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Department of Microbiology, Moyne Institute of Preventive Medicine, Trinity College, Dublin 2, Ireland
Correspondence
Charles J. Dorman
cjdorman{at}tcd.ie
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ABSTRACT |
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 TOP ABSTRACT INTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
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Present address: Division of Infectious Diseases, Children's Hospital, Enders Building, Harvard University, 300 Longwood Avenue, Boston, MA 02115, USA.
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INTRODUCTION |
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TOPABSTRACTINTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
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) and the archaea (Bell & Jackson, 2000; Brinkman et al., 2002). In Salmonella and Escherichia coli, Lrp is a DNA-binding protein with a molecular mass of 18.8 kDa whose activity can be modulated by L-leucine. This amino acid influences the oligomeric state of Lrp, with octamer formation being favoured in its presence (Chen et al., 2001, 2005; Chen & Calvo, 2002; De los Rios & Perona, 2007; Peterson et al., 2007). Lrp can activate or repress gene expression, and depending on the system subject to Lrp regulation, leucine can play a stimulatory, inhibitory or neutral role (Brinkman et al., 2003; Calvo & Matthews, 1994). Lrp activity is not restricted to transcriptional control: it can affect other DNA transactions, such as site-specific recombination (Kelly et al., 2006; Roesch & Blomfield, 1998).
Lrp has roles in Salmonella that are distinct from its contributions in E. coli. These include regulating the expression of the spv virulence genes on the Salmonella virulence plasmid (Marshall et al., 1999), which are important for the establishment of a systemic infection in the host (Libby et al., 2000; Rotger & Casadesús, 1999; Paesold et al., 2002). Lrp is also a regulator of the pef fimbrial genes on the same plasmid (Nicholson & Low, 2000). The mechanism by which Lrp regulates pef genes involves competition with the DNA adenine methylase for access to key 5'-GATC-3' sites in the pef regulatory region that determine the expression state of the fimbrial operon (Nicholson & Low, 2000). This mechanism is similar to the one that controls expression of the pap fimbrial genes in uropathogenic strains of E. coli (Braaten et al., 1992; Hernday et al., 2002; van der Woude et al., 1992). In addition, Lrp controls the expression of the virulence plasmid traJ gene (Camacho & Casadesús, 2005), allowing Lrp to influence the conjugal transfer of this episome (Camacho & Casadesús, 2002). On the Salmonella chromosome, the Lrp protein regulates type 1 fimbrial gene expression through a mechanism that is completely distinct from that used to control type 1 fimbrial genes in E. coli (Kelly et al., 2006; McCusker et al., 2008; McFarland et al., 2008; Roesch & Blomfield, 1998). Instead of influencing the efficiency of a site-specific recombination process that moves the E. coli fim genes between ON and OFF expression states, Lrp in Salmonella enterica controls fim gene expression by governing the supply of a regulatory protein, FimZ (McFarland et al., 2008).
Despite its involvement in many important regulatory processes, the S. enterica lrp gene has not been previously characterized. Bioinformatic analysis of the S. enterica LT2 genome sequence has suggested locations for the lrp transcription start signals; however, the inferred binding sites for RNA polymerase differ from the consensus sequences for promoters used by RNA polymerase holoenzyme containing sigma-70 or any other Salmonella sigma factor (McClelland et al., 2001). In this study we cloned the lrp gene from the S. enterica serovar Typhimurium mouse-virulent strain SL1344 and characterized its regulation at the level of transcription using both in vivo and in vitro methods. The transcription control elements were found to be in a different location from that suggested by the available bioinformatic information. The S. enterica lrp gene was found to share several regulatory features with its well-characterized counterpart in E. coli K-12, although there were also some features that were specific to S. enterica.
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METHODS |
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TOPABSTRACTINTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
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Strains, plasmids and oligonucleotides.
