Repression and induction of the nag regulon of Escherichia coll K‐12: the roles of nagC and nagA in maintenance of the uninduced state

Plumbridge, Jacqueline

doi:10.1111/j.1365-2958.1991.tb00828.x

Cited by 119 publications

(170 citation statements)

References 42 publications

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“…By analogous procedures, the non-polar cat (Cm r ) cassette (cf. Plumbridge, 1991) from pCM7 was inserted into the same HindIII of kpsF as that used for mutagenesis with the Km r cassette to yield pSX390, followed by construction of the suicide vector pSX405 and allelic exchange with the wild-type copy in EV36 to generate strain EV508. Southern hybridization analysis of ScaI-digested DNA isolated from EV36, EV500 and EV508 confirmed that the mutants contained the expected kpsF disruptions (results not shown).…”

Section: Insertion Mutagenesis Of Kpsfmentioning

confidence: 99%

Reduced polysialic acid capsule expression in Escherichia coli K1 mutants with chromosomal defects in kpsF

Cieslewicz

Vimr

1997

Molecular Microbiology

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SummaryNeuroinvasive Escherichia coli K1 synthesizes and assembles a polysialic acid capsule virulence factor on the external leaflet of the outer membrane. This capsule functions in pathogenesis by blocking nonimmune host defence mechanisms and acting as a relatively non-immunogenic molecular mimic of the polysialic acid chains found in high concentrations on neural cell adhesion molecules of the human embryo and neonate. The synthetic, regulatory and export components for capsule expression are encoded in three functionally distinct gene blocks or regions of the 20 kb kps 'pathogenicity island'. These regions are organized as two convergently transcribed operons inserted into the monocistronic tRNA gene, pheV. The six genes of the so-called region 1 operon are transcribed in the same direction as pheV, and at least four of these genes are required for polysialic acid export. Expression of this operon is thermoregulated by transcriptional control of its first gene, kpsF. To investigate the function of region 1 further, two independent chromosomal disruptions were engineered by inserting promoterless, terminatorless kanamycin or chloramphenicol resistance cassettes into the HindIII site of the kpsF coding sequence. The chromosomal insertions were regulated by temperature in the same way as the wild-type operon, demonstrating that this control mechanism remained intact in these mutants. Chemical, immunological and ultrastructural microscopical methods demonstrated that full-length polysialic acid chains were synthesized but not exported by the kpsF mutants. This phenotype was correlated with decreased plaque diameter when the mutants were infected with the capsule-specific bacteriophage K1F. The export defect could not be complemented in trans with kpsF þ containing its cis-regulatory region because of titration of an apparent positive regulator of region 1 expression, whereas complementation was observed with a plasmid expressing kpsF from a physiologically irrelevant promoter. An N-terminal polyhistidine peptide was attached to KpsF and used to purify the overproduced polypeptide. Antibodies raised against KpsF identified at least one of its paralogues in E. coli, GutQ, suggesting that KpsF and its homologues are membrane associated. The results indicate the requirement for a precise balance between region 1 components of the capsule export machinery, and that KpsF plays a positive role in the assembly, operation or regulation of this apparatus.

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Section: Insertion Mutagenesis Of Kpsfmentioning

confidence: 99%

Reduced polysialic acid capsule expression in Escherichia coli K1 mutants with chromosomal defects in kpsF

Cieslewicz

Vimr

1997

Molecular Microbiology

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“…Strain NM522 was used for cloning experiments. Other E. coli strains used are 11814 (MG1655 ⌬lac) (A. Vianney), IBPC536 (thrA1 leuB6 hisG4 argE3 ⌬(gpt-lac)62 thi-1 galK2 ara-14 xyl-5 mtl-1 tsx-33 kdgK51 rpsL31 supE44 recB21 recC22 sbcB15 sbcC201 nagC::tc) (19), TK821 (MC4100 ompR::Tn10) (8), MV679 (MG1655 ⌬lacZYA ⌬nanR) (I. Blomfield), PHL1242 (MG1655 ⌬cpxR) (G. Jubelin), pop4129 (zhd::Tn10 ⌬crp18) (J.-C. Lazzaroni), and IBPC1016 (JM101 mlc::tc nanR6) (20).…”

