Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The toxT gene encodes an AraC family transcriptional activator that is responsible for regulating virulence gene expression in Vibrio cholerae. Analysis of ToxT by dominant/negative assays and a LexA-based reporter system demonstrated that the N-terminus of the protein contains dimerization determinants, indicating that ToxT likely functions as a dimer. Additionally, a natural variant of ToxT with only 60% identity in the N-terminus, as well as a mutant form of ToxT with an altered amino acid in the N-terminus (L107F), exhibited altered transcriptional responses to bile, suggesting that the N-terminus is involved in environmental sensing. The C-terminus of ToxT functions to bind DNA and requires dimerization for stable binding in vitro, as demonstrated by gel shift analysis. Interestingly, a dimerized form of the ToxT C-terminus is able to bind DNA in vitro but is transcriptionally inactive in vivo, indicating that the N-terminus contains determinants that are required for transcriptional activation. These results provide a model for a two-domain structure for ToxT, with an N-terminal dimerization and environmental sensing domain and a C-terminal DNA-binding domain; unlike other well-studied AraC family proteins, both domains of ToxT appear to be required for transcriptional activation.
Mol Microbiol 2005 Nov
PMID:Characterization of functional domains of the Vibrio cholerae virulence regulator ToxT. 1626 96

The Gram-negative, curved rod Vibrio cholerae causes the severe diarrhoeal disease cholera. The two major virulence factors produced by V. cholerae during infection are the cholera toxin (CT) and the toxin-coregulated pilus (TCP). Transcription of the genes encoding both CT and the components of the TCP is directly activated by ToxT, a transcription factor in the AraC/XylS family. ToxT binds upstream of the ctxAB genes, encoding CT, and upstream of tcpA, the first gene in a large operon encoding the components of the TCP. The DNA sequences upstream of ctxAB and tcpA that contain ToxT binding sites do not have any significant similarity other than being AT-rich. Extensive site-directed mutagenesis was performed on the region upstream of tcpA previously shown to be protected by ToxT, and we identified specific base pairs important for activation of tcpA transcription by ToxT. This genetic approach was complemented by copper-phenanthroline footprinting experiments that showed protection by ToxT of the base pairs identified as most important for transcription activation in the mutagenesis experiments. Based on this new information and on previous work, we propose the presence of a ToxT-binding motif - the 'toxbox'- in promoters regulated by ToxT. At tcpA, two toxbox elements are present in a direct repeat configuration and both are required for activation of transcription by ToxT. The identity of only a few of the base pairs within the toxbox is important for activation by ToxT, and we term these the core toxbox elements. Lastly, we examined ToxT binding to a mutant having 5 bp inserted between the two toxboxes at tcpA and found that occupancy of both binding sites is retained regardless of the positions of the binding sites relative to each other on the face of the DNA. This suggests that ToxT binds independently as a monomer to each toxbox in the tcpA direct repeat, in accordance with what we observed previously with the inverted repeat ToxT sites between acfA and acfD.
Mol Microbiol 2006 Mar
PMID:The toxbox: specific DNA sequence requirements for activation of Vibrio cholerae virulence genes by ToxT. 1655 83

