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A W Johnston

Publications and source records attributed to A W Johnston.

At least 19 recordsLinked to original sources
PubMed1998

Is the fur gene of Rhizobium leguminosarum essential?

Using primers corresponding to conserved regions of the bacterial regulatory gene fur, a homologue of this gene from the genome of Rhizobium leguminosarum biovar viciae, the nitrogen-fixing symbiont of peas, was isolated and sequenced. The fur gene is normally expressed constitutively, independent of the presence of Fe in the medium, but in one Rhizobium strain it was transcribed at a low level. Attempts to isolate a fur knockout mutant failed, suggesting that the gene is essential for free-living growth. In other bacteria, certain fur mutations confer manganese resistance; however, none of the manganese-resistant mutants of R. leguminosarum which we isolated was corrected by the cloned fur gene. When the cloned R. leguminosarum fur gene was introduced into a fur mutant of Escherichia coli, it caused some Fe-dependent reduction in the amount of siderophore, indicating that it can function heterologously.

Amino Acid Sequence↗
PubMed1997

Iron-dependent transcription of the regulatory gene ros of Agrobacterium radiobacter.

Transcription of the regulatory gene ros of Agrobacterium radiobacter requires growth in the presence of Fe although this regulation was not mediated by ros itself. The ros gene repressed its own transcription, independently of the Fe status of the growth media. It was shown that the two cysteine residues in the Ros protein were essential for the complementation of the exopolysaccharide synthesis defect of ros mutant strains. It was found that the mutation in one exo mutant strain that was complemented both by ros and by the "structural" exoY gene was not, in fact, in ros but is in some other, unknown gene. The two cysteines were also essential for the correction of this mutant. This mutant affected the expression of exoY but not of ros.

Alleles↗
PubMed1996

A region of a Sym plasmid of Rhizobium leguminosarum biovar phaseoli has similarity to prokaryotic insertion sequences and to eukaryotic integrases.

Near the nod and nif genes of the Sym plasmid pRP2JI of Rhizobium leguminosarum biovar phaseoli are three open reading frames whose deduced polypeptide products have similarities to those of genes in bacterial insertion sequences. The similarity of one of these ORFs was significantly greater to that of the integrase region of pol proteins of eukaryotic retroviruses and transposable elements in animals and plants than it was to the transposases of prokaryotic insertion sequences. In the noncoding region of the IS-like element, there was a sequence similar to that which had been identified close to nod genes in Azorhizobium caulinodans.

Amino Acid Sequence↗
PubMed1996

Histoplasmosis--a ten year follow-up.

A patient with chronic disseminated histoplasmosis was followed after successful treatment with amphotericin B for 10 years until his death from chronic obstructive airways disease. Necropsy showed that Histoplasma organisms could still be identified in the adrenal and pancreatic abscesses, though none appeared viable on electron microscopy. Excision of such abscesses should be considered in view of their potential as a reservoir of organisms.

Aged↗
PubMed1996

The C-terminal domain of the Rhizobium leguminosarum chitin synthase NodC is important for function and determines the orientation of the N-terminal region in the inner membrane.

The nodC genes from rhizobia encode an N-acetylglucosaminyl transferase (chitin synthase) involved in the formation of lipo-chito-oligosaccharide Nod factors that initiate root nodule morphogenesis in legume plants. NodC proteins have two hydrophobic domains, one of about 21 residues at the N-terminus and a longer one, which could consist of two or three transmembrane spans, near the C-terminus. These two hydrophobic domains flank a large hydrophilic region that shows extensive homology with other beta -glycosyl transferases. The topology NodC in the inner membrane of Rhizobium leguminosarum biovar viciae was analysed using a series of gene fusions encoding proteins in which NodC was fused to alkaline phosphatase (PhoA) lacking an N-terminal transit sequence or to beta-galactosidase (LacZ). Our data support a model in which the N-terminal hydrophobic domain spans the membrane in a Nout-Cin orientation, with the adjacent large hydrophilic domain being exposed to the cytoplasm. This orientation appears to depend upon the presence of the hydrophobic region near the C-terminus. We propose that this hydrophobic region contains three transmembrane spans, such that the C-terminus of NodC is located in the periplasm. A short region of about 40 amino acids, encompassing the last transmembrane span, is essential for the function of NodC. Our model for NodC topology suggests that most of NodC, including the region showing most similarity to other beta-glycosyl transferases, is exposed to the cytoplasm, where it is likely that polymerization of N-acetyl glucoasamine occurs. Such a model is incompatible with previous reports suggesting that NodC spans both inner and other membranes.