The strains and plasmids are described in Table 1 and the oligonucleotides used in this study are listed in Table 2. Plasmid pZEPlrp was produced by PCR amplification of a 612 bp fragment, including 61 bp of the lrp ORF and 533 bp of the lrp regulatory region of SL1344, using primer pair lrp-gfp-F and lrp-gfp-R (Table 1). This fragment and the parental vector pZEP08 were each digested with SmaI and XbaI, purified and ligated. The plasmid pBSK.lrp was used for DNase I footprinting and associated DNA sequencing reactions. It was generated by PCR of SL1344 DNA with primer pair lrp-F.BamHI and lrp-R.EcoRI, followed by cloning into the multi-cloning site of pBluescript II SK (–). Plasmid pKMC101 was used to determine the lrp transcription start site by primer extension analysis. This plasmid was produced by PCR amplification of the lrp gene and its flanking regions using primer pair Fwd1.3-EcoRI and Rev1.3-PstI followed by restriction enzyme digestion and ligation of the resulting DNA fragment into the EcoRI and PstI sites in pCL1921.
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Electrophoretic mobility shift assay (EMSA).
DNA probes were amplified using biotinylated primers to the region of interest (Table 2
) and were subsequently purified by gel electrophoresis. Approximately 100 pg biotinylated DNA probe was incubated with binding buffer for 5 min at room temperature as described by Camacho & Casadesús (2002). Increasing concentrations of Lrp protein were added, followed by incubation for 20 min at room temperature. Protein–DNA complexes were formed in a reaction volume of 20 µl, including leucine where applicable. A 10 µl aliquot of the protein–DNA mixture, plus loading dye, was subjected to electrophoresis at 100 V on Novex 6 % DNA Retardation gels (Invitrogen) at room temperature. Gels were transferred onto membrane using the Novex XCell II blot module (Invitrogen), UV-cross-linked, and developed using the Chemiluminescent Nucleic Acid Detection Module (Pierce).
DNase I footprinting.
DNase I footprinting was carried out as previously described (McFarland et al., 2008). The probes used for DNase I footprinting were generated by PCR using primer pairs lrp-F.BamHI and lrp-R.EcoRI (Table 2) with pBSK.lrp as the DNA template. The sequences complementary to the T7 oligonucleotides are located externally to the multi-cloning site of pBluescript II SK (–). T4 polynucleotide kinase was used to label the DNA probes at both ends with [
-32P]ATP, which were then digested to remove the label from one end, depending on the strand to be analysed. Lrp protein–DNA binding reactions used for DNase I footprinting were identical to those used for EMSA analysis, except that these reactions were performed in 50 µl volumes. DNA sequencing reactions using dideoxy chain-terminators were performed as previously described (McFarland et al., 2008), using the T4 DNA sequencing kit (USB) and appropriate sequencing primers. The products of the DNase I footprinting and the DNA sequencing reactions were resolved by electrophoresis through a 6 % urea-polyacrylamide gel.
Primer extension analysis.
RNA was extracted from SL1344 that had been grown to OD600 3.5 in LB broth (approx. 5.5 h growth), and to OD600 0.5 in MOPS minimal broth, using the SV Total RNA isolation kit (Promega). The oligonucleotide PE-lrp (Table 2) is complementary to 32 bp of the lrp mRNA, 58 nt into the ORF. PE-lrp was labelled with [-32P]ATP using T4 phosphonucleotide kinase (New England Biolabs) according to the manufacturer's instructions. The SuperScript III Reverse Transcriptase kit (Invitrogen) was used to perform the primer extension reaction; the product was then purified by ethanol precipitation, resuspended in formamide loading dye, and heated to 95 °C for 10 min. Samples were analysed by electrophoresis on denaturing 6 % polyacrylamide gels alongside sequencing reactions that were generated using primer PE-lrp and pKMC101 as template.
