cheW Genes Research Articles

Comparative Genomics Reveals Pathogenicity-Related Loci in Shewanella algae.

Shewanella algae is an emerging marine zoonotic pathogen and accounts for considerable mortality and morbidity in compromised hosts. However, there is scarce literature related to the understanding of the genetic background of virulence determinants in S. algae. In this study, we aim to determine the occurrence of common virulence genes in S. algae using whole-genome sequence and comparative genomic analysis. Comparative genomics reveals putative-virulence genes related to bile resistance, chemotaxis, hemolysis, and motility. We detected the existence of hlyA, hlyD, and hlyIII involved in hemolysis. We also found chemotaxis gene cluster cheYZA operon and cheW gene. The results provide insights into the genetic basis underlying pathogenicity in S. algae.

Two Agrobacterium tumefaciens CheW Proteins Are Incorporated into One Chemosensory Pathway with Different Efficiencies.

Agrobacterium tumefaciens is the agent that causes crown gall tumor disease on more than 140 species of dicotyledonous plants. Chemotaxis of A. tumefaciens toward the wound sites of the host plant is the first step to recognize the host. CheW is a coupling protein that bridges the histidine kinase CheA and the chemoreceptors to form the chemotaxis core signaling complex and plays a crucial role in the assembly and function of the large chemosensory array. Unlike all previously reported chemotaxis systems, A. tumefaciens has only one major che operon but two cheW homologs (atu2075 as cheW1 and atu2617 as cheW2) on unlinked loci. The in-frame deletion of either cheW gene significantly affects A. tumefaciens chemotaxis but does not abolish the chemotaxis, unless both cheW genes were deleted. The effect of cheW2 deletion on the chemotaxis is more severe than that of cheW1 deletion. Either CheW can interact with CheA and couple it to the cell poles. The promoter activity of cheW2 is always higher than that of cheW1 under all of the tested conditions. When two cheW genes were adjusted to the same expression level by using the identical promoter, the difference between the effects of two CheW proteins on the chemotaxis still existed. Therefore, we envision that both the different molecular ratio of two CheW proteins in cell and the different affinities of two CheW proteins with CheA and chemoreceptors result in the efficiency difference of two CheW proteins in functioning in the large chemosensory array.

A mutation in the Xanthomonas oryzae pv. oryzae wxoD gene affects xanthan production and chemotaxis

Xanthomonas oryzae pv. oryzae causes bacterial blight in rice (Oryza sativa L.). The effect of a mutation in the wxoD gene, that encodes a putative O-antigen acetylase, on xanthan production as well as bacterial chemotaxis was investigated. The mutation increased xanthan production by 52 %. The mutant strain was non-motile on semi-solid agar swarm plates. In addition, several genes involved in chemotaxis, including the cheW, cheV, cheR, and cheD genes, were down-regulated by a mutation in the wxoD gene. Thus, the mutation in the wxoD gene affects xanthan production as well as bacterial chemotaxis. However, the wxoD gene is not essential for the virulence of X. oryzae.

Two CheW coupling proteins are essential in a chemosensory pathway of Borrelia burgdorferi

In the model organism Escherichia coli, the coupling protein CheW, which bridges the chemoreceptors and histidine kinase CheA, is essential for chemotaxis. Unlike the situation in E. coli, Borrelia burgdorferi, the causative agent of Lyme disease, has three cheW homologues (cheW(1) , cheW(2) and cheW(3) ). Here, a comprehensive approach is utilized to investigate the roles of the three cheWs in chemotaxis of B. burgdorferi. First, genetic studies indicated that both the cheW(1) and cheW(3) genes are essential for chemotaxis, as the mutants had altered swimming behaviours and were non-chemotactic. Second, immunofluorescence and cryo-electron tomography studies suggested that both CheW(1) and CheW(3) are involved in the assembly of chemoreceptor arrays at the cell poles. In contrast to cheW(1) and cheW(3) , cheW(2) is dispensable for chemotaxis and assembly of the chemoreceptor arrays. Finally, immunoprecipitation studies demonstrated that the three CheWs interact with different CheAs: CheW(1) and CheW(3) interact with CheA(2) whereas CheW(2) binds to CheA(1) . Collectively, our results indicate that CheW(1) and CheW(3) are incorporated into one chemosensory pathway that is essential for B. burgdorferi chemotaxis. Although many bacteria have more than one homologue of CheW, to our knowledge, this report provides the first experimental evidence that two CheW proteins coexist in one chemosensory pathway and that both are essential for chemotaxis.

Polar localization of CheA2 in Rhodobacter sphaeroides requires specific Che homologs.