Enteroaggregative Escherichia coli (EAEC) is defined by aggregative adherence (AA) to HEp-2 cells, where bacteria display adherence to cell surfaces and also to the intervening substratum in a stacked-brick configuration. We previously showed that an AraC homologue designated AggR is required for the expression of plasmid-encoded genes that mediate AA of EAEC strain 042. In this study, we hypothesized that AggR also controls the expression of other virulence determinants in EAEC 042. Using proteomic and microarray analysis, we identified for the first time that AggR activates the expression of chromosomal genes, including 25 contiguous genes (aaiA-Y), which are localized to a 117 kb pathogenicity island (PAI) inserted at pheU. Many of these genes have homologues in other Gram-negative bacteria and were recently proposed to constitute a type VI secretion system (T6SS). AaiC was identified as a secreted protein that has no apparent homologues within GenBank. EAEC strains carrying in-frame deletions of aaiB, aaiG, aaiO or aaiP still synthesized AaiC; however, AaiC secretion was abolished. Cloning of aai genes into E. coli HB101 suggested that aaiA-P are sufficient for AaiC secretion. A second T6SS was identified within the pheU PAI that secretes a protein unrelated by sequence identity to AaiC. Distribution studies indicated that aaiA and aaiC are commonly found in EAEC isolates worldwide, particularly in strains defined as typical EAEC. These data support the hypothesis that AggR is a global regulator of EAEC virulence determinants, and builds on the hypothesis that T6SS is an importance mediator of pathogenesis.
Mol Microbiol 2006 Sep
PMID:Proteomic and microarray characterization of the AggR regulon identifies a pheU pathogenicity island in enteroaggregative Escherichia coli. 1692 58

IkappaBalpha is the major regulator of transcription factor NF-kappaB function. The ankyrin repeat region of IkappaBalpha mediates specific interactions with NF-kappaB dimers, but ankyrin repeats 1, 5 and 6 display a highly dynamic character when not in complex with NF-kappaB. Using chemical denaturation, we show here that IkappaBalpha displays two folding transitions: a non-cooperative conversion under weak perturbation, and a major cooperative folding phase upon stronger insult. Taking advantage of a native Trp residue in ankyrin repeat (AR) 6 and engineered Trp residues in AR2, AR4 and AR5, we show that the cooperative transition involves AR2 and AR3, while the non-cooperative transition involves AR5 and AR6. The major structural transition can be affected by single amino acid substitutions converging to the "consensus" ankyrin repeat sequence, increasing the native state stability significantly. We further characterized the structural and dynamic properties of the native state ensemble of IkappaBalpha and the stabilized mutants by H/(2)H exchange mass spectrometry and NMR. The solution experiments were complemented with molecular dynamics simulations to elucidate the microscopic origins of the stabilizing effect of the consensus substitutions, which can be traced to the fast conformational dynamics of the folded ensemble.
J Mol Biol 2007 Jan 26
PMID:Stabilizing IkappaBalpha by "consensus" design. 1717 35

Corynebacterium glutamicum is a gram-positive soil microorganism able to utilize a large variety of aromatic compounds as the sole carbon source. The corresponding catabolic routes are associated with multiple ring-fission dioxygenases and among other channeling reactions, include the gentisate pathway, the protocatechuate and catechol branches of the beta-ketoadipate pathway and two potential hydroxyquinol pathways. Genes encoding the enzymatic machinery for the bioconversion of aromatic compounds are organized in several clusters in the C. glutamicum genome. Expression of the gene clusters is under specific transcriptional control, apparently including eight DNA-binding proteins belonging to the AraC, IclR, LuxR, PadR, and TetR families of transcriptional regulators. Expression of the gentisate pathway involved in the utilization of 3-hydroxybenzoate and gentisate is positively regulated by an IclR-type activator. The metabolic channeling of ferulate, vanillin and vanillate into the protocatechuate branch of the beta-ketoadipate pathway is controlled by a PadR-like repressor. Regulatory proteins of the IclR and LuxR families participate in transcriptional regulation of the branches of the beta-ketoadipate pathway that are involved in the utilization of benzoate, 4-hydroxybenzoate and protocatechuate. The channeling of phenol into this pathway may be under positive transcriptional control by an AraC-type activator. One of the potential hydroxyquinol pathways of C. glutamicum is apparently repressed by a TetR-type regulator. This global analysis revealed that transcriptional regulation of aromatic compound utilization is mainly controlled by single regulatory proteins sensing the presence of aromatic compounds, thus representing single input motifs within the transcriptional regulatory network of C. glutamicum.
Genet Mol Res 2006 Dec 07
PMID:Transcriptional regulation of catabolic pathways for aromatic compounds in Corynebacterium glutamicum. 1718 85

