Mutual Regulation of Crl and Fur in Escherichia coli W3110

Lelong, Cécile; Rolland, Magali; Louwagie, Mathilde; Garin, Jérôme; Geiselmann, Johannes

doi:10.1074/mcp.m600192-mcp200

Cited by 10 publications

(51 citation statements)

References 23 publications

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“…6C). Cells were grown to stationary phase in LB medium at 30°C and 37°C and also on LB agar at 30°C, a growth condition used to study the regulation of crl expression by Fur in E. coli W3110 (28). The amount of Crl detected in the fur strain was the same as in the wild-type strain under all three conditions (Fig.…”

Section: Fig 5 the Rdar Morphotypes Of Salmonellamentioning

confidence: 99%

“…The Fur hybrid proteins were stable, as indicated by the fact that the expected dimerization of Fur-T18 and T25-Fur (reference 8 and references therein) gave a positive signal (Table 3). crl transcription in E. coli W3110 is repressed by Fur (by 100-fold) and by Crl (by more than 10-fold) (28). To assess the possible relationship between Crl and Fur in Salmonella ATCC 14028, we examined the effects of Fur and Crl on crl expression.…”

Section: Fig 5 the Rdar Morphotypes Of Salmonellamentioning

confidence: 99%

“…Expression of the gene fusions in the Salmonella strains indicated was determined in cultures grown under different conditions. Cultures were grown overnight in LB medium (LB) at 37°C or 30°C or on LB agar at 30°C, as described previously (28). M9, minimal M9 medium with 20 mM glucose; M9ϩFeCl 2 , M9 with 100 M ferrous chloride; M9ϩDP, M9 with the iron chelator 2,2Ј-dipyridyl (100 M).…”

Section: Fig 8 Expression Of Crl-laczmentioning

confidence: 99%

“…In contrast, Crl does not bind 70 and does not modify the affinity of 70 for the core enzyme (12). Lelong et al (28) reported that in E. coli W3110, Crl can interact with the ferric uptake regulator Fur, a key protein for the control of intracellular iron concentration (see reference 8 and references therein) and that Fur represses crl transcription (28).…”

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Identification of Conserved Amino Acid Residues of the Salmonella σ ^S Chaperone Crl Involved in Crl-σ ^S Interactions

et al. 2010

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Proteins that bind factors typically attenuate the function of the factor by restricting its access to the RNA polymerase (RNAP) core enzyme. An exception to this general rule is the Crl protein that binds the stationary-phase sigma factor S (RpoS) and enhances its affinity for the RNAP core enzyme, thereby increasing expression of S -dependent genes. Analyses of sequenced bacterial genomes revealed that crl is less widespread and less conserved at the sequence level than rpoS. Seventeen residues are conserved in all members of the Crl family. Site-directed mutagenesis of the crl gene from Salmonella enterica serovar Typhimurium and complementation of a ⌬crl mutant of Salmonella indicated that substitution of the conserved residues Y22, F53, W56, and W82 decreased Crl activity. This conclusion was further confirmed by promoter binding and abortive transcription assays. We also used a bacterial two-hybrid system (BACTH) to show that the four substitutions in Crl abolish Crl-S interaction and that residues 1 to 71 in S are dispensable for Crl binding. In Escherichia coli, it has been reported that Crl also interacts with the ferric uptake regulator Fur and that Fur represses crl transcription. However, the Salmonella Crl and Fur proteins did not interact in the BACTH system. In addition, a fur mutation did not have any significant effect on the expression level of Crl in Salmonella. These results suggest that the relationship between Crl and Fur is different in Salmonella and E. coli.In bacteria, transcription depends on a multisubunit RNA polymerase (RNAP) consisting of a catalytically active core enzyme (E) with a subunit structure ␣ 2 ␤␤Ј that associates with any one of several factors to form different holoenzyme (E) species. The subunit is required for specific promoter binding, and different factors direct RNAP to different classes of promoters, thereby modulating the gene expression patterns (17). The RNA polymerase holoenzyme containing the 70 subunit is responsible for the transcription of most genes during exponential growth (17). When cells enter stationary phase or are under specific stress conditions (high osmolarity, low pH, or high and low temperatures) during exponential growth, S , which is encoded by the rpoS gene, becomes more abundant, associates with the core enzyme, and directs the transcription of genes essential for the general stress response and for stationary phase survival (17,20,25).Sigma factors compete for binding to a limited amount of the core polymerase (16,17,20,34).70 is abundant throughout the growth cycle and has the highest affinity of all sigma factors for E in vitro (20). In contrast, levels of S reach only about one-third of the 70 levels upon entry into stationary phase, and S exhibits the lowest affinity for E of all sigma factors in vitro (20). The cell uses at least two strategies to ensure the switch between 70 -and S -associated RNA polymerases and to allow gene expression to be reprogrammed upon entry into stationary phase. Several factors (Rsd, 6S RNA, ppGpp, and D...