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RESULTS |
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TOPABSTRACTINTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
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). It was important to assess the impact of the lrp mutation on the physiology of the bacterium. For this reason, the growth of the lrp knockout mutant was compared with the wild-type in the complex medium LB and in MOPS minimal medium (with and without leucine) over a 24 h period. In LB, the growth curves of the wild-type and mutant were essentially superimposable, although the wild-type had a slightly shorter doubling time (28 min) than the lrp mutant (30 min). In MOPS minimal medium, the mutant had a doubling time that was 
30 % longer than that of the wild-type during the exponential phase of growth; the two strains achieved approximately similar culture densities in stationary phase. This pattern was mildly affected by the addition of leucine: there was some reduction in the growth of the wild-type in exponential phase and the lrp mutant had a shortened lag phase. These data implied an important role for Lrp in the physiology of S. Typhimurium growing in minimal medium. They also showed that, in contrast to the situation in E. coli (Bouloc et al., 1992), leucine did not abolish the growth of the bacteria in minimal medium. The reason for this is not known.
Negative autoregulation of lrp transcription
The putative lrp transcription signals were cloned into the plasmid pZEP08 to create an lrp-gfp transcriptional fusion (Methods). This plasmid, pZEPlrp, allowed the green fluorescent protein to be used as a reporter of lrp promoter activity. The plasmid was introduced into the SL1344 wild-type and the SL1344 lrp knockout mutant. This allowed the effect of the lrp mutation on the activity of the lrp promoter to be measured. Strains SL1344 and SL1344 lrp grew with similar growth curves in LB when harbouring the pZEPlrp plasmid (Fig. 1a). In MOPS minimal medium, the lrp mutant grew more slowly than the wild-type and the presence of leucine had a minor negative influence on growth (Fig. 2a).
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) or in minimal MOPS (Fig. 2b). When leucine was added to the MOPS culture, the level of lrp-gfp expression increased by between 1.5- and 2-fold, provided the Lrp protein was present; leucine did not have this effect in the lrp knockout mutant (Fig. 2b). These data showed that Lrp repressed the transcription of the lrp gene and that this negative effect was partially alleviated by the presence of leucine.
Identifying the lrp transcription start site
It was necessary to identify the transcription start site of the S. Typhimurium lrp gene by molecular methods. Previous estimates based on bioinformatic techniques suggested that the lrp promoter had several non-standard features, in particular very poor matches to the –35 and –10 hexanucleotide motifs that are bound by the RpoD sigma factor of RNA polymerase (McClelland et al., 2001). If these were confirmed, they would have to be taken into account in assembling a model of lrp gene regulation.
The transcription start site was determined using a standard primer extension procedure (Methods) and mapped to a cluster of A bases located 257 bp upstream from the translation start codon of the lrp ORF (Fig. 3). The start site was 10 bases further downstream of the position described previously for the lrp gene in E. coli K-12 (Wang et al., 1994). Moreover, the likely binding sites for RNA polymerase were poor matches to the standard features associated with sigma-70 promoters, in keeping with the weak nature of the promoter. The –35 hexamer matched the consensus sequence in three out of six positions and the –10 had only two matches out of six to the consensus (Fig. 3). These data showed that the S. Typhimurium lrp transcription start site determined previously using in silico methods was likely to be incorrect.
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), in agreement with the data obtained using the lrp-gfp reporter fusion (Figs 2b and 3b).
Purified Lrp binds to the lrp promoter region
Purified Lrp protein (McFarland et al., 2008) was used in a series of EMSAs to assess Lrp interaction with the transcriptional regulatory region of the SL1344 lrp gene (Fig. 4). Experiments were performed initially with a DNA fragment that extended from 115 bp upstream of the translation initiation codon to 418 bp further upstream, spanning from –275 to +143 with respect to the transcription start site (Fig. 4a). The labelled DNA fragment was incubated with Lrp protein at a range of concentrations, and parallel incubations were performed in the presence of leucine or glycine. Two Lrp–DNA complexes were formed in all three cases (Fig. 4b). Leucine encouraged the formation of the slowest-migrating Lrp–DNA complex at the expense of the faster-migrating one; glycine did not have this effect. These results showed that Lrp could bind to the lrp regulatory region in the absence of amino acids and that leucine could modulate this process.
