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1.
Fig 4

Fig 4. From: BqsR/BqsS Constitute a Two-Component System That Senses Extracellular Fe(II) in Pseudomonas aeruginosa.

Fe(II) titration curve. Duplicate trials showing induction of a transcriptional response for bqsR (gray lines) and bqsS (dashed black lines) in response to increasing Fe(II) concentrations. The minimum Fe(II) concentration to elicit a response was ∼20 μM, and the maximum fold change was seen with a concentration of ∼60 μM. Additional replicates show similar trends.

Naomi N. K. Kreamer, et al. J Bacteriol. 2012 March;194(5):1195-1204.
2.
Fig 2

Fig 2. From: BqsR/BqsS Constitute a Two-Component System That Senses Extracellular Fe(II) in Pseudomonas aeruginosa.

Extracellular Fe(II) upregulates bqsR and bqsS. Wild-type strain PA14 (gray bars) and mutant strain feoB::MAR2xT7 (white bars) were cultured anaerobically in MMM. The feoB mutant strain has a severe defect in Fe(II) uptake. A 30-min Fe(II) shock was performed, and qRT-PCR was performed to analyze expression of bqsR and bqsS. Since the wild type and the mutant show the same levels of transcriptional response, Fe(II) must be extracellular. The bars represent the averages of biological triplicates, and the individual points indicate the fold changes of each replicate.

Naomi N. K. Kreamer, et al. J Bacteriol. 2012 March;194(5):1195-1204.
3.
Fig 3

Fig 3. From: BqsR/BqsS Constitute a Two-Component System That Senses Extracellular Fe(II) in Pseudomonas aeruginosa.

BqsS and BqsR are required for extracellular Fe(II) regulation. (A) WT (gray bars) and ΔbqsS mutant (white bars) transcriptional fold change after Fe(II) shock, as measured by qRT-PCR. (B) WT (gray bars), ΔbqsR mutant (white bars), ΔbqsR mutant with pMQ64-bqsR (bqsR complement) (diagonal bars), and ΔbqsR mutant with pMQ64 (vector-only control) (hashed bars) transcriptional fold change after Fe(II) shock, as measured by qRT-PCR. Both bqsR and bqsS are required for the Fe(II)-induced upregulation of the bqs operon and upregulation of the genes in the bqs regulon. ΔbqsR::pMQ64-bqsR shows WT levels of upregulation. The bars are the averages of biological triplicates, and the individual points indicate the fold changes of each replicate. *, values obtained within background error.

Naomi N. K. Kreamer, et al. J Bacteriol. 2012 March;194(5):1195-1204.
4.
Fig 6

Fig 6. From: BqsR/BqsS Constitute a Two-Component System That Senses Extracellular Fe(II) in Pseudomonas aeruginosa.

bqsR is transcribed over the course of growth as Fe(II) accumulates. Correlation between growth, phenazine production, iron reduction, and bqsR transcription in PA14. Growth curves (OD500) averaged for triplicate cultures of PA14 are plotted alongside the percentages of total Fe that were Fe(II). Bars indicate the average upregulation of bqsS (relative to recA) compared to that in cells grown without Fe and normalized to OD500. For the complete qPCR data set, see Table S2 in the supplemental material. Filled circles, PA14 OD500; open circles, percentage of total Fe that was Fe(II).

Naomi N. K. Kreamer, et al. J Bacteriol. 2012 March;194(5):1195-1204.
5.
Fig 5

Fig 5. From: BqsR/BqsS Constitute a Two-Component System That Senses Extracellular Fe(II) in Pseudomonas aeruginosa.

bqsS responds specifically to Fe(II). Expression of bqsS under both anaerobic (gray bars) and aerobic (white bars) conditions after shock with Fe(II) and other cations. Metal concentrations of 100 μM FeCl3, 100 μM Fe(NH4)2(SO4)2, 10 mM MgCl2, 5 mM CaCl2, 100 μM ZnCl2, 100 μM CuCl2, and 100 μM MnCl2 were used in the 30-min metal shocks for Fe(III), Fe(II), Mg(II), Ca(II), Zn(II), Cu(II), and Mn(II), respectively. The bars are the averages of biological triplicates, and the individual points indicate the fold changes of each replicate.

Naomi N. K. Kreamer, et al. J Bacteriol. 2012 March;194(5):1195-1204.
6.
Fig 1

Fig 1. From: BqsR/BqsS Constitute a Two-Component System That Senses Extracellular Fe(II) in Pseudomonas aeruginosa.

The bqs operon. (A) Genomic organization of the bqs operon, which contains genes encoding two putative PepSY domain-containing membrane proteins (bqsP [PA14_29710] and bqsQ [PA14_29720]), a response regulator (bqsR [PA14_29730]), a histidine kinase (bqsS [PA14_29740]), and a hypothetical protein (bqsT [PA14_29750]). The length of each arrow represents the size of the gene relative to those of the other genes in the operon. (B) A 1% agarose gel confirming that bqs is an operon. The notation bqsP-bqsQ indicates, for example, that in lanes 2 to 4, PCR products were obtained using a forward primer within bqsP and a reverse primer within bqsQ. G, genomic DNA run as a positive control. mRNA −RT (reverse transcriptase) is a negative control to ensure there is no genomic DNA contamination in the mRNA. The presence of products in all of the cDNA lanes (mRNA +RT) reveals this is polycistronic mRNA that contains bqsP, bqsQ, bqsR, bqsS, and bqsT.

Naomi N. K. Kreamer, et al. J Bacteriol. 2012 March;194(5):1195-1204.
7.
Fig 7

Fig 7. From: BqsR/BqsS Constitute a Two-Component System That Senses Extracellular Fe(II) in Pseudomonas aeruginosa.

Iron acquisition model. Fe(III). Outside of the cell, Fe(III) binds to the siderophore pyoverdine, and this complex enters the periplasm via the TonB-dependent outer membrane transporter FpvA (59). The reduction of Fe(III) to Fe(II) frees the iron from pyoverdine. A pyoverdine efflux pump, OmpQ, recycles free pyoverdine by transporting it out of the cell (69). The Fe(II) then enters the cytoplasm through an unidentified ABC transporter. Upon entering the cytoplasm, Fe(II) binds the ferric uptake regulator (Fur), which subsequently represses multiple iron acquisition and iron storage genes (17). Fe(II). Fe(II) enters the periplasm through a nonspecific porin in the outer membrane. While in the periplasm, Fe(II) activates the histidine kinase BqsS, which subsequently activates the response regulator BqsR. BqsR then goes on to induce its regulon. Fe(II) enters the cytoplasm through the ferrous iron transporter FeoB (31). Once in the cytoplasm, Fe(II) binds Fur, which results in the repression of iron acquisition genes. Solid arrows indicate known pathways, dashed arrows indicate putative pathways, and three parallel lines indicates binding. Pvd, pyoverdine; OM, outer membrane; CM, cytoplasmic membrane.

Naomi N. K. Kreamer, et al. J Bacteriol. 2012 March;194(5):1195-1204.

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