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Section: Fig 5 the Rdar Morphotypes Of Salmonellamentioning

confidence: 99%

Section: Fig 5 the Rdar Morphotypes Of Salmonellamentioning

confidence: 99%

Section: Fig 8 Expression Of Crl-laczmentioning

confidence: 99%

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Identification of Conserved Amino Acid Residues of the Salmonella σ ^S Chaperone Crl Involved in Crl-σ ^S Interactions

et al. 2010

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“…Another important checkpoint is the formation of E S , which in Escherichia coli is restricted by the competition of S with six other factors for binding to a limited amount of E (9 -11). Even in stationary phase, when S is most abundant, the concentration of the primary factor, 70 , remains 3-fold higher (12). Furthermore, of all of the E. coli factors, S exhibits the lowest affinity for E (13).…”

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Binding of the Unorthodox Transcription Activator, Crl, to the Components of the Transcription Machinery

England

Westblade

Karimova

et al. 2008

Journal of Biological Chemistry

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The small regulatory protein Crl binds to S , the RNA polymerase stationary phase factor. Crl facilitates the formation of the S -associated holoenzyme (E S ) and thereby activates Sdependent genes. Using a real time surface plasmon resonance biosensor, we characterized in greater detail the specificity and mode of action of Crl. Crl specifically forms a 1:1 complex with S , which results in an increase of the association rate of S to core RNA polymerase without any effect on the dissociation rate of E S . Crl is also able to associate with preformed E S with a higher affinity than with S alone. Furthermore, even at saturating S concentrations, Crl significantly increases E S association with the katN promoter and the productive isomerization of the E S -katN complex, supporting a direct role of Crl in transcription initiation. Finally, we show that Crl does not bind to 70 itself but is able at high concentrations to form a weak and transient 1:1 complex with both core RNA polymerase and the 70 -associated holoenzyme, leaving open the possibility that Crl might also exert a side regulatory role in the transcriptional activity of additional non-S holoenzymes.In Enterobacteria, S , encoded by rpoS, is the master regulator of the general stress response and is also responsible for the transcription of stationary phase-specific genes. S accumulates at the onset of the stationary phase and in response to harsh environmental conditions, including carbon and phosphate starvation and acidic and osmotic stress (1, 2). When associated with the RNA polymerase core enzyme (E), 3 the S -associated holoenzyme (E S ) transcribes rpoS-dependent genes and endows the cells with the ability to endure stationary phase and tolerate a multitude of stress conditions (3-5). The acquisition of this multiple stress resistance status, which is dependent on S , has an energetic cost and decreases bacterial fitness in environments containing nutrients at low concentrations (6). Therefore, S abundance and activity are tightly controlled by the interplay of a complex set of regulators that affect transcription, translation, and the stability of the protein. Indeed, the levels of S are controlled primarily by the ClpXP protease, which, together with the catalytic adaptor protein RssB, targets free S for degradation during exponential growth in the absence of stress (7,8). S concentration is not the sole parameter controlling the expression of rpoS-dependent genes. Another important checkpoint is the formation of E S , which in Escherichia coli is restricted by the competition of S with six other factors for binding to a limited amount of E (9 -11). Even in stationary phase, when S is most abundant, the concentration of the primary factor, 70 , remains 3-fold higher (12). Furthermore, of all of the E. coli factors, S exhibits the lowest affinity for E (13). E. coli has developed different strategies that enable S to capture enough E to transcribe its target genes, and several factors favor the formation of E S during stationary phase or in resp...

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Regulators of oxidative stress response genes in Escherichia coli and their functional conservation in bacteria

Chiang

Schellhorn

2012

Archives of Biochemistry and Biophysics

292

222

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Mutual Regulation of Crl and Fur in Escherichia coli W3110

Cited by 10 publications

References 23 publications

Identification of Conserved Amino Acid Residues of the Salmonella σ ^S Chaperone Crl Involved in Crl-σ ^S Interactions

Identification of Conserved Amino Acid Residues of the Salmonella σ ^S Chaperone Crl Involved in Crl-σ ^S Interactions

Binding of the Unorthodox Transcription Activator, Crl, to the Components of the Transcription Machinery

Regulators of oxidative stress response genes in Escherichia coli and their functional conservation in bacteria

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Mutual Regulation of Crl and Fur in Escherichia coli W3110

Cited by 10 publications

References 23 publications

Identification of Conserved Amino Acid Residues of the Salmonella σ S Chaperone Crl Involved in Crl-σ S Interactions

Identification of Conserved Amino Acid Residues of the Salmonella σ S Chaperone Crl Involved in Crl-σ S Interactions

Binding of the Unorthodox Transcription Activator, Crl, to the Components of the Transcription Machinery

Regulators of oxidative stress response genes in Escherichia coli and their functional conservation in bacteria

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Identification of Conserved Amino Acid Residues of the Salmonella σ ^S Chaperone Crl Involved in Crl-σ ^S Interactions

Identification of Conserved Amino Acid Residues of the Salmonella σ ^S Chaperone Crl Involved in Crl-σ ^S Interactions