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). The six DNA fragments, F102, F200, F311, R111, R202 and R310, were used in EMSAs with purified Lrp protein. Only F311 and R310 formed complexes with the protein (Fig. 4c). These two fragments shared a common 203 bp stretch of DNA that included the lrp promoter. This region of DNA was amplified and used in a further EMSA with Lrp. Three Lrp–DNA complexes were detected, with a fourth being visible at the highest concentration of protein (Fig. 4d). The formation of the slowest-migrating Lrp–DNA complexes was unaffected by the presence of leucine or glycine.
Taken together, the data from the EMSAs showed that Lrp bound to DNA sequences located between positions –167 and +36, numbered with respect to the transcription start site (+1) of the S. Typhimurium lrp gene. DNase I footprinting was used to examine Lrp–DNA interaction in this region in more detail. The 418 bp DNA fragment (Fig. 4a, b) was labelled at one end with 32P, incubated with increasing concentrations of Lrp protein, and then treated with DNase I. The samples were then separated by electrophoresis on a DNA sequencing gel (Fig. 5). Increasing concentrations of Lrp protein were accompanied by a pattern of protection from, and periodic hypersensitivity to, digestion by DNase I on both the coding (Fig. 5a) and the non-coding DNA strands (Fig. 5b) over a region that extended from –135 to +15. The region of protection included the lrp promoter, a finding that was consistent with its repression by the Lrp protein. There was an abrupt transition between DNase I sensitivity and protection and vice versa between 50 and 100 nM Lrp. The pattern of protection and hypersensitivity to DNase I was subtly altered by the presence and absence of 15 mM leucine. Addition of the amino acid was accompanied by hypersensitivity to DNase I in the promoter region (–40 to +1) on the coding and non-coding strands, a region that was protected by Lrp in the absence of leucine (Fig. 5a, b). Leucine also reversed the sensitivity of bases to DNase I in the regions –90 to –100 and –120 to –130 (Fig. 5a). Several matches to the degenerate consensus sequence for Lrp binding sites (Wang et al., 1994) were identified in the region between –110 and +15; these are summarized in Fig. 6. In several cases, these sequences were flanked by ‘T’ residues that had shown hypersensitivity to DNase I digestion in the presence of Lrp.
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DISCUSSION |
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TOPABSTRACTINTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
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).
The lrp promoter in E. coli has been described as leucine-sensitive (Borst et al., 1996) and leucine-insensitive (Lin et al., 1992; Wang et al., 1994). In our study, the lrp promoter in S. Typhimurium consistently displayed between a 1.5- and 2-fold higher activity in the presence of leucine in bacteria growing in minimal MOPS medium (Fig. 1). This modest positive effect of leucine on lrp transcription was contingent on the presence of the Lrp protein; the lrp knockout mutant expressed the lrp-gfp fusion to the same level in the presence or the absence of leucine (Fig. 1). We noted a dip in lrp transcription at the end of exponential growth (Fig. 1b). While we cannot explain this in mechanistic terms, a similar effect was noted previously in the case of the E. coli lrp gene (Landgraf et al., 1996).
EMSAs showed that Lrp binds to a 418 bp DNA fragment and that this binding is not abrogated by the addition of leucine (Fig. 4). It has been shown previously that leucine does not alter the binding of Lrp to the regulatory region of the E. coli lrp gene (Wang et al., 1994). However, in the case of the S. Typhimurium gene, leucine (but not glycine) alters the relative abundances of the different Lrp–DNA complexes in favour of the slowest-migrating complex. This suggests that leucine changes the nature of the Lrp–DNA complex without displacing the protein. The details of this alteration to the nucleoprotein complex are unknown but are likely to be subtle given the very modest effect of leucine on lrp promoter activity.
The 418 bp fragment encompasses a 203 bp subfragment that binds Lrp, forming up to four Lrp–DNA complexes in EMSAs (Fig. 4). The number and relative abundances of these complexes do not respond to leucine or glycine. This shows that the subtle leucine-sensitivity of the Lrp–DNA complexes formed with the 418 bp fragment involves DNA sequences not found in this smaller piece of DNA, perhaps due to a requirement for long-range protein–DNA interactions that can only occur on the longest DNA fragments used.
