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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The ability of Salmonella enterica serovar Typhimurium to cause disease depends upon the co-ordinated expression of many genes located around the Salmonella chromosome. Specific pathogenicity loci, termed Salmonella pathogenicity islands, have been shown to be crucial for the invasion and survival of Salmonella within host cells. Salmonella pathogenicity island 1 (SPI-1) harbours the genes required for the stimulation of Salmonella uptake across the intestinal epithelia of the infected host. Regulation of SPI-1 genes is complex, as invasion gene expression responds to a number of different signals, presumably signals similar to those found within the environment of the intestinal tract. As a result of our continued studies of SPI-1 gene regulation, we have discovered that the nucleoid-binding protein Fis plays a pivotal role in the expression of HilA and InvF, two activators of SPI-1 genes. A S. typhimurium fis mutant demonstrates a two- to threefold reduction in hilA:Tn5lacZY and a 10-fold reduction in invF:Tn5lacZY expression, as well as a 50-fold decreased ability to invade HEp-2 tissue culture cells. This decreased expression of hilA and invF resulted in an altered secreted invasion protein profile in the fis mutant. Furthermore, the virulence of a S. typhimurium fis mutant is attenuated 100-fold when administered orally, but has wild-type virulence when administered intraperitoneally. Expression of hilA:Tn5lacZY and invF:Tn5lacZY in the fis mutant could be restored by introducing a plasmid containing the S. typhimurium fis gene or a plasmid containing hilD, a gene encoding an AraC-like regulator of Salmonella invasion genes.
Mol Microbiol 2001 Jan
PMID:Fis, a DNA nucleoid-associated protein, is involved in Salmonella typhimurium SPI-1 invasion gene expression. 1112 90

The cupin superfamily is a group of functionally diverse proteins that are found in all three kingdoms of life, Archaea, Eubacteria, and Eukaryota. These proteins have a characteristic signature domain comprising two histidine- containing motifs separated by an intermotif region of variable length. This domain consists of six beta strands within a conserved beta barrel structure. Most cupins, such as microbial phosphomannose isomerases (PMIs), AraC- type transcriptional regulators, and cereal oxalate oxidases (OXOs), contain only a single domain, whereas others, such as seed storage proteins and oxalate decarboxylases (OXDCs), are bi-cupins with two pairs of motifs. Although some cupins have known functions and have been characterized at the biochemical level, the majority are known only from gene cloning or sequencing projects. In this study, phylogenetic analyses were conducted on the conserved domain to investigate the evolution and structure/function relationships of cupins, with an emphasis on single- domain plant germin-like proteins (GLPs). An unrooted phylogeny of cupins from a wide spectrum of evolutionary lineages identified three main clusters, microbial PMIs, OXDCs, and plant GLPs. The sister group to the plant GLPs in the global analysis was then used to root a phylogeny of all available plant GLPs. The resulting phylogeny contained three main clades, classifying the GLPs into distinct subfamilies. It is suggested that these subfamilies correlate with functional categories, one of which contains the bifunctional barley germin that has both OXO and superoxide dismutase (SOD) activity. It is proposed that GLPs function primarily as SODs, enzymes that protect plants from the effects of oxidative stress. Closer inspection of the DNA sequence encoding the intermotif region in plant GLPs showed global conservation of thymine in the second codon position, a character associated with hydrophobic residues. Since many of these proteins are multimeric and enzymatically inactive in their monomeric state, this conservation of hydrophobicity is thought to be associated with the need to maintain the various monomer- monomer interactions. The type of structure-based predictive analysis presented in this paper is an important approach for understanding gene function and evolution in an era when genomes from a wide range of organisms are being sequenced at a rapid rate.
Mol Biol Evol 2001 Apr
PMID:Phylogeny, function, and evolution of the cupins, a structurally conserved, functionally diverse superfamily of proteins. 1126 12

