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Gene regulation by H-NS as a function of growth conditions depends on chromosomal position in Escherichia coli.

Brambilla E, Sclavi B - G3 (Bethesda) (2015)

Bottom Line: Cellular adaptation to changing environmental conditions requires the coordinated regulation of expression of large sets of genes by global regulatory factors such as nucleoid associated proteins.Our results show that the activity of the Phns promoter depends on whether it is placed within the AT-rich regions of the genome that are known to be bound preferentially by H-NS.Genomic position can thus play a significant role in the adaptation of the cells to environmental changes, providing a fitness advantage that can explain the selection of a gene's position during evolution.

View Article: PubMed Central - PubMed

Affiliation: LBPA, UMR 8113 du CNRS, Ecole Normale Supérieure de Cachan, Cachan, France School of Engineering and Science, Jacobs University Bremen, Bremen, Germany.

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Schematic representation of the Phns promoter and of the insertions in the E. coli chromosome. (A) The boxes indicate the binding sites for different proteins in the Phns promoter region (black for FIS, gray for H-NS, white for CspA) as derived from the literature (La Teana et al. 1991; Ueguchi et al. 1993; Falconi et al. 1996). Stars indicate the H-NS high-affinity DNA binding sites (Lang et al. 2007). The -10, -35 regions and the transcription starting site, +1, also are annotated. (B) The promoter-yfp unit is flanked by two T1 terminators from the E. coli rrnB coding sequence. (C) Representation of the six different mirror sites on the E. coli chromosome in which the yfp gene was inserted under the control of the Phns promoter next to the gene conferring resistance to chloramphenicol. The symbols used here are the ones used to indicate these positions in Figure 2B, Figure 5 and Supporting Information, Figure S6. Details about the insertion positions can be found in Table 1. CspA, Cold shock protein A; FIS, factor for inversion stimulation; H-NS, histone-like nucleoid-structuring protein.
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fig1: Schematic representation of the Phns promoter and of the insertions in the E. coli chromosome. (A) The boxes indicate the binding sites for different proteins in the Phns promoter region (black for FIS, gray for H-NS, white for CspA) as derived from the literature (La Teana et al. 1991; Ueguchi et al. 1993; Falconi et al. 1996). Stars indicate the H-NS high-affinity DNA binding sites (Lang et al. 2007). The -10, -35 regions and the transcription starting site, +1, also are annotated. (B) The promoter-yfp unit is flanked by two T1 terminators from the E. coli rrnB coding sequence. (C) Representation of the six different mirror sites on the E. coli chromosome in which the yfp gene was inserted under the control of the Phns promoter next to the gene conferring resistance to chloramphenicol. The symbols used here are the ones used to indicate these positions in Figure 2B, Figure 5 and Supporting Information, Figure S6. Details about the insertion positions can be found in Table 1. CspA, Cold shock protein A; FIS, factor for inversion stimulation; H-NS, histone-like nucleoid-structuring protein.

Mentions: The reporter construction, comprising the Phns promoter upstream of the YFP gene next to an antibiotic resistance cassette, was inserted at six sites along the genome in three sets of mirror sites on each side of the origin of replication (Figure 1 and Table 1). Gene expression from the hns promoter is mainly regulated by FIS and the H-NS protein itself (Ueguchi et al. 1993; Falconi et al. 1993, 1996); therefore, these reporter strains can be used to measure the relative changes in activity of these two nucleoid proteins along the chromosome as a function of growth phase and growth rate. The strains containing the reporter construct in different positions were grown in a 96-well plate overnight to monitor the changes in OD and fluorescence as a function of time in growth media of different composition resulting in different growth rates. To control for the emergence of heterogeneity in the bacterial population, possibly leading to a decreased average amount of measured fluorescence from YFP, the amount of fluorescence per cell was also measured in parallel experiments by flow cytometry, for cells growing in exponential phase, in a flask, in a shaking incubator, confirming the results obtained in the plate reader (Supporting Information, Figure S1).