DNase I footprinting reveals Lrp-mediated protection and distortion of the DNA structure between approximately positions –130 and +15 (Fig. 5). Protection is seen most clearly in the absence of leucine on the coding (Fig. 5a, lanes 1 to 5) and non-coding strands (Fig. 5b, lanes 1 to 5). The zones of protection are interrupted by bases showing strong Lrp-mediated hypersensitivity to DNase I. Adding leucine has a subtle effect on the patterns of protection and hypersensitivity. It reduces the overall degree of protection afforded by Lrp and it modifies the pattern of DNase I hypersensitivity. In particular, the protection seen between bases –40 and +1 on the coding strand is replaced by a pattern of periodic hypersensitivity (Fig. 5a). This pattern is also detectable on the non-coding strand (Fig. 5b). New regions of hypersensitivity are also seen further upstream on the coding strand, between bases –95 and –130 (Fig. 5a).
The markedly periodic pattern of hypersensitivity to DNase I digestion is consistent with wrapping of the DNA around the Lrp protein, resulting in enhanced exposure of certain bases to DNase I (Nickerson & Achberger, 1995; Wagner, 2000), and has been reported previously for other DNA sequences that bind Lrp (McFarland et al., 2008; Wang & Calvo, 1993; Wiese et al., 1997). The 3D structure of the E. coli Lrp protein has been solved (de los Rios & Perona, 2007). This protein, which is almost identical to Lrp in S. Typhimurium, can form octamers capable of wrapping approximately 120 bp of DNA, a stretch that is similar in length to that shown here to be affected by Lrp. In the Lrp octamer, the eight helix–turn–helix (H-T-H) DNA-binding motifs located in the amino-terminal domains of each monomer are arranged around the circumference of the octameric structure. This allows the protein to dock with Lrp binding sites distributed along the DNA with a spacing of approximately three helical turns, or 30 bp. The fit of each H-T-H motif in the DNA requires adjustment to the Lrp protein structure, suggesting that an induced-fit mechanism operates. It is possible that the binding of leucine to the Regulation of Amino acid Metabolism (RAM) domain in the carboxyl terminus of each Lrp monomer may provide the structural adjustment required to strengthen or to weaken Lrp–DNA interaction, depending on the gene regulatory sequence involved (de los Rios & Perona, 2007). The modifications to the patterns of protection and hypersensitivity seen in the lrp regulatory region in the presence and absence of leucine are consistent with an adjustment to the Lrp–DNA contacts due to the influence of the amino acid on Lrp protein structure.
Data from the present study show that Lrp can form up to four complexes with the S. Typhimurium lrp promoter region, with one being located downstream of the transcription start site (Fig. 4). DNase I protection data suggest that Lrp binding occurs as far downstream as base +15, and a leucine-sensitive and DNase-I-hypersensitive base is located at position +35. This pattern of protein–DNA interaction is consistent with repression of the lrp promoter by Lrp promoter occlusion. The effect of adding leucine is not to abolish Lrp–DNA interaction, but to remodel it so that the negative influence of the Lrp protein on lrp promoter function is attenuated.
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ACKNOWLEDGEMENTS |
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Edited by: L. S. Frost
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REFERENCES |
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Borst, D. W., Blumenthal, R. M. & Matthews, R. G. (1996). Use of an in vivo titration method to study a global regulator: effect of varying Lrp levels on expression of gltBDF in Escherichia coli. J Bacteriol 178, 6904–6912.
Bouloc, P., Vinella, D. & D'Ari, R. (1992). Leucine and serine induce mecillinam resistance in Escherichia coli. Mol Gen Genet 235, 242–246.[CrossRef][Medline]
Braaten, B. A., Platko, J. V., van der Woude, M. W., Simons, B. H., de Graaf, F. K., Calvo, J. M. & Low, D. A. (1992). Leucine-responsive regulatory protein controls the expression of both pap and fan pili operons in Escherichia coli. Proc Natl Acad Sci U S A 89, 4250–4254.