AraC protein, the regulator of the l-arabinose operon in Escherichia coli has been postulated to function by a light switch mechanism. According to this mechanism, it should be possible to find mutations in the DNA-binding domain of AraC that result in weaker arm-DNA-binding domain interactions and which make the protein constitutive, that is, it no longer requires arabinose to activate transcription. We isolated such mutations by randomizing three contiguous leucine residues in the DNA-binding domain, and then by systematically scanning surface residues of the DNA-binding domain with alanine and glutamic acid. As a result, a total of 20 constitutive mutations were found at ten different positions. They form a contiguous trail on the DNA-distal face of the DNA-binding domain, and likely define the region where the N-terminal arm that extends from the N-terminal dimerization domain contacts the C-terminal DNA-binding domain.
J Mol Biol 2001 Apr 06
PMID:Mapping arm-DNA-binding domain interactions in AraC. 1128 51

SoxS is the direct transcriptional activator of at least 15 genes of the Escherichia coli superoxide regulon. SoxS is small (107 amino acids), binds DNA as a monomer and recognizes a highly degenerate DNA binding site, termed 'soxbox'. Like other members of the AraC/XylS family, SoxS has two putative helix-turn-helix (HTH) DNA-binding motifs, and it has been proposed that each HTH motif recognizes a highly conserved recognition element of the soxbox. To determine which nucleotides are important for SoxS binding, we conducted a systematic mutagenesis of the DNA binding sites for SoxS in the zwf and fpr promoters and determined the effect of the soxbox mutations on SoxS DNA binding and transcription activation in vivo by measuring beta-galactosidase activity in strains with fusions to lacZ. We found that the sequences GCAC and CAAA, termed recognition elements 1 and 2 (RE 1 and RE 2), respectively, are critical for SoxS binding, as mutations within these elements severely hinder or eliminate SoxS-dependent transcription activation; substitutions within RE 2 (CAAA), however, are tolerated better than changes within RE 1 (GCAC). Although substitutions at the seven positions separating the two REs had only a modest effect on SoxS binding, AT basepairs were favoured within this 'spacer' region, presumably because, by facilitating DNA bending, they help bring the two recognition elements into proper juxtaposition. We also found that the 'invariant A' present at position 1 of 14/15 functional soxboxes identified thus far is important for SoxS binding, as a change to any other nucleotide at this position reduced SoxS-dependent transcription by approximately 50%. In addition, positions surrounding the REs seem to show a context effect, in that certain substitutions there have little or no effect when the RE has the optimal binding sequence, but produce a pronounced effect when the RE has a suboptimal sequence. We propose that these nucleotides play an important role in effecting differential expression from the various promoters. Lastly, we used gel retardation assays to show that alterations in transcription activation in vivo are caused by effects on DNA binding. Based on this exhaustive mutagenesis, we propose the following optimal sequence for SoxS binding: AnVGCACWWWnKRHCAAAHn (n = A, C, G, T; V = A, C, G; W = A, T; K = G, T; R = A, G; H = A, C, T).
Mol Microbiol 2001 Jun
PMID:Systematic mutagenesis of the DNA binding sites for SoxS in the Escherichia coli zwf and fpr promoters: identifying nucleotides required for DNA binding and transcription activation. 1140 18

During infection, Salmonella enterica serovar Typhimurium colonizes the small intestine of its hosts. This process requires a type III secretion system encoded by several genes on Salmonella pathogenicity island 1 (SPI1), a 40 kb region of DNA near centisome 63 of the Salmonella chromosome. SPI1 gene expression is controlled by a complex regulatory cascade. HilA, a member of the OmpR/ToxR family of transcriptional regulators, directly activates the expression of two SPI1 operons encoding type III apparatus components. hilA transcription is repressed by many environmental conditions and regulatory mutations. This repression requires an upstream repressing sequence (URS) located between -314 and -68 relative to the hilA transcription start site. The repressing activity of the URS is counteracted by two AraC/XylS family members named HilC and HilD. We show that HilC and HilD bind directly to the hilA promoter region in vitro. We also provide evidence that HilC and HilD bind to the same or overlapping sites within the URS. Our data are consistent with a model in which HilC and HilD derepress hilA expression by binding directly to the URS and counteracting its repressing effect in vivo.
Mol Microbiol 2001 Jun
PMID:AraC/XylS family members, HilC and HilD, directly bind and derepress the Salmonella typhimurium hilA promoter. 1144 28