Alkaline Phosphatase↗
PubMed1996

Expression of the exoY gene, required for exopolysaccharide synthesis in Agrobacterium, is activated by the regulatory ros gene.

Some mutants of Agrobacterium radiobacter, defective in exopolysaccharide synthesis, were phenotypically complemented by two different regions of cloned chromosomal DNA. One of these had been shown to contain a gene termed ros, a novel class of transcriptional regulator. The other contains a gene termed exoY which encodes a glycosyltransferase that is involved in one of the early steps in exopolysaccharide synthesis. Mutations in ros reduced the expression of exoY and a model to account for the complementation of certain exo alleles by both ros and exoY is presented. TnphoA insertions into exoY which expressed alkaline phosphatase activity were isolated and mapped, confirming the membrane location of the exoY gene product. Some of these mutations were dominant, causing merodiploids to be non-mucoid. exoY is linked to two genes, one encoding an omega-aminotransferase and the other encoding an aldehyde dehydrogenase.

ATP Binding Cassette Transporter, Subfamily D, Mem↗
PubMed1995

Rhizobium leguminosarum nodulation gene (nod) expression is lowered by an allele-specific mutation in the dicarboxylate transport gene dctB.

To identify host genes that might influence nod (nodulation) gene expression in Rhizobium leguminosarum, a nodC-phoA reporter plasmid (carrying nodD) was introduced into a chemically mutagenized population of a R. leguminosarum strain lacking a symbiotic plasmid. The transconjugants were screened for expression of alkaline phosphatase (PhoA) on plates containing hesperetin, an inducer of nod genes, and a mutant with reduced expression was identified. When the nodC-phoA plasmid was cured from the mutant and the symbiotic plasmid pRL1Jl introduced, the mutant formed nodules, but symbiotic nitrogen fixation was less than 20% of normal. When the nodC-phoA allele was introduced on pRL1Jl a low level of nod gene induction was found. The reduced nodC expression appeared to be caused by a decrease in expression of the regulatory gene nodD, since expression of a nodD-lacZ fusion was also lower in the mutant than in the control. These mutant phenotypes and the low nitrogen fixation were complemented with a plasmid (plJ1848) from a R. leguminosarum cosmid library. DNA hybridization confirmed that plJ1848 was not from the symbiotic plasmid and showed that a DNA insertion was present in the mutant. The complementing region of plJ1848 was defined by transposon mutagenesis; DNA sequencing revealed that it carried the dicarboxylic acid transport (dct) genes. However, the mutant grew well with succinate as sole C-source. Genetic analysis revealed that the mutant appeared to contain IS50 in the regulatory gene dctB and that this mutation caused the reduction in nod gene expression. The effect was allele-specific since other mutations in dctB did not influence nod gene expression. Surprisingly, the mutant had a constitutive high level of succinate transport, indicating that the mutation caused unregulated expression of dctA the structural gene for dicarboxylic acid transport. This in some way appears to have lowered the expression of nodD, indicating that the nodD promoter may be influenced by the metabolic status of the cells or by expression of dctD in the absence of dctB.

Alkaline Phosphatase↗
PubMed1995

Characterization of the cycHJKL genes involved in cytochrome c biogenesis and symbiotic nitrogen fixation in Rhizobium leguminosarum.

