieszkowska, Sabina Potrykus, Katarzyna Arent, Zbigniew Krajewska, Joanna Int J Mol Sci Article There is limited information on gene expression in the pathogenic spirochaete Leptospira interrogans and genetic mechanisms controlling its virulence. Transcription is the first step in gene expression that is often determined by environmental effects, including infection-induced stresses. Alterations in the environment result in significant changes in the transcription of many genes, allowing effective adaptation of Leptospira to mammalian hosts. Thus, promoter and transcriptional start site identification are crucial for determining gene expression regulation and for the understanding of genetic regulatory mechanisms existing in Leptospira. Here, we characterized the promoter region of the L. interrogans clpB gene (clpB(Li)) encoding an AAA(+) molecular chaperone ClpB essential for the survival of this spirochaete under thermal and oxidative stresses, and also during infection of the host. Primer extension analysis demonstrated that transcription of clpB in L. interrogans initiates at a cytidine located 41 bp upstream of the ATG initiation codon, and, to a lesser extent, at an adenine located 2 bp downstream of the identified site. Transcription of both transcripts was heat-inducible. Determination of clpB(Li) transcription start site, combined with promoter transcriptional activity assays using a modified two-plasmid system in E. coli, revealed that clpB(Li) transcription is controlled by the ECF σ(E) factor. Of the ten L. interrogans ECF σ factors, the factor encoded by LIC_12757 (LA0876) is most likely to be the key regulator of clpB gene expression in Leptospira cells, especially under thermal stress. Furthermore, clpB expression may be mediated by ppGpp in Leptospira. MDPI 2019-12-15 /pmc/articles/PMC6941012/ /pubmed/31847479 http://dx.doi.org/10.3390/ijms20246325 Text en © 2019 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Kędzierska-Mieszkowska, Sabina
Potrykus, Katarzyna
Arent, Zbigniew
Krajewska, Joanna
Identification of σ(E)-Dependent Promoter Upstream of clpB from the Pathogenic Spirochaete Leptospira interrogans by Applying an E. coli Two-Plasmid System
title Identification of σ(E)-Dependent Promoter Upstream of clpB from the Pathogenic Spirochaete Leptospira interrogans by Applying an E. coli Two-Plasmid System
title_full Identification of σ(E)-Dependent Promoter Upstream of clpB from the Pathogenic Spirochaete Leptospira interrogans by Applying an E. coli Two-Plasmid System
title_fullStr Identification of σ(E)-Dependent Promoter Upstream of clpB from the Pathogenic Spirochaete Leptospira interrogans by Applying an E. coli Two-Plasmid System
title_full_unstemmed Identification of σ(E)-Dependent Promoter Upstream of clpB from the Pathogenic Spirochaete Leptospira interrogans by Applying an E. coli Two-Plasmid System
title_short Identification of σ(E)-Dependent Promoter Upstream of clpB from the Pathogenic Spirochaete Leptospira interrogans by Applying an E. coli Two-Plasmid System
title_sort identification of σ(e)-dependent promoter upstream of clpb from the pathogenic spirochaete leptospira interrogans by applying an e. coli two-plasmid system
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6941012/
https://www.ncbi.nlm.nih.gov/pubmed/31847479
http://dx.doi.org/10.3390/ijms20246325
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