Enteropathogenic Escherichia coli (EPEC) is a major cause of infantile diarrhoea in a number of developing countries and is the prototype of pathogenic bacteria that cause attaching and effacing (A/E) intestinal lesions. A chromosomal pathogenicity island, termed the locus of enterocyte effacement (LEE), contains all the genes necessary for the A/E phenotype as well as genes for a type III secretion system and intimate adhesion. Genes in the LEE and genes involved in the synthesis of bundle-forming pili (BFP) are positively regulated by the plasmid-encoded regulator (Per) and comprise the per regulon. In order to identify factors that control the per regulon, we screened an EPEC genomic library for clones that modulate the expression of per. A plasmid clone that decreased the expression of per was isolated using a lacZ reporter gene fused to the per promoter. Subcloning revealed that YhiX, a putative AraC/XylR family transcriptional regulator, was the effector of per repression. Through downregulation of per, a plasmid overproducing YhiX reduced the synthesis of intimin, BfpA, Tir, and CesT, factors important for EPEC virulence. yhiX is located downstream of gadA, which encodes glutamate decarboxylase, an enzyme involved in acid resistance of E. coli. YhiX was found to be an activator of gadA, and the cloned yhiX gene increased production of glutamate decarboxylases (GAD) and activated the transcription of the gadA and gadB promoters. Therefore, yhiX was renamed gadX. Analysis of a gadX mutant grown in the different culture media with acidic and alkaline pH showed that regulation of perA, gadA and gadB by GadX was altered by the external pH and the culture media condition. Under conditions in which EPEC infects cultured epithelial cells, GadX negatively regulated perA expression, and the derepression in the gadX mutant increased translocation of Tir into epithelial cells relative to wild-type EPEC. DNA mobility shift experiments showed that purified GadX protein bound to the perA, gadA and gadB promoter regions in vitro, indicating that GadX is a transcriptional regulator of these genes. On the basis of these results, we propose that GadX may be involved in the appropriate expression of genes required for acid resistance and virulence of EPEC. Our data are consistent with a model in which environmental changes resulting from passage from the stomach to the proximal small intestine induce the functional effect of GadX on per and GAD expression in order to prevent inappropriate expression of the products of these two systems.
Mol Microbiol 2001 Sep
PMID:An activator of glutamate decarboxylase genes regulates the expression of enteropathogenic Escherichia coli virulence genes through control of the plasmid-encoded regulator, Per. 1155 93

The Hrp type III protein secretion system is essential for pathogenicity of the Gram-negative plant pathogen Xanthomonas campestris pv. vesicatoria. Expression of the hrp gene cluster is controlled by HrpG, a two-component response regulator, and HrpX, an AraC-type transcriptional activator. Using the cDNA-AFLP technique, 30 hrpG-induced (hgi) and five hrpG-repressed (hgr) cDNA fragments were identified, defining a large hrpG-regulon in X. campestris pv. vesicatoria. Expression of most genes in the hrpG-regulon was dependent on hrpX. Seven cDNA fragments map to the known hrp gene cluster and flanking regions. All other genes appear to be scattered over the chromosome and endogenous plasmids. Sequence analysis identified genes encoding putative extracellular proteases, a putative transcriptional regulator and XopJ and XopB (Xanthomonas outer proteins), homologues of YopJ from Yersinia spp. and the avirulence protein AvrPphD of Pseudomonas syringae respectively. XopB is secreted by the Hrp type III secretion system. Analysis of deletion mutants in several hgi genes revealed a new virulence locus. This study demonstrates that cDNA-AFLP is a powerful tool to study prokaryotic transcriptomes and to identify genes contributing to Xanthomonas virulence and putative effector proteins.
Mol Microbiol 2001 Sep
PMID:cDNA-AFLP analysis unravels a genome-wide hrpG-regulon in the plant pathogen Xanthomonas campestris pv. vesicatoria. 1158 Aug 33