Brinkman, A. B., Bell, S. D., Lebbink, R. J., de Vos, W. M. & van der Oost, J. (2002). The Sulfolobus solfataricus Lrp-like protein LysM regulates lysine biosynthesis in response to lysine availability. J Biol Chem 277, 29537–29549.
Brinkman, A. B., Ettema, T. J. G., de Vos, W. M. & van der Oost, J. (2003). The Lrp family of transcriptional regulators. Mol Microbiol 48, 287–294.[CrossRef][Medline]
Calvo, J. M. & Matthews, R. G. (1994). The leucine-responsive regulatory protein, a global regulator of metabolism in Escherichia coli. Microbiol Rev 58, 466–490.
Camacho, E. M. & Casadesús, J. (2002). Conjugal transfer of the virulence plasmid of Salmonella enterica is regulated by the leucine-responsive regulatory protein and DNA adenine methylation. Mol Microbiol 44, 1589–1598.[CrossRef][Medline]
Camacho, E. M. & Casadesús, J. (2005). Regulation of traJ transcription in the Salmonella virulence plasmid by strand-specific DNA adenine hemimethylation. Mol Microbiol 57, 1700–1718.[CrossRef][Medline]
Chen, S. & Calvo, J. M. (2002). Leucine-induced dissociation of Escherichia coli Lrp hexadecamers to octamers. J Mol Biol 318, 1031–1042.[CrossRef][Medline]
Chen, S., Rosner, M. H. & Calvo, J. M. (2001). Leucine-regulated self-association of leucine-responsive regulatory protein (Lrp) from Escherichia coli. J Mol Biol 312, 625–635.[CrossRef][Medline]
Chen, S., Iannolo, M. & Calvo, J. M. (2005). Cooperative binding of the leucine-responsive regulatory protein (Lrp) to DNA. J Mol Biol 345, 251–264.[CrossRef][Medline]
de los Rios, S. & Perona, J. J. (2007). Structure of the Escherichia coli leucine-responsive regulatory protein (Lrp) reveals a novel octameric assembly. J Mol Biol 366, 1589–1602.[CrossRef][Medline]
Hautefort, I., Proenca, M. J. & Hinton, J. C. D. (2003). Single-copy green fluorescent protein gene fusions allow accurate measurement of Salmonella gene expression in vitro and during infection of mammalian cells. Appl Environ Microbiol 69, 7480–7491.
Hernday, A., Krabbe, M., Braaten, B. & Low, D. (2002). Self-perpetuating epigenetic pili switches in bacteria. Proc Natl Acad Sci U S A 99, 16470–16476.
Hoiseth, S. K. & Stocker, B. A. (1981). Aromatic-dependent Salmonella typhimurium are non-virulent and effective as live vaccines. Nature 291, 238–239.[CrossRef][Medline]
Kelly, A., Conway, C., Cróinín, T. O., Smith, S. G. J. & Dorman, C. J. (2006). DNA supercoiling and the Lrp protein determine the directionality of fim switch DNA inversion in Escherichia coli K-12. J Bacteriol 188, 5356–5363.
Landgraf, J. R., Wu, J. & Calvo, J. (1996). Effects of nutrition and growth rate on Lrp levels in Escherichia coli. J Bacteriol 178, 6930–6936.
Lerner, C. G. & Inouye, M. (1990). Low copy number plasmids for regulated low-level expression of cloned genes in Escherichia coli with blue/white insert screening capability. Nucleic Acids Res 18, 4631
Libby, S. J., Lesnick, M., Hasegawa, P., Weidenhammer, E. & Guiney, D. G. (2000). The Salmonella virulence plasmid spv genes are required for cytopathology in human monocyte-derived macrophages. Cell Microbiol 2, 49–58.[CrossRef][Medline]
Lin, R., D'Ari, R. & Newman, E. B. (1992). 
placMu insertions in genes of the leucine regulon: extension of the regulon to genes not regulated by leucine. J Bacteriol 174, 1948–1955.