Gene regulation by H-NS as a function of growth conditions depends on chromosomal position in Escherichia coli.

Brambilla E, Sclavi B - G3 (Bethesda) (2015)

Schematic representation of the Phns promoter and of the insertions in the E. coli chromosome. (A) The boxes indicate the binding sites for different proteins in the Phns promoter region (black for FIS, gray for H-NS, white for CspA) as derived from the literature (La Teana et al. 1991; Ueguchi et al. 1993; Falconi et al. 1996). Stars indicate the H-NS high-affinity DNA binding sites (Lang et al. 2007). The -10, -35 regions and the transcription starting site, +1, also are annotated. (B) The promoter-yfp unit is flanked by two T1 terminators from the E. coli rrnB coding sequence. (C) Representation of the six different mirror sites on the E. coli chromosome in which the yfp gene was inserted under the control of the Phns promoter next to the gene conferring resistance to chloramphenicol. The symbols used here are the ones used to indicate these positions in Figure 2B, Figure 5 and Supporting Information, Figure S6. Details about the insertion positions can be found in Table 1. CspA, Cold shock protein A; FIS, factor for inversion stimulation; H-NS, histone-like nucleoid-structuring protein.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4390576&req=5

fig1: Schematic representation of the Phns promoter and of the insertions in the E. coli chromosome. (A) The boxes indicate the binding sites for different proteins in the Phns promoter region (black for FIS, gray for H-NS, white for CspA) as derived from the literature (La Teana et al. 1991; Ueguchi et al. 1993; Falconi et al. 1996). Stars indicate the H-NS high-affinity DNA binding sites (Lang et al. 2007). The -10, -35 regions and the transcription starting site, +1, also are annotated. (B) The promoter-yfp unit is flanked by two T1 terminators from the E. coli rrnB coding sequence. (C) Representation of the six different mirror sites on the E. coli chromosome in which the yfp gene was inserted under the control of the Phns promoter next to the gene conferring resistance to chloramphenicol. The symbols used here are the ones used to indicate these positions in Figure 2B, Figure 5 and Supporting Information, Figure S6. Details about the insertion positions can be found in Table 1. CspA, Cold shock protein A; FIS, factor for inversion stimulation; H-NS, histone-like nucleoid-structuring protein.
Mentions: The reporter construction, comprising the Phns promoter upstream of the YFP gene next to an antibiotic resistance cassette, was inserted at six sites along the genome in three sets of mirror sites on each side of the origin of replication (Figure 1 and Table 1). Gene expression from the hns promoter is mainly regulated by FIS and the H-NS protein itself (Ueguchi et al. 1993; Falconi et al. 1993, 1996); therefore, these reporter strains can be used to measure the relative changes in activity of these two nucleoid proteins along the chromosome as a function of growth phase and growth rate. The strains containing the reporter construct in different positions were grown in a 96-well plate overnight to monitor the changes in OD and fluorescence as a function of time in growth media of different composition resulting in different growth rates. To control for the emergence of heterogeneity in the bacterial population, possibly leading to a decreased average amount of measured fluorescence from YFP, the amount of fluorescence per cell was also measured in parallel experiments by flow cytometry, for cells growing in exponential phase, in a flask, in a shaking incubator, confirming the results obtained in the plate reader (Supporting Information, Figure S1).

Bottom Line: Cellular adaptation to changing environmental conditions requires the coordinated regulation of expression of large sets of genes by global regulatory factors such as nucleoid associated proteins.Our results show that the activity of the Phns promoter depends on whether it is placed within the AT-rich regions of the genome that are known to be bound preferentially by H-NS.Genomic position can thus play a significant role in the adaptation of the cells to environmental changes, providing a fitness advantage that can explain the selection of a gene's position during evolution.

View Article: PubMed Central - PubMed

Affiliation: LBPA, UMR 8113 du CNRS, Ecole Normale Supérieure de Cachan, Cachan, France School of Engineering and Science, Jacobs University Bremen, Bremen, Germany.

Show MeSH
Related in: MedlinePlus