Mutants of Rhizobium leguminosarum bv. viciae unable to respire via the cytochrome aa3 pathway were identified by the inability to oxidize N,N'-dimethyl-p-phenylenediamine. Two mutants which were complemented by cosmid pIJ1942 from an R. leguminosarum clone bank were identified. Although pea nodules induced by these mutants contained many bacteroids, no symbiotic nitrogen fixation was detected. Heme staining of cellular proteins revealed that all cytochrome c-type heme proteins were absent. These mutants lacked spectroscopically detectable cytochrome c, but cytochromes aa3 and d were present, the latter at a higher-than-normal level. DNA sequence analysis of complementing plasmids revealed four apparently cotranscribed open reading frames (cycH, cycJ, cycK, and cycL). CycH, CycJ, CycK, and CycL are homologous to Bradyrhizobium japonicum and Rhizobium meliloti proteins thought to be involved in the attachment of heme to cytochrome c apoproteins; CycK and CycL are also homologous to the Rhodobacter capsulatus ccl1 and ccl2 gene products and the Escherichia coli nrfE and nrfF gene products involved in the assembly of c-type cytochromes. The absence of cytochrome c heme proteins in these R. leguminosarum mutants is consistent with the view that the cycHJKL operon could be involved in the attachment of heme to apocytochrome c.

Amino Acid Sequence↗
PubMed1995

Recording eye movements using coaxial cameras--applications for visual ergonomics and reading studies.

We developed a system of coaxial video cameras that records monocular eye position and scene, and superimposes these images using a digital video mixer. We mounted miniature video cameras above and below a cube beam-splitting prism in the spectacle plane. An infrared emitting diode was imaged in the cornea to locate eye position. The technique was accurate to about 0.5 degrees within 15 degrees of primary gaze; however, we see its main advantages as being its low cost and simple design that, for some applications, does not require complex computer analysis and data manipulation. With improved camera optics, it has the potential for helmet mounting and use remote from a recording console. We used the instrument to monitor a reader's eye position when using low vision devices, and see applications of the technique in the field of visual ergonomics and sports vision.

Calibration↗
PubMed1994

Sequence and regulation of psrA, a gene on the Sym plasmid of Rhizobium leguminosarum biover phaseoli which inhibits transcription of the psi genes.

The psr region of Rhizobium leguminosarum biovar phaseoli had originally been recognized on the basis of its ability to repress the transcription of the psi genes, one of which, psiA, inhibits exopolysaccharide synthesis when cloned in multi-copy plasmids. Both psr and psi are located on the symbiotic plasmid pRP2JI. The psrA gene was localized and sequenced. The deduced amino acid sequence of PsrA was shown to have similarity to the DNA-binding region of a family of other transcriptional regulators, consistent with its known effects on the expression of psi. The transcription of psrA itself appears to be constitutive in free-living Rhizobium, but is regulated by another gene on the Sym plasmid pRP2JI.

Amino Acid Sequence↗
PubMed1994

The psi operon of Rhizobium leguminosarum biovar phaseoli: identification of two genes whose products are located at the bacterial cell surface.

We have delineated three short open reading frames, psiA, ORF-P and psiB within the psi operon of Rhizobium leguminosarum biovar phaseoli. psiA, in a multi-copy plasmid, causes inhibition of exopolysaccharide synthesis in R. leguminosarum. In addition, the suppression of exopolysaccharide synthesis due to the multi-copy psiA caused R. leguminosarum strains to stain with the dye calcofluor, a response that does not occur with wild-type strains of this species. Insertions of a defective phoA gene (lacking its promoter, ribosomal binding site and leader sequence) into psiA and psiB were isolated and the precise locations of the insertions were established. PsiA-PhoA and PsiB-PhoA protein fusions were found to express alkaline phosphatase activity indicating that PsiA and PsiB span the inner membrane or are translocated across it.

Alkaline Phosphatase↗
PubMed1994

Tuberous sclerosis--an unusual cause of seizures in an 18 year old.

An 18 year old man presenting with seizures was found to have hypomelanotic macules and a cardiac rhabdomyoma. These features suggested a diagnosis of tuberous sclerosis (TS) but there were no other clinical signs, no family history and cranial imaging failed to reveal the characteristic appearances. The diagnostic criteria for TS are reviewed and the importance of thorough clinical examination and appropriate investigation in this disease is stressed.

