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Analysis of a taurine-dependent promoter in Sinorhizobium meliloti that offers tight modulation of gene expression
BACKGROUND: Genetic models have been developed in divergent branches of the class Alphaproteobacteria to help answer a wide spectrum of questions regarding bacterial physiology. For example, Sinorhizobium meliloti serves as a useful representative for investigating rhizobia-plant symbiosis and nitro...
Autores principales: | , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
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BioMed Central
2014
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4254191/ https://www.ncbi.nlm.nih.gov/pubmed/25420869 http://dx.doi.org/10.1186/s12866-014-0295-2 |
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author | Mostafavi, Mina Lewis, Jainee Christa Saini, Tanisha Bustamante, Julian Albert Gao, Ivan Thomas Tran, Tuyet Thi King, Sean Nicholas Huang, Zhenzhong Chen, Joseph C |
author_facet | Mostafavi, Mina Lewis, Jainee Christa Saini, Tanisha Bustamante, Julian Albert Gao, Ivan Thomas Tran, Tuyet Thi King, Sean Nicholas Huang, Zhenzhong Chen, Joseph C |
author_sort | Mostafavi, Mina |
collection | PubMed |
description | BACKGROUND: Genetic models have been developed in divergent branches of the class Alphaproteobacteria to help answer a wide spectrum of questions regarding bacterial physiology. For example, Sinorhizobium meliloti serves as a useful representative for investigating rhizobia-plant symbiosis and nitrogen fixation, Caulobacter crescentus for studying cell cycle regulation and organelle biogenesis, and Zymomonas mobilis for assessing the potentials of metabolic engineering and biofuel production. A tightly regulated promoter that enables titratable expression of a cloned gene in these different models is highly desirable, as it can facilitate observation of phenotypes that would otherwise be obfuscated by leaky expression. RESULTS: We compared the functionality of four promoter regions in S. meliloti (P(araA), P(tauA), P(rhaR), and P(melA)) by constructing strains carrying fusions to the uidA reporter in their genomes and measuring beta-glucuronidase activities when they were induced by arabinose, taurine, rhamnose, or melibiose. P(tauA) was chosen for further study because it, and, to a lesser extent, P(melA), exhibited characteristics suitable for efficient modulation of gene expression. The levels of expression from P(tauA) depended on the concentrations of taurine, in both complex and defined media, in S. meliloti as well as C. crescentus and Z. mobilis. Moreover, our analysis indicated that TauR, TauC, and TauY are each necessary for taurine catabolism and substantiated their designated roles as a transcriptional activator, the permease component of an ABC transporter, and a major subunit of the taurine dehydrogenase, respectively. Finally, we demonstrated that P(tauA) can be used to deplete essential cellular factors in S. meliloti, such as the PleC histidine kinase and TatB, a component of the twin-arginine transport machinery. CONCLUSIONS: The P(tauA) promoter of S. meliloti can control gene expression with a relatively inexpensive and permeable inducer, taurine, in diverse alpha-proteobacteria. Regulated expression of the same gene in different hosts can be achieved by placing both tauR and P(tauA) on appropriate vectors, thus facilitating inspection of conservation of gene function across species. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12866-014-0295-2) contains supplementary material, which is available to authorized users. |
format | Online Article Text |
id | pubmed-4254191 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2014 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-42541912014-12-04 Analysis of a taurine-dependent promoter in Sinorhizobium meliloti that offers tight modulation of gene expression Mostafavi, Mina Lewis, Jainee Christa Saini, Tanisha Bustamante, Julian Albert Gao, Ivan Thomas Tran, Tuyet Thi King, Sean Nicholas Huang, Zhenzhong Chen, Joseph C BMC Microbiol Research Article BACKGROUND: Genetic models have been developed in divergent branches of the class Alphaproteobacteria to help answer a wide spectrum of questions regarding bacterial physiology. For example, Sinorhizobium meliloti serves as a useful representative for investigating rhizobia-plant symbiosis and nitrogen fixation, Caulobacter crescentus for studying cell cycle regulation and organelle biogenesis, and Zymomonas mobilis for assessing the potentials of metabolic engineering and biofuel production. A tightly regulated promoter that enables titratable expression of a cloned gene in these different models is highly desirable, as it can facilitate observation of phenotypes that would otherwise be obfuscated by leaky expression. RESULTS: We compared the functionality of four promoter regions in S. meliloti (P(araA), P(tauA), P(rhaR), and P(melA)) by constructing strains carrying fusions to the uidA reporter in their genomes and measuring beta-glucuronidase activities when they were induced by arabinose, taurine, rhamnose, or melibiose. P(tauA) was chosen for further study because it, and, to a lesser extent, P(melA), exhibited characteristics suitable for efficient modulation of gene expression. The levels of expression from P(tauA) depended on the concentrations of taurine, in both complex and defined media, in S. meliloti as well as C. crescentus and Z. mobilis. Moreover, our analysis indicated that TauR, TauC, and TauY are each necessary for taurine catabolism and substantiated their designated roles as a transcriptional activator, the permease component of an ABC transporter, and a major subunit of the taurine dehydrogenase, respectively. Finally, we demonstrated that P(tauA) can be used to deplete essential cellular factors in S. meliloti, such as the PleC histidine kinase and TatB, a component of the twin-arginine transport machinery. CONCLUSIONS: The P(tauA) promoter of S. meliloti can control gene expression with a relatively inexpensive and permeable inducer, taurine, in diverse alpha-proteobacteria. Regulated expression of the same gene in different hosts can be achieved by placing both tauR and P(tauA) on appropriate vectors, thus facilitating inspection of conservation of gene function across species. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12866-014-0295-2) contains supplementary material, which is available to authorized users. BioMed Central 2014-11-25 /pmc/articles/PMC4254191/ /pubmed/25420869 http://dx.doi.org/10.1186/s12866-014-0295-2 Text en © Mostafavi et al.; licensee BioMed Central Ltd. 2014 This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. |
spellingShingle | Research Article Mostafavi, Mina Lewis, Jainee Christa Saini, Tanisha Bustamante, Julian Albert Gao, Ivan Thomas Tran, Tuyet Thi King, Sean Nicholas Huang, Zhenzhong Chen, Joseph C Analysis of a taurine-dependent promoter in Sinorhizobium meliloti that offers tight modulation of gene expression |
title | Analysis of a taurine-dependent promoter in Sinorhizobium meliloti that offers tight modulation of gene expression |
title_full | Analysis of a taurine-dependent promoter in Sinorhizobium meliloti that offers tight modulation of gene expression |
title_fullStr | Analysis of a taurine-dependent promoter in Sinorhizobium meliloti that offers tight modulation of gene expression |
title_full_unstemmed | Analysis of a taurine-dependent promoter in Sinorhizobium meliloti that offers tight modulation of gene expression |
title_short | Analysis of a taurine-dependent promoter in Sinorhizobium meliloti that offers tight modulation of gene expression |
title_sort | analysis of a taurine-dependent promoter in sinorhizobium meliloti that offers tight modulation of gene expression |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4254191/ https://www.ncbi.nlm.nih.gov/pubmed/25420869 http://dx.doi.org/10.1186/s12866-014-0295-2 |
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