Section: Methodsmentioning

confidence: 99%

Function and Expression of an N -Acetylneuraminic Acid-Inducible Outer Membrane Channel in Escherichia coli

et al. 2005

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The Escherichia coli yjhA (renamed nanC) gene encodes a protein of the KdgM family of outer membranespecific channels. It is transcribed divergently from fimB, a gene involved in the site-specific inversion of the region controlling transcription of the fimbrial structural genes but is separated from it by one of the largest intergenic regions in E. coli. We show that nanC expression is induced by N-acetylneuraminic acid and modulated by N-acetylglucosamine. This regulation occurs via the NanR and NagC regulators, which also control fimB expression. nanC expression is also activated by the regulators cyclic AMP-catabolite activator protein, OmpR, and CpxR. When the NanC protein was reconstituted into liposomes, it formed channels with a conductance of 450 pS at positive potential and 300 to 400 pS at negative potential in 800 mM KCl. The channels had a weak anionic selectivity. In an ompR background, where the general porins OmpF and OmpC are absent, NanC is required for growth of E. coli on N-acetylneuraminic acid as the sole carbon source. All these results suggest that NanC is an N-acetylneuraminic acid outer membrane channel protein.Porins are proteins that form water-filled channels across the outer membrane of gram-negative bacteria. These channels allow the diffusion of hydrophilic molecules (Ͻ600 Da for the general porins OmpF and OmpC) into the periplasm with no particular substrate specificity. In Escherichia coli this family of proteins is represented by the trimeric porins OmpC, OmpF, and PhoE. Elucidation of the structure of these proteins showed that each monomeric barrel is formed by 16 ␤-strands (5). Other outer membrane channels catalyze the specific diffusion of a particular class of nutrient and thus allow the uptake of compounds that would diffuse too slowly through the general porins (15).The E. coli maltose-specific channel protein LamB and the sucrose channel protein ScrY are the best studied of this class of channel proteins. These proteins also form trimers, and each monomer consists of 18 transmembrane ␤-strands (24). A few other specific channels are known in E. coli: the aryl-␤-

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“…However, the molecular details of curli gene repression by GlcNAc-6P remain unknown. NagC is thought to be a functional regulator only in the absence of GlcNAc since NagC-GlcNAc-6P binds DNA poorly (40). In the absence of nagA, GlcNAc-6P accumulates in the cell (55) and binds to NagC, preventing NagC-mediated repression at the nag promoters (40).…”

Section: Vol 188 2006mentioning

confidence: 99%

“…NagC acts as a transcriptional repressor of the nag genes when GlcNAc is not available (36,38,40). A role for NagD in GlcNAc metabolism and transport has not been identified.…”

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confidence: 99%

GlcNAc-6P Levels Modulate the Expression of Curli Fibers byEscherichia coli

2006

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Curli are extracellular surface fibers that are produced by many members of the Enterobacteriaceae and contribute to biofilm formation. The environmental cues that promote biofilm formation are poorly understood. We found that deletion of the N-acetylglucosamine-6-phosphate (GlcNAc-6P) deacetylase gene, nagA, resulted in decreased transcription from the curli-specific promoters csgBA and csgDEFG and a corresponding decrease in curli production in Escherichia coli. nagA is in an operon that contains nagB, nagC, nagD, and nagE, whose products are required for utilization of GlcNAc as a carbon source. NagC is a repressor of the nagBACD and nagE genes in the absence of intracellular GlcNAc-6P. We found that nagC mutants were also defective in curli production. Growth of a wild-type strain on media containing additional GlcNAc reduced curli gene transcription to a level similar to the level observed when nagA was deleted. The defect in curli production in nagA or nagC mutants was alleviated by deletion of the GlcNAc transporter gene, nagE. Curli-producing ⌬nagA suppressor mutants whose cells were unable to take up GlcNAc were isolated. These results suggest that elevated levels of intracellular GlcNAc-6P signal cells to down-regulate curli gene expression.

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Repression and induction of the nag regulon of Escherichia coll K‐12: the roles of nagC and nagA in maintenance of the uninduced state

Cited by 119 publications

References 42 publications

Reduced polysialic acid capsule expression in Escherichia coli K1 mutants with chromosomal defects in kpsF

Reduced polysialic acid capsule expression in Escherichia coli K1 mutants with chromosomal defects in kpsF

Function and Expression of an N -Acetylneuraminic Acid-Inducible Outer Membrane Channel in Escherichia coli

GlcNAc-6P Levels Modulate the Expression of Curli Fibers byEscherichia coli

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