Location and sequence of the promoter of the gene for the NADH-dependent nitrite reductase of Escherichia coli and its regulation by oxygen, the Fnr protein and nitrite

Jayaraman, Padma-Sheela; Peakman, Timothy C.; Busby, Stephen J. W.; Quincey, R. V.; Cole, Jeffrey A.

doi:10.1016/0022-2836(87)90404-9

Cited by 109 publications

(61 citation statements)

References 22 publications

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“…To assay expression from pnrf derivatives cloned into the lac expression vector pRW50, different host strains were transformed, and ␤-galactosidase activity was measured as described by Jayaraman et al (12), using the Miller protocol (19). Cells were grown in minimal medium (minimal salts with 0.4% glycerol, 10% Lennox broth, and 40 mM fumarate) (22).…”

Section: Methodsmentioning

confidence: 99%

TheEscherichia coliK-12 NarL and NarP Proteins Insulate thenrfPromoter from the Effects of Integration Host Factor

et al. 2006

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The Escherichia coli K-12 nrf operon promoter can be activated fully by the FNR protein (regulator of fumarate and nitrate reduction) binding to a site centered at position ؊41.5. FNR-dependent transcription is suppressed by integration host factor (IHF) binding at position ؊54, and this suppression is counteracted by binding of the NarL or NarP response regulator at position ؊74.5. The E. coli acs gene is transcribed from a divergent promoter upstream from the nrf operon promoter. Transcription from the major acsP2 promoter is dependent on the cyclic AMP receptor protein and is modulated by IHF and Fis binding at multiple sites. We show that IHF binding to one of these sites, located at position ؊127 with respect to the nrf promoter, has a positive effect on nrf promoter activity. This activation is dependent on the face of the DNA helix, independent of IHF binding at other locations, and found only when NarL/NarP are not bound at position ؊74.5. Binding of NarL/NarP appears to insulate the nrf promoter from the effects of IHF. The acs-nrf regulatory region is conserved in other pathogenic E. coli strains and related enteric bacteria but differs in Salmonella enterica serovar Typhimurium.The Escherichia coli K-12 nrf operon encodes a periplasmic nitrite reductase, which is responsible for the reduction of nitrite to ammonium ions (8). Transcription of this operon is driven from a single promoter (pnrf), and expression is induced in the absence of oxygen by the FNR protein (regulator of fumarate and nitrate reduction), a global transcription activator that controls the expression of many genes required for anaerobic respiration (2). FNR binding to a single site, centered at position Ϫ41.5, is sufficient for maximum induction of pnrf (3, 25). However, FNR-dependent activation is suppressed by the binding of integration host factor (IHF), a nucleoidassociated protein, to a site located at position Ϫ54 (IHF I). This suppression is reversed by the binding of NarL or NarP at position Ϫ74.5. NarL and NarP are homologous response regulators that are controlled by the NarX and NarP sensor kinases in response to the presence of nitrite or nitrate (reviewed in references 9 and 24). NarL/NarP binding at position Ϫ74.5 results in displacement of IHF from the IHF I site and, hence, in nitrite/nitrate-dependent activation of pnrf (6).Directly upstream of the nrf operon is the divergently transcribed acs gene, which encodes acetyl coenzyme A synthetase (16). Expression of acs is principally dependent on the acsP2 promoter, which controls a divergent transcript that initiates at a location 280 bp upstream of the nrf transcription start point (1) (Fig. 1). Transcription from acsP2 is totally dependent on activation by the cyclic AMP receptor protein (CRP), which binds at two sites (CRP I and CRP II) (Fig. 1). CRP-dependent activation of acsP2 is modulated by binding of the nucleoidassociated proteins IHF and Fis to tandem sites, namely, IHF II and III and Fis II and III (4) (Fig. 1). Most of our previous studies of pnrf activity...

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Section: Methodsmentioning

confidence: 99%

TheEscherichia coliK-12 NarL and NarP Proteins Insulate thenrfPromoter from the Effects of Integration Host Factor

et al. 2006

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“…Quantification of transcription from the icsP promoter. Transcription from the icsP promoter was determined by measuring ␤-galactosidase activity (as described previously [20] by using the Miller protocol) in AWY8 or strains carrying pHJW7. Routinely, transcription was analyzed in mid-exponentialphase cultures.…”

