Repression of <i>tonB</i> transcription during anaerobic growth requires Fur binding at the promoter and a second factor binding upstream

Young, Glenn M.; Postle, Kathleen

doi:10.1111/j.1365-2958.1994.tb00373.x

Cited by 31 publications

(30 citation statements)

References 59 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Although we did not attempt to quantitate the iron-dependent repression of TonB expression, it is clear that neisserial TonB is induced under iron-stressed conditions. This is consistent with observations of E. coli TonB, which is repressed threefold by growth under high-iron conditions and is additionally regulated by anaerobiosis (48).…”

Section: Discussionsupporting

confidence: 80%

Demonstration and Characterization of a Specific Interaction between Gonococcal Transferrin Binding Protein A and TonB

Kenney¹,

Cornelissen

2002

J Bacteriol

View full text Add to dashboard Cite

Iron scavenging by Neisseria gonorrhoeae is accomplished by the expression of receptors that are specific for host iron-binding proteins, such as transferrin and lactoferrin. Efficient transferrin-iron acquisition is dependent on the combined action of two proteins, designated TbpA and TbpB. TbpA is a TonB-dependent outer membrane receptor, whereas TbpB is lipid modified and serves to increase the efficiency of transferrin-iron uptake. Both proteins, together or separately, can be isolated from the gonococcal outer membrane by using affinity chromatography techniques. In the present study, we identified an additional protein in transferrinaffinity preparations, which had an apparent molecular mass of 45 kDa. The ability to copurify this protein by transferrin affinity was dependent upon the presence of TbpA and not TbpB. The amino-terminal sequence of the 45-kDa protein was identical to the amino terminus of gonococcal TonB, indicating that TbpA stably interacted with TonB, without the addition of chemical cross-linkers. Using immunoprecipitation, we could recover TbpA-TonB complexes without the addition of transferrin, suggesting that ligand binding was not a necessary prerequisite for TonB interaction. In contrast, a characterized TonB box mutant of TbpA did not facilitate interaction between these two proteins such that complexes could be isolated. We generated an in-frame deletion of gonococcal TonB, which removed 35 amino acids, including a Neisseria-specific, glycinerich domain. This mutant protein, like the parental TonB, energized TbpA to enable growth on transferrin. Consistent with the functionality of this deletion derivative, TbpA-TonB complexes could be recovered from this strain. The results of the present study thus begin to define the requirements for a functional interaction between gonococcal TbpA and TonB.Bacterial pathogens infecting humans must propagate in an environment that maintains a very small reservoir of soluble iron (38), and since iron is required for the growth of nearly all microorganisms, efficient iron scavenging systems are essential. Most bacteria acquire this scarce but necessary nutrient by synthesis of low-molecular-weight siderophores in addition to membrane-bound transporters for ferric-siderophore uptake (35). However, the pathogenic Neisseria species are not known to secrete iron-scavenging siderophores (46) but have evolved outer membrane receptors that directly bind to host iron sources, such as transferrin and lactoferrin, and relieve them of their bound iron. The transferrin receptor is composed of two proteins: an integral outer membrane, TonB-dependent receptor (TbpA), and a surface-exposed lipoprotein (TbpB) (4, 15, 17). Together, these proteins specifically bind human transferrin and remove and internalize the iron in an energy-dependent fashion (13, 16). By analogy with the well-defined activities of the TonB-dependent siderophore receptors, we proposed that receptor-dependent iron removal from transferrin is accomplished at the expense of the cytoplasmic memb...

show abstract

Section: Discussionsupporting

confidence: 80%

Demonstration and Characterization of a Specific Interaction between Gonococcal Transferrin Binding Protein A and TonB

Kenney¹,

Cornelissen

2002

J Bacteriol

View full text Add to dashboard Cite

show abstract

“…In the presence of iron, expression was threefold repressible aerobically and at least 10-fold repressible anaerobically. This regulatory pattern was similar to those of both exbB and tonB (2,35). The presence of an upstream Tn10 insertion in exbB reduced phoA activity to the iron-insensitive background levels produced by the repressed endogenous phoA genes of W3110 and KP1036, suggesting the absence of an independent exbD promoter.…”