Adolescent↗
PubMed1992

Secretion of the Rhizobium leguminosarum nodulation protein NodO by haemolysin-type systems.

The Rhizobium leguminosarum biovar viciae nodulation protein NodO is partially homologous to haemolysin of Escherichia coli and, like haemolysin, is secreted into the growth medium. The NodO protein can be secreted by a strain of E. coli carrying the cloned nodO gene plus the haemolysin secretion genes hlyBD, in a process that also requires the outer membrane protein encoded by tolC. The related protease secretion genes, prtDEF, from Erwinia chrysanthemi also enable E. coli to secrete NodO. The Rhizobium genes encoding the proteins required for NodO secretion are unlinked to nodO and are unlike other nod genes, since they do not require flavonoids or NodO for their expression. Although proteins similar to NodO were not found in rhizobia other than R. leguminosarum bv. viciae, several rhizobia and an Agrobacterium strain containing the cloned nodO gene were found to have the ability to secrete NodO. These observations indicate that a wide range of the Rhizobiaceae have a protein secretion mechanism analogous to that which secretes haemolysin and related toxins and proteases in the ENterobacteriaceae.

Bacterial Proteins↗
PubMed1992

One hundred requests for predictive testing for Huntington's disease.

In Grampian region, the first 100 requests for predictive testing for Huntington's disease have been analysed. Forty-five presymptomatic predictive test results have been given, eleven families have made a decision to undergo prenatal exclusion tests, and so far eleven pregnancies have been investigated using chorion villus biopsy. This paper presents the protocol used and a descriptive analysis of those receiving results, those choosing to leave the programme, and those unable to complete it because of an unsuitable family structure or unco-operative relatives.

Adult↗
PubMed1992

Molecular characterization and regulation of the rhizosphere-expressed genes rhiABCR that can influence nodulation by Rhizobium leguminosarum biovar viciae.

A group of four rhi (rhizosphere-expressed) genes from the symbiotic plasmid of Rhizobium leguminosarum biovar viciae has been characterized. Although mutation of the rhi genes does not normally affect nodulation, in the absence of the closely linked nodulation genes nodFEL, mutations in the rhi genes can influence the nodulation of the vetch Vicia hirsuta. The DNA sequence of the rhi gene region reveals four large open reading frames, three of them constituting an operon (rhiABC) transcribed convergently toward the fourth gene, rhiR. rhiABC are under the positive control of RhiR, the expression of which is repressed by flavonoids that normally induce nod gene expression. This repression, which requires the nodD gene product (the transcriptional activator of nod gene expression), may be due to a cis effect caused by a high level of NodD-dependent expression from the adjacent nodO promoter, which is transcribed divergently from rhiR. RhiR shows significant similarities to a subfamily of transcriptional regulators that includes the LuxR and UvrC-28K proteins. RhiA shows limited homology to a short domain of the lactose permease, LacY, close to a region thought to be involved in substrate binding. No strong homologies were found for the other rhi gene products. It appears that RhiA and RhiB are cytoplasmic, whereas RhiC is a periplasmic protein, since it has a typical N-terminal transit sequence and a rhiC-phoA protein fusion expresses alkaline phosphatase activity. The biochemical role of the rhi genes has not been established, but it appears that they may play a role in the plant-microbe interaction, possibly by allowing the bacteria to metabolize a plant-made metabolite.

Amino Acid Sequence↗
PubMed1992

Cardiofaciocutaneous syndrome with new ectodermal manifestations.

We describe a 7 year old girl whose features satisfy the diagnosis of cardiofaciocutaneous syndrome. Her ectodermal features consist of fine, sparse hair, thin, opalescent nails, finger tip pads, generalised pigmentation of the skin, but no hyperkeratosis. Skin pigmentation and finger tip pads have not been previously reported in this syndrome. Twenty-two cases of CFC have been described but there is debate as to whether it is distinct from Noonan syndrome.

Abnormalities, Multiple↗