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Expression of the translocator protein (TSPO) from Pseudomonas fluorescens Pf0-1 requires the stress regulatory sigma factors AlgU and RpoH.

Leneveu-Jenvrin C, Bouffartigues E, Maillot O, Cornelis P, Feuilloley MG, Connil N, Chevalier S - Front Microbiol (2015)

Bottom Line: In silico analysis of this promoter region failed to detect an AlgU consensus binding site; however, a putative binding site for the heat shock response RpoH sigma factor was detected.Accordingly, the promoter activity of the region containing this sequence is increased in response to high growth temperature and slightly lowered in a P. aeruginosa rpoH mutant strain.Taken together, our data suggest that P. fluorescens tspo gene may belong at least partly to the cell wall stress response.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Microbiology Signals and Microenvironment, University of Rouen Evreux, France.

ABSTRACT
The translocator protein (TSPO), previously designated as peripheral-type benzodiazepine receptor, is an evolutionary conserved protein that is found in many Eukarya, Archae, and Bacteria, in which it plays several important functions including for example membrane biogenesis, signaling, and stress response. A tspo homolog gene has been identified in several members of the Pseudomonas genus, among which the soil bacterium P. fluorescens Pf0-1. In this bacterium, the tspo gene is located in the vicinity of a putative hybrid histidine kinase-encoding gene. Since tspo has been involved in water stress related response in plants, we explored the effects of hyperosmolarity and temperature on P. fluorescens Pf0-1 tspo expression using a strategy based on lux-reporter fusions. We show that the two genes Pfl01_2810 and tspo are co-transcribed forming a transcription unit. The expression of this operon is growth phase-dependent and is increased in response to high concentrations of NaCl, sucrose and to a D-cycloserine treatment, which are conditions leading to activity of the major cell wall stress responsive extracytoplasmic sigma factor AlgU. Interestingly, the promoter region activity is strongly lowered in a P. aeruginosa algU mutant, suggesting that AlgU may be involved at least partly in the molecular mechanism leading to Pfl01_2810-tspo expression. In silico analysis of this promoter region failed to detect an AlgU consensus binding site; however, a putative binding site for the heat shock response RpoH sigma factor was detected. Accordingly, the promoter activity of the region containing this sequence is increased in response to high growth temperature and slightly lowered in a P. aeruginosa rpoH mutant strain. Taken together, our data suggest that P. fluorescens tspo gene may belong at least partly to the cell wall stress response.

No MeSH data available.


Related in: MedlinePlus

The activity of the promoter region containing the putative RpoH binding site is increased in response to enhanced growth temperature. (A) Schematic representation of the four transcriptional fusions. The position of the studied promoter region is indicated relatively to the translational initiation start of each ORF (+1). Putative RpoH binding sequence (black star). Gray bars represent the beginning of the luxCDABE reporter cassette of the promoterless pAB133 vector. (B,C) Growth in microtiter plates at 28°C (B) or 32°C (C), and transcriptional activity of P. fluorescens Pf0-1 containing pHK-TSPO-88, pHK-TSPO-157, and pHK-TSPO-227 plasmids.
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Figure 5: The activity of the promoter region containing the putative RpoH binding site is increased in response to enhanced growth temperature. (A) Schematic representation of the four transcriptional fusions. The position of the studied promoter region is indicated relatively to the translational initiation start of each ORF (+1). Putative RpoH binding sequence (black star). Gray bars represent the beginning of the luxCDABE reporter cassette of the promoterless pAB133 vector. (B,C) Growth in microtiter plates at 28°C (B) or 32°C (C), and transcriptional activity of P. fluorescens Pf0-1 containing pHK-TSPO-88, pHK-TSPO-157, and pHK-TSPO-227 plasmids.

Mentions: To monitor tspo transcription, tspo and Pfl01_2810-tspo promoter regions were fused to the promoter-less luxCDABE cassette in the replicative low-copy-number pAB133 vector (Gmr) (Bazire et al., 2005). The promoter regions of interest are located in Figures 1A and 5A, relatively to the tspo and Pfl01_2810 genes. The primer sequences, which were used for PCR amplifications, are shown in Table 2. The SacI-SpeI-digested PCR products were inserted into pAB133, yielding pTSPO, pHK-TSPO, pHK-TSPO-227, pHK-TSPO-157, and pHK-TSPO-88. The inserts were verified by DNA sequencing (Beckman Coulter Genomics).