Section: Fig 2 Effects Of Exb Mutations Onmentioning

confidence: 57%

Characterization of the exbBD operon of Escherichia coli and the role of ExbB and ExbD in TonB function and stability

et al. 1995

View full text Add to dashboard Cite

TonB protein appears to couple the electrochemical potential of the cytoplasmic membrane to active transport across the essentially unenergized outer membrane of gram-negative bacteria. ExbB protein has been identified as an auxiliary protein in this process. In this paper we show that ExbD protein, encoded by an adjacent gene in the exb cluster at 65, was also required for TonB-dependent energy transduction and, like ExbB, was required for the stability of TonB. The phenotypes of exbB exbD ؉ strains were essentially indistinguishable from the phenotypes of exbB ؉ exbD strains. Mutations in either gene resulted in the degradation of TonB protein and in decreased, but not entirely absent, sensitivities to colicins B and Ia and to bacteriophage 80. Evidence that the absence of ExbB or ExbD differentially affected the half-lives of newly synthesized and steady-state TonB was obtained. In the absence of ExbB or ExbD, newly synthesized TonB was degraded with a half-life of 5 to 10 min, while the half-life of TonB under steady-state conditions was significantly longer, approximately 30 min. These results were consistent with the idea that ExbB and ExbD play roles in the assembly of TonB into an energy-transducing complex. While interaction between TonB and ExbD was suggested by the effect of ExbD on TonB stability, interaction of ExbD with TonB was detected by neither in vivo cross-linking assays nor genetic tests for competition. Assays of a chromosomally encoded exbD::phoA fusion showed that exbB and exbD were transcribed as an operon, such that ExbD-PhoA levels in an exbB::Tn10 strain were reduced to 4% of the levels observed in an exbB ؉ strain under iron-limiting conditions. Residual ExbD-PhoA expression in an exbB::Tn10 strain was not iron regulated and may have originated from within the Tn10 element in exbB.The acquisition of Fe(III) siderophores and vitamin B 12 by gram-negative bacteria appears to be energized by the transduction of the cytoplasmic membrane proton motive force to a set of outer membrane proteins (termed receptors) that actively transport those nutrients into the periplasmic space. In this process, the cytoplasmic membrane protein, TonB, serves as the energy transducer (1). An auxiliary cytoplasmic membrane protein, ExbB, is required for the stability of TonB (9, 33) and plays a direct role in energy transduction (20,32).ExbB is encoded by the first of a closely positioned pair of genes, with the second gene encoding ExbD (8). ExbB is a 26-kDa protein that traverses the cytoplasmic membrane three times, with the majority of the protein occupying the cytoplasm (9,16,17). ExbD, a 17.8-kDa cytoplasmic membrane protein, has a single transmembrane region at its amino terminus, with the majority of the protein extending into the periplasmic space (15). On the basis of this topological partitioning, it has been proposed that the two proteins may function in concert as a signal transduction molecule (26).The process of transcriptional regulation of the two proteins is not clear. Since the open readin...

show abstract

“…This increased inhibition of Fc uptake by CN-Cbl in iron-repressed cells could result from the reduced ratio of FhuA relative to BtuB and from the increased competition for the iron-repressed levels of TonB protein. Expression of TonB is repressed two-to threefold by iron under aerobic conditions (26,37). When these cells were grown in the presence of both ferric citrate and CN-Cbl, CN-Cbl transport was repressed to about the level in a haploid cell, and CN-Cbl no longer inhibited Fc uptake.…”

Section: Resultsmentioning

confidence: 99%

“…However, when BtuB was overexpressed, growth with excess iron resulted in a fourfold reduction in CN-Cbl uptake with no significant change in CN-Cbl binding. This repression is likely to reflect the Fur-mediated repression of TonB levels, which has been demonstrated with the use of transcriptional fusions and immunoquantitation of the TonB protein (26,37). It was not possible previously to demonstrate iron repression of TonB function because of the concomitant Fur-mediated repression of the iron transporters.…”