Virulence factor expression in Vibrio cholerae is controlled by the transcriptional regulatory protein ToxT. ToxT activates transcription of the genes encoding cholera toxin (ctx) and the toxin co-regulated pilus (tcp), as well as accessory colonization factor (acf) genes. Previous studies of ToxT, a member of the AraC family of proteins, have revealed that it consists of two domains, an N-terminal dimerization and environmental sensing domain, and a C-terminal DNA binding domain. In this study, comprehensive scanning alanine mutagenesis was utilized to identify amino acids critical for the function of ToxT. Forty-eight proteins with Ala substitutions (of 267 total) exhibited defects in ToxT-dependent activation (>90% reduction) in both a V. cholerae acfA-phoA reporter strain and a Salmonella typhimurium ctxAp-lacZ reporter strain. Most of these mutant proteins also caused reductions in cholera toxin (CT) and toxin coregulated pilus (TCP) expression in a DeltatoxT V cholerae strain under in vitro virulence factor inducing conditions. Further analysis with a LexA-based reporter system revealed that one of the 20 Ala substitutions in the N terminus (F151A) diminishes dimerization, and this residue is located in a region of predicted alpha-helical structure, thus identifying a putative dimer interface. Ala substitutions in two putative helix-turn-helix (HTH) recognition helices that caused differential promoter activation (K203A and S249A) did not appear to alter specific DNA binding, suggesting these residues contribute to other aspects of transcriptional activation. A number of Ala substitutions were also found that result in a higher level of ToxT transcriptional activity, and these mutations were almost exclusively found within the N terminus, consistent with this domain being involved in modulation of ToxT activity. This study illuminates the contribution of specific amino acids to the dimerization, DNA binding, and transcriptional activity of ToxT.
J Mol Biol 2007 Apr 13
PMID:Identification of residues critical for the function of the Vibrio cholerae virulence regulator ToxT by scanning alanine mutagenesis. 1732 Jan 5

When iron is scarce, Bacillus subtilis expresses genes involved in the synthesis and uptake of the siderophore bacillibactin (BB) and uptake systems to pirate other microbial siderophores. Here, we demonstrate that transcriptional induction of the feuABCybbA operon, encoding the Fe-BB uptake system, is mediated by Btr (formerly YbbB), which is encoded by the immediately upstream gene. Btr contains an AraC-type DNA binding domain fused to a substrate binding protein (SBP) domain related to FeuA, the SBP for Fe-BB uptake. When cells are iron-limited, the Fur-mediated repression of btr is relieved and Btr binds to a conserved direct repeat sequence adjacent to feuA to activate transcription. If BB is present, Btr further activates feuA expression. Btr binds with high affinity to both apo-BB and Fe-BB, and the resulting complex displays a significantly increased efficacy as a transcriptional activator relative to Btr alone. Btr can also activate transcription in response to the structurally similar siderophore enterobactin, although genetic analyses indicate that the two siderophores make distinct interactions with the Btr substrate binding domain. Thus, the FeuABC transporter is optimally expressed under conditions of iron starvation, when Fur-mediated repression is relieved, and in the presence of its cognate substrate.
Mol Microbiol 2007 Oct
PMID:Substrate induction of siderophore transport in Bacillus subtilis mediated by a novel one-component regulator. 1772 65

Streptomyces scabies is the best studied of those streptomycetes that cause an economically important disease known as potato scab. The phytotoxin thaxtomin is made exclusively by these pathogens and is required for virulence. Here we describe regulation of thaxtomin biosynthesis by TxtR, a member of the AraC/XylS family of transcriptional regulators. The txtR gene is imbedded in the thaxtomin biosynthetic pathway and is located on a conserved pathogenicity island in S. scabies, S. turgidiscabies and S. acidiscabies. Thaxtomin biosynthesis was abolished and virulence was almost eliminated in the txtR deletion mutant of S. scabies 87.22. Accumulation of thaxtomin biosynthetic gene (txtA, txtB, txtC, nos) transcripts was reduced compared with the wild-type S. scabies 87.22. NOS-dependent nitric oxide production by S. scabies was also reduced in the mutant. The TxtR protein bound cellobiose, an inducer of thaxtomin production, and transcription of txtR and thaxtomin biosynthetic genes was upregulated in response to cellobiose. TxtR is the first example of an AraC/XylS family protein regulated by cellobiose. Together, these data suggest that cellobiose, the smallest oligomer of cellulose, may signal the availability of expanding plant tissue, which is the site of action of thaxtomin.
Mol Microbiol 2007 Nov
PMID:The AraC/XylS regulator TxtR modulates thaxtomin biosynthesis and virulence in Streptomyces scabies. 1791 90