Marshall, D. G., Sheehan, B. J. & Dorman, C. J. (1999). A role for the leucine-responsive regulatory protein and integration host factor in the regulation of the Salmonella plasmid virulence (spv) locus in Salmonella typhimurium. Mol Microbiol 34, 134–145.[CrossRef][Medline]
McClelland, M., Sanderson, K. E., Spieth, J., Clifton, S. W., Latreille, P., Courtney, L., Porwollik, S., Ali, J., Dante, M. & other authors (2001). Complete genome sequence of Salmonella enterica serovar Typhimurium LT2. Nature 413, 852–856.[CrossRef][Medline]
McCusker, M. P., Turner, E. C. & Dorman, C. J. (2008). DNA sequence heterogeneity in Fim tyrosine-integrase recombinase-binding elements and functional motif asymmetries determine the directionality of the fim genetic switch in Escherichia coli K-12. Mol Microbiol 67, 171–187.[Medline]
McFarland, K. A., Lucchini, S., Hinton, J. C. D. & Dorman, C. J. (2008). The leucine-responsive regulatory protein, Lrp, activates transcription of the fim operon in Salmonella enterica serovar Typhimurium via the fimZ regulatory gene. J Bacteriol 190, 602–612.
Nicholson, B. & Low, D. (2000). DNA methylation-dependent regulation of pef expression in Salmonella typhimurium. Mol Microbiol 35, 728–742.[CrossRef][Medline]
Nickerson, C. A. & Achberger, E. C. (1995). Role of curved DNA in binding of Escherichia coli RNA polymerase to promoters. J Bacteriol 177, 5756–5761.
Paesold, G., Guiney, D. G., Eckmann, L. & Kagnoff, M. F. (2002). Genes in the Salmonella pathogenicity island 2 and the Salmonella virulence plasmid are essential for Salmonella-induced apoptosis in intestinal epithelial cells. Cell Microbiol 4, 771–781.[CrossRef][Medline]
Peterson, S. N., Dahlquist, F. W. & Reich, N. O. (2007). The role of high affinity non-specific DNA binding by Lrp in transcriptional regulation and DNA organization. J Mol Biol 369, 1307–1317.[CrossRef][Medline]
Roesch, P. L. & Blomfield, I. C. (1998). Leucine alters the interaction of the leucine-responsive regulatory protein (Lrp) with the fim switch to stimulate site-specific recombination in Escherichia coli. Mol Microbiol 27, 751–761.[CrossRef][Medline]
Rotger, R. & Casadesús, J. (1999). The virulence plasmids of Salmonella. Int Microbiol 2, 177–184.[Medline]
Tuan, L. R., D'Ari, R. & Newman, E. B. (1990). The leucine regulon of Escherichia coli K-12: a mutation in rblA alters expression of L-leucine-dependent metabolic operons. J Bacteriol 172, 4529–4535.
van der Woude, M. W., Braaten, B. A. & Low, D. A. (1992). Evidence for global regulatory control of pilus expression in Escherichia coli by Lrp and DNA methylation: model building based on analysis of pap. Mol Microbiol 6, 2429–2435.[Medline]
Wagner, R. (2000). Transcription Regulation in Prokaryotes. Oxford, UK: Oxford University Press.
Wang, Q. & Calvo, J. M. (1993). Lrp, a major regulatory protein in Escherichia coli, bends DNA and can organize the assembly of a higher-order nucleoprotein structure. EMBO J 12, 2495–2501.[Medline]
Wang, Q., Wu, J., Friedberg, D., Platko, J. & Calvo, J. M. (1994). Regulation of the Escherichia coli lrp gene. J Bacteriol 176, 1831–1839.
Wiese, D. E., Jr, Ernsting, B. R., Blumenthal, R. M. & Matthews, R. G. (1997). A nucleoprotein activation complex between the leucine-responsive regulatory protein and DNA upstream of the gltBDF operon in Escherichia coli. J Mol Biol 270, 152–168.[CrossRef][Medline]
Received 10 March 2008;
revised 17 April 2008;
accepted 23 April 2008.
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) and SL1344 lrp(pZEPlrp) (
) in LB. (b) Gfp expression from lrp-gfp in the wild-type (
) or without (
) or without (