Section: Methodsmentioning

confidence: 99%

Regulation of IcsP, the Outer Membrane Protease of the Shigella Actin Tail Assembly Protein IcsA, by Virulence Plasmid Regulators VirF and VirB

et al. 2004

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The Shigella outer membrane protease IcsP removes the actin assembly protein IcsA from the bacterial surface, and consequently modulates Shigella actin-based motility and cell-to-cell spread. Here, we demonstrate that IcsP expression is undetectable in mutants lacking either of two transcriptional activators, VirF and VirB. In wild-type Shigella spp., virB expression is entirely dependent on VirF; therefore, to circumvent this regulatory cascade, we independently expressed VirF or VirB in Shigella strains lacking both activators and measured both IcsP levels and transcription from the icsP promoter. Our results show that VirB significantly enhanced icsP transcription, even in the absence of VirF. In contrast, when VirF was induced in the absence of VirB, VirF had variable effects. The regulation of icsP is distinctly different from the regulation of the gene encoding its major substrate, icsA, which is activated by VirF and not VirB. We propose that the different pathways regulating icsA and icsP may be critical to the modulation of IcsA-mediated actin-based motility by IcsP.Shigella spp., gram-negative bacterial pathogens cause severe and bloody diarrhea in their human hosts by invading and spreading through the colonic epithelium. Shigella movement within the host cell cytoplasm is dependent on the ability of the bacterium to recruit host cell actin to its surface to form an "actin tail," which propels the bacterium from one cell to another (5,16,29). Actin tail assembly is mediated by a single bacterial protein, IcsA, which is found on the outer surface at one pole of the bacterium (17). This asymmetric localization of IcsA ensures that actin assembly occurs in a directional manner. In its mature form, IcsA is comprised of two domains: the ␣ domain (residues 53 to 758) contains the determinant for actin assembly (14) and extends from the bacterial surface into the extracellular environment, whereas the ␤ domain (residues 759 to 1102) is embedded in the outer membrane (33). The amount of IcsA ␣ domain exposed on the bacterial surface correlates with the efficiency of actin tail formation in the cytoplasm of infected cells (21).IcsP, an outer membrane protease of Shigella, cleaves IcsA between Arg 758 and Arg 759 , removing the entire IcsA ␣ domain from the bacterial surface (8, 13, 15a, 31). Overexpression of IcsP leads to complete removal of the IcsA ␣ domain from the bacterial cell surface (32), whereas genetic disruption of icsP increases the total amount of cell associated IcsA ␣ domain, leading to an increase in the rate of actin-based movement of Shigella (31). Although IcsP is not required for polar localization of IcsA (6,28), it contributes to the maintenance of a tight polar cap of IcsA on the bacterial surface (31). Furthermore, as Shigella enter stationary phase, the amount of cell-associated IcsA ␣ domain decreases dramatically, an effect due at least in part to IcsP (18,32).These data demonstrate that IcsP plays an important role in modulating the amount of the IcsA ␣ domain present on the bacter...

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“…faecalis and serve to enhance the transcription of these genes under anaerobic growth conditions. E. coli possesses NADH-dependent NIR, which may account for the low NIR activity even in the untransformed E. coli cells (see Table 1 ) ; the expression of the nirB gene encoding the NIR is regulated not only by oxygen but also by nitrite (Cole, 1968;Jayaraman et al, 1987). The production of E. coli NIR was stimulated by two-to threefold by the addition of nitrite.…”

Section: Discussionmentioning

confidence: 99%

Cloning and characterization of a nitrite reductase gene from Alcaligenes faecalis and its expression in Escherichia coli

Nishiyama

Suzuki

Kukimoto

et al. 1993

Journal of General Microbiology

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The gene (nir) encoding the copper-containing nitrite reductase (NIR) of a denitrifying bacterium, Alcaligenes faecalis S-6, was cloned by a synthetic oligonucleotide-probing method. The nucleotide sequence of the cloned DNA fragment revealed the primary structure of the NIR precursor containing the N-terminal signal sequence for secretion. A nucleotide sequence, possibly recognized by a transcriptional regulator resembling FNR was found upstream of the structural gene. When the cloned gene was expressed in Escherichia coli under the control of the lac promoter at 37 "C, NIR was produced as large inclusion bodies and little activity was detected. When cultivation was at 20 "C, most of the NIR was detected in the soluble fraction and a significant portion of the protein was translocated into the periplasmic space, accompanied by removal of its signal sequence.

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Location and sequence of the promoter of the gene for the NADH-dependent nitrite reductase of Escherichia coli and its regulation by oxygen, the Fnr protein and nitrite

Cited by 109 publications

References 22 publications

TheEscherichia coliK-12 NarL and NarP Proteins Insulate thenrfPromoter from the Effects of Integration Host Factor

TheEscherichia coliK-12 NarL and NarP Proteins Insulate thenrfPromoter from the Effects of Integration Host Factor

Regulation of IcsP, the Outer Membrane Protease of the Shigella Actin Tail Assembly Protein IcsA, by Virulence Plasmid Regulators VirF and VirB

Cloning and characterization of a nitrite reductase gene from Alcaligenes faecalis and its expression in Escherichia coli

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