Rhodobacter sphaeroides is a motile bacterium that has multiple chemotaxis genes organized predominantly in three major operons (cheOp(1), cheOp(2), and cheOp(3)). The chemoreceptor proteins are clustered at two distinct locations, the cell poles and in one or more cytoplasmic clusters. One intriguing possibility is that the physically distinct chemoreceptor clusters are each composed of a defined subset of specific chemotaxis proteins, including the chemoreceptors themselves plus specific CheW and CheA proteins. Here we report the subcellular localization of one such protein, CheA(2), under aerobic and photoheterotrophic growth conditions. CheA(2) is predominantly clustered and localized at the cell poles under both growth conditions. Furthermore, its localization is dependent upon one or more genes in cheOp(2) but not those of cheOp(1) or cheOp(3). In E. coli, the polar localization of CheA depends upon CheW. The R. sphaeroides cheOp(2) contains two cheW genes. Interestingly, CheW(2) is required under both aerobic and photoheterotrophic conditions, whereas CheW(3) is not required under aerobic conditions but appears to play a modest role under photoheterotrophic conditions. This suggests that R. sphaeroides contains at least two distinct chemotaxis complexes, possibly composed of proteins dedicated for each subcellular location. Furthermore, the composition of these spatially distinct complexes may change under different growth conditions.

Pathogenesis of enteric Campylobacter infection.

Campylobacter jejuni and related species are important human pathogens, causing acute human enterocolitis, and they are the most common cause of food-borne diarrhoea in many industrialized countries. Previous infection with certain strains of C. jejuni is also linked with the development of the neurological disorder Guillain-Barre syndrome (GBS). Despite its importance as a human pathogen, relatively little is understood of the mechanisms of C. jejuni-associated disease. The recent release of the complete genome sequence of C. jejuni strain NCTC 11168, together with new strategies for random and directed mutagenesis, have allowed a better insight into some of the genetic determinants of C. jejuni virulence. In this review current knowledge on the pathogenesis of C. jejuni infection is summarized, and possible directions of future research indicated.

Identification of a chemotaxis operon with two cheY genes in Rhodobacter sphaeroides.

A large chemotaxis operon was identified in Rhodobacter sphaeroides WS8-N using a probe based on the 3' terminal portion of the Rhizobium meliloti cheA gene. Two genes homologous to the enteric cheY were identified in an operon also containing cheA, cheW, and cheR homologues. The deduced protein sequences of che gene products were aligned with those from Escherichia coli and shown to be highly conserved. A mutant with an interrupted copy of cheA showed normal patterns of swimming, unlike the equivalent mutants in E. coli which are smooth swimming. Tethered cheA mutant cells showed normal responses to changes in organic acids, but increased, inverted responses to sugars. The unusual behaviour of the cheA mutant and the identification of two homologues of cheY suggests that R. sphaeroides has at least two pathways controlling motor activity. To identify functional similarity between the newly identified R. sphaeroides Che pathway and the methyl-accepting chemotaxis protein (MCP)-dependent pathway in enteric bacteria, the R. sphaeroides cheW gene was expressed in a cheW mutant strain of E. coli and found to complement, causing a partial return to a swarming phenotype. In addition, expression of the R. sphaeroides gene in wild-type E. coli resulted in the same increased tumbling and reduced swarming as seen when the native gene is overexpressed in E. coli. The identification of che homologues in R. sphaeroides and complementation by cheW suggests the presence of MCPs in an organism previously considered to use only MCP-independent sensing. The MCP-dependent pathway, appears conserved.(ABSTRACT TRUNCATED AT 250 WORDS)

Role of the CheW protein in bacterial chemotaxis: overexpression is equivalent to absence.

The cheW gene from Escherichia coli has been cloned an inducible promoter, and the effects of the overproduction of the CheW protein on chemotactic behavior and receptor covalent modification have been examined. Plasmids that contain the cheW gene behind a regulatable promoter complement a cheW mutation when the CheW protein is produced at low levels. However, when the CheW protein is greatly overproduced in either a wild-type strain or a cheW mutant, chemotaxis is greatly inhibited, cheW null mutant cells swim smoothly as if they were constantly responding to an attractant. Surprisingly, cells in which the CheW protein is overproduced also swim smoothly. The behavioral defect produced by overproduction of the CheW protein does not require the presence of the cheR, cheB, or cheZ gene. Receptor demethylation is also inhibited by overproduction of the CheW protein, as it is by a mutation in the cheW gene or a response to an attractant. In all respects, therefore, overproduction of the CheW protein has the same consequences as does a mutation in the cheW gene or a response to an attractant. A model involving two states of the CheW protein is proposed to explain its role in bacterial chemotaxis.