Section: Discussionmentioning

confidence: 99%

Mutual inhibition of cobalamin and siderophore uptake systems suggests their competition for TonB function

Kadner

Heller

1995

J Bacteriol

View full text Add to dashboard Cite

Vitamin B 12 (CN-Cbl) and iron-siderophore complexes are transported into Escherichia coli in two energydependent steps. The first step is mediated by substrate-specific outer membrane transport proteins and the energy-coupling TonB protein complex, and the second step uses separate periplasmic permeases for transport across the cytoplasmic membrane. Genetic and biochemical evidence suggests that the TonB-dependent outer membrane transporters contact TonB directly, and thus they might compete for limiting amounts of functional TonB. The transport of iron-siderophore complexes, such as ferrichrome, causes a partial decrease in the rate of CN-Cbl transport. Although CN-Cbl uptake does not inhibit ferrichrome uptake in wild-type cells, in which the amount of the outer membrane ferrichrome transporter FhuA far exceeds that of the cobalamin transporter BtuB, CN-Cbl does inhibit ferrichrome uptake when BtuB is overexpressed from a multicopy plasmid. This inhibition by CN-Cbl is increased when the expression of FhuA and TonB is repressed by growth with excess iron and is eliminated when BtuB synthesis is repressed by CN-Cbl. The mutual inhibition of CN-Cbl and ferrichrome uptake is overcome by increased expression of TonB. Additional evidence for interaction of the Cbl and iron transport systems is provided by the strong stimulation of the BtuB-and TonB-dependent transport of CN-Cbl into a nonexchangeable, presumably cytoplasmic pool by preincubation of cells with the iron(II) chelator 2,2-dipyridyl. Other metal ion chelators inhibited CN-Cbl uptake across the outer membrane. Although the effects of chelators are multiple and complex, they indicate competition or interaction among TonB-dependent transport systems.Uptake of vitamin B 12 (cyanocobalamin, [CN-Cbl]) and iron-siderophore complexes in Escherichia coli occurs by their sequential energy-dependent transport across both cell membranes. The unusual operation of specific transport systems across the outer membrane is necessary for acquisition of these valuable nutrients, whose passage through the porin channels is restricted by their size and low concentrations in natural environments (reviewed in references 6, 18, 27, and 28). Outer membrane proteins BtuB and FhuA mediate high-affinity binding and transport of cobalamins and ferrichrome (Fc), respectively. The TonB-ExbB-ExbD protein complex couples the action of these transporters to the proton motive force across the cytoplasmic membrane (3, 11). The TonB protein is anchored in the cytoplasmic membrane through a nonpolar segment near its N-terminal end (12,16,32) and extends across the periplasmic space to interact directly with the outer membrane transporters (2, 33, 35).The btuCD and fhuCDB loci encode the ATP-driven periplasmic permeases for unidirectional transport of CN-Cbl and Fc, respectively, across the cytoplasmic membrane (7,8,10,34). In a btuC mutant, CN-Cbl is accumulated in the periplasmic space by BtuB and TonB action and is released in unaltered form during a chase with an unlabeled substrate (31)...

show abstract

Repression of tonB transcription during anaerobic growth requires Fur binding at the promoter and a second factor binding upstream

Cited by 31 publications

References 59 publications

Demonstration and Characterization of a Specific Interaction between Gonococcal Transferrin Binding Protein A and TonB

Demonstration and Characterization of a Specific Interaction between Gonococcal Transferrin Binding Protein A and TonB

Characterization of the exbBD operon of Escherichia coli and the role of ExbB and ExbD in TonB function and stability

Mutual inhibition of cobalamin and siderophore uptake systems suggests their competition for TonB function

Contact Info

Product

Resources

About