Transcription activation of anaerobically induced genes in Escherichia coli is mediated through the action of the global anaerobic regulator FNR. Although regions of FNR involved in FNR-dependent transcription activation have been identified, the side-chains critical to the function of these regions are not known. In this study, alanine-scanning of amino acid residues 80-89 of FNR-activating region 3 (FNR-AR3) was used to determine which amino acid side-chains are required for transcription activation of class II FNR-dependent promoters. In vivo beta-galactosidase assays and in vitro transcription activation assays showed that Ala substitution of Ile81, Gly85 and Asp86 had the largest transcription activation defects, while comparison of the activity of single and double mutants indicated that Thr82, Glu83, Glu87 and Gln88 may contribute in a minor way to FNR-AR3 function. Site-directed mutagenesis of positions 81 and 86 showed that the hydrophobicity of Ile81 and the negative charge of Asp86 were important to FNR-AR3's function. Lastly, substitution of residues of E. coli FNR-AR3 with those more basic residues found in a subset of FNR homologs, such as Rhodobacter sphaeroides FnrL, resulted in a mutant strain that was unable to activate transcription from E. coli class II FNR-dependent promoters. In conclusion, this study demonstrates a requirement for negatively charged and hydrophobic side-chain residues in E. coli FNR-AR3 function, although there is likely to be some variability in the characteristics of this region in other members of the FNR family.
J Mol Biol 2002 Jan 18
PMID:Characterization of activating region 3 from Escherichia coli FNR. 1178 11

A large number of bacteria are able to degrade aromatic carbon sources employing different strategies. All these pathways are objects of regulatory control at the level of gene expression. This includes specific control in response to the availability of the respective substrate and in many cases global control responding to other available carbon sources or to the metabolic status of the cell. Here, the regulatory proteins responsible for gene regulation are reviewed in particular in correlation to other proteins with a similar primary structure. Most common is the appearance of regulators of the LysR family; other abundant regulator types are NtrC/XyIR-type proteins, AraC/XyIS-type proteins and the IcIR-type proteins. Almost all of the regulators exert their effects as activators of gene expression with the exception of the GntR-type proteins, which are exclusively described as repressors. Factors involved in individual cases of global regulatory mechanisms are enterobacterial CAP, (p)ppGpp, Crc protein, and direct modification of a specific regulator. However, for most pathways of aromatic compound degradation, the molecular mechanisms causing global regulation are not understood.
J Mol Microbiol Biotechnol 2002 Mar
PMID:Specific and global regulation of genes associated with the degradation of aromatic compounds in bacteria. 1187 6

The virulence plasmid-encoded type III secretion system of Shigella flexneri consists of the Mxi-Spa secretion apparatus, secreted proteins IpaA-D and IpgD involved in entry of bacteria into epithelial cells,cytoplasmic chaperones IpgC and IpgE and 15 other secreted proteins of unknown function, including VirA and members of the IpaH family. The activity of the Mxi-Spa apparatus is regulated by external signals, and transcription of virA and IpaH genes is specifically induced in conditions of active secretion. We present genetic evidence that regulation of these genes involves both MxiE, the transcriptional activator of the AraC family encoded by the mxi operon, and IpgC, the chaperone for IpaB and IpaC. We also show that together MxiE and IpgC are sufficient to activatevirA and IpaH 9.8 promoters in Escherichia coli. InS. flexneri, increasing the expression of IpgC led to a concomitant increase in IpaH production in conditions of non-secretion. This suggests that the activity of secretion is sensed by the presence of free IpgC, which acts as a coactivator to allow MxiE to activate transcription at its target promoters.
Mol Microbiol 2002 Mar
PMID:Regulation of transcription by the activity of the Shigella flexneri type III secretion apparatus. 1197 Dec 64


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