Control of transducer methylation levels in Escherichia coli: investigation of components essential for modulation of methylation and demethylation reactions.

During bacterial chemotaxis in Escherichia coli, adaptation is accomplished by reversible methylation of the transmembrane signal transducers. Methyl groups are added by the CheR protein in a slow response to attractants and removed by the CheB protein in response to repellents. The methylesterase activity of the CheB protein is modulated by a factor that is controlled in a global fashion throughout the cell. By controlling the level of expression of the cheR, cheB, and transducer genes with exogenous promoters on multicopy plasmids, we demonstrate that the modulating factor exists in stoichiometric concentrations relative to CheB protein and that the generation or efficacy of this factor requires the cheA and/or cheW gene products, suggesting that phosphorylation of the methylesterase by CheA may be involved in its global activation. We show that in the absence of any modulation of the CheB activity, the CheR methyltransferase activity is modulated in a local fashion at the transducers, most likely as a result of a conformational change in the transducer protein brought about by the binding of ligand, and does not require CheA or CheW.

Receptor interactions through phosphorylation and methylation pathways in bacterial chemotaxis.

The effects of messages initiated by one receptor on the covalent modification of a second receptor were studied by use of a technique for rapidly separating the receptors. Methylation of the bacterial-chemotactic serine receptor increases as a result of aspartate binding to the aspartate receptor. The aspartate-induced methylation on the serine receptor is absent in a strain that lacks cheA and cheW genes and is not the result of physical interaction, such as the formation of heterodimers between the aspartate and serine receptors, or of alterations in the affinity of the serine receptor for the methyltransferase and the methylesterase. Serine-induced methylation of the serine receptor did not require cheA and cheW. A model is presented in which the receptor methylation level depends on the combination of (i) a ligand-induced conformational change on the receptor substrate of the methylation enzymes and (ii) an indirect cytoplasmic signal that operates through the methylesterase.

Identification of a possible nucleotide binding site in CheW, a protein required for sensory transduction in bacterial chemotaxis.

CheW is an essential component of the system which mediates chemotaxis in Salmonella typhimurium and Escherichia coli. Here we report the nucleotide sequence of the cheW gene as well as the purification and characterization of the CheW protein. The DNA sequence predicts a protein of 18,000 molecular weight. The pure protein exhibits an apparent molecular weight of 18,000 during sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Molecular sieve chromatography under nondenaturing conditions indicates a molecular weight of approximately 35,000, however. This result suggests that CheW is a homodimer. The predicted amino acid sequence between Thr-128 and Asp-160 fits a consensus exhibited by many proteins which bind purine nucleotides.

Nucleotide sequence corresponding to five chemotaxis genes in Escherichia coli.

The nucleotide sequence of DNA which contains five chemotaxis-related genes of Escherichia coli, cheW, cheR, cheB, cheY, and cheZ, and part of the cheA gene was determined. Molecular weights of the polypeptides encoded by these genes were calculated from translated amino acid sequences, and they were 18,100 for cheW, 32,700 for cheR, 37,500 for cheB, 14,100 for cheY, and 24,000 for cheZ. Nucleotide sequences which could act as ribosome-binding sites were found in the upstream region of each gene. After the termination codon of the cheW gene, a typical rho-independent transcription termination signal was observed. There are no other open reading frames long enough to encode polypeptides in this region except those which code for the two previously reported genes tar and tap.

Phase variation in Salmonella: genetic analysis of a recombinational switch.

The alternative expression of Salmonella genes H1 and H2, which specify different flagellar antigens, results in the oscillation of phenotype known as phase variation. This alternation is controlled by the inversion of an 800-base-pair sequence of DNA adjacent to, or including part of, the H2 gene. The invertable region was presumed to regulate the function of a promoter and to include specific sites at which a recombinational event, resulting in the inversion, could occur. Here we report genetic manipulations of hybrid lambda phage carrying the H2 gene that were used to define the H2 promoter region and the recombinational sites. The H2 gene fragment was inserted on a hybrid lambda phage next to the cheW gene, which lacked a promoter element. In the resulting fusion, cheW gene activity was restored, the expression of the H2 and cheW genes was controlled coordinately by the inversion, and the polarity of transcription and location of the H2 gene could be determined. Evidence from this type of gene fusion suggested that the H2 gene promoter is included in the inversion region. Hybrid H2 phage were constructed that contained substitutions for regions of the H2 gene. In contrast to hybrid lambda containing the H2 gene, which alternate between "on" and "off" states, several substituted lambdaH2 were fixed in the "on" state. A site necessary for the recombinational event must have been removed in these fixed lambdaH2.