The XylS protein is the positive transcription regulator of the TOL plasmid meta-cleavage pathway operon Pm. XylS belongs to the AraC family of transcriptional regulators and exhibits an N-terminal domain involved in effector recognition, and a C-terminal domain, made up of seven alpha-helices conforming two helix-turn-helix DNA-binding domains. alpha-Helix 3 and alpha-helix 6 are the recognition helices. In consonance with XylS structural organization, Pm exhibits a bipartite DNA-binding motif consisting of two boxes, called A and B, whose sequences are TGCA and GGNTA, respectively. This bipartite motif is repeated at the Pm promoter so that one of the XylS monomers binds to each of the repeats. An extensive series of genetic epistasis assays combining mutant Pm promoters and XylS single substitution mutant proteins revealed that alpha-helix 3 contacts A box nucleotides, whereas alpha-helix 6 residues contact B box nucleotides. In alpha-helix 3, Asn246 and Arg242 are involved in specific contacts with the TG dinucleotide at box A, whereas Arg296 and Glu299 contact the second G and T nucleotides at box B. On the basis of our results and of the three-dimensional model of the XylS C-terminal domain, we propose that Ser243, Glu249 and Lys250 in alpha-helix 3, and Asn299 and Arg302 in alpha-helix 6 contact the phosphate backbones. Alanine substitutions at the predicted phosphate backbone-contacting residues yielded mutants with low levels of activity, suggesting that XylS-Pm binding specificity not only involves specific amino acid-base interactions, but also relies on secondary DNA structure, which, although at another level, also comes into play. We propose a model in which a XylS dimer binds to the direct repeats in Pm in a head-to-tail conformation that allows the direct interaction of the XylS proximal subunit with the RNA polymerase sigma factor.
J Mol Biol 2008 Jan 04
PMID:XylS-Pm promoter interactions through two helix-turn-helix motifs: identifying XylS residues important for DNA binding and activation. 1800 85

Regulation of virulence gene expression plays a central role in the pathogenesis of enteric bacteria as they encounter diverse environmental conditions in the gastrointestinal tract of their hosts. In this study, we investigated environmental regulation of two putative virulence determinants adcA and kfc by RegA, an AraC/XylS-like regulator, from Citrobacter rodentium, and identified bicarbonate as the environmental signal which induced transcription of adcA and kfc through RegA. Primer extension experiments showed that adcA and kfc were divergently transcribed from sigma(70) promoters. In vivo and in vitro experiments demonstrated that bicarbonate facilitated and stabilized the binding of RegA to an operator located between the two promoters. The interaction of RegA with its DNA target resulted in the formation of a nucleosome-like structure, which evidently displaced the histone-like proteins, H-NS and StpA, from the adcA and kfc promoter regions, leading to transcriptional derepression. In addition, our results indicated that RegA also behaved as a Class I activator by directly stimulating transcription initiation by RNA polymerase. This is the first report to describe the molecular mechanism by which an environmental chemical stimulates transcription of virulence-associated genes of an enteric pathogen through an AraC/XlyS-like activator.
Mol Microbiol 2008 Apr
PMID:Bicarbonate-mediated transcriptional activation of divergent operons by the virulence regulatory protein, RegA, from Citrobacter rodentium. 1828 89


<< Previous 1 2 3 4 5 6 7 8 9 10