Expression of the translocator protein (TSPO) from Pseudomonas fluorescens Pf0-1 requires the stress regulatory sigma factors AlgU and RpoH.

Leneveu-Jenvrin C, Bouffartigues E, Maillot O, Cornelis P, Feuilloley MG, Connil N, Chevalier S - Front Microbiol (2015)

The activity of the promoter region containing the putative RpoH binding site is increased in response to enhanced growth temperature. (A) Schematic representation of the four transcriptional fusions. The position of the studied promoter region is indicated relatively to the translational initiation start of each ORF (+1). Putative RpoH binding sequence (black star). Gray bars represent the beginning of the luxCDABE reporter cassette of the promoterless pAB133 vector. (B,C) Growth in microtiter plates at 28°C (B) or 32°C (C), and transcriptional activity of P. fluorescens Pf0-1 containing pHK-TSPO-88, pHK-TSPO-157, and pHK-TSPO-227 plasmids.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4585239&req=5

Figure 5: The activity of the promoter region containing the putative RpoH binding site is increased in response to enhanced growth temperature. (A) Schematic representation of the four transcriptional fusions. The position of the studied promoter region is indicated relatively to the translational initiation start of each ORF (+1). Putative RpoH binding sequence (black star). Gray bars represent the beginning of the luxCDABE reporter cassette of the promoterless pAB133 vector. (B,C) Growth in microtiter plates at 28°C (B) or 32°C (C), and transcriptional activity of P. fluorescens Pf0-1 containing pHK-TSPO-88, pHK-TSPO-157, and pHK-TSPO-227 plasmids.
Mentions: To monitor tspo transcription, tspo and Pfl01_2810-tspo promoter regions were fused to the promoter-less luxCDABE cassette in the replicative low-copy-number pAB133 vector (Gmr) (Bazire et al., 2005). The promoter regions of interest are located in Figures 1A and 5A, relatively to the tspo and Pfl01_2810 genes. The primer sequences, which were used for PCR amplifications, are shown in Table 2. The SacI-SpeI-digested PCR products were inserted into pAB133, yielding pTSPO, pHK-TSPO, pHK-TSPO-227, pHK-TSPO-157, and pHK-TSPO-88. The inserts were verified by DNA sequencing (Beckman Coulter Genomics).

Bottom Line: In silico analysis of this promoter region failed to detect an AlgU consensus binding site; however, a putative binding site for the heat shock response RpoH sigma factor was detected.Accordingly, the promoter activity of the region containing this sequence is increased in response to high growth temperature and slightly lowered in a P. aeruginosa rpoH mutant strain.Taken together, our data suggest that P. fluorescens tspo gene may belong at least partly to the cell wall stress response.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Microbiology Signals and Microenvironment, University of Rouen Evreux, France.

ABSTRACT
The translocator protein (TSPO), previously designated as peripheral-type benzodiazepine receptor, is an evolutionary conserved protein that is found in many Eukarya, Archae, and Bacteria, in which it plays several important functions including for example membrane biogenesis, signaling, and stress response. A tspo homolog gene has been identified in several members of the Pseudomonas genus, among which the soil bacterium P. fluorescens Pf0-1. In this bacterium, the tspo gene is located in the vicinity of a putative hybrid histidine kinase-encoding gene. Since tspo has been involved in water stress related response in plants, we explored the effects of hyperosmolarity and temperature on P. fluorescens Pf0-1 tspo expression using a strategy based on lux-reporter fusions. We show that the two genes Pfl01_2810 and tspo are co-transcribed forming a transcription unit. The expression of this operon is growth phase-dependent and is increased in response to high concentrations of NaCl, sucrose and to a D-cycloserine treatment, which are conditions leading to activity of the major cell wall stress responsive extracytoplasmic sigma factor AlgU. Interestingly, the promoter region activity is strongly lowered in a P. aeruginosa algU mutant, suggesting that AlgU may be involved at least partly in the molecular mechanism leading to Pfl01_2810-tspo expression. In silico analysis of this promoter region failed to detect an AlgU consensus binding site; however, a putative binding site for the heat shock response RpoH sigma factor was detected. Accordingly, the promoter activity of the region containing this sequence is increased in response to high growth temperature and slightly lowered in a P. aeruginosa rpoH mutant strain. Taken together, our data suggest that P. fluorescens tspo gene may belong at least partly to the cell wall stress response.

No MeSH data available.


Related in: MedlinePlus