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Jonathan D Walton

Publications and source records attributed to Jonathan D Walton.

9 recordsLinked to original sources
PubMed2006

HC-toxin.

HC-toxin is a cyclic tetrapeptide of structure cyclo(D-Pro-L-Ala-D-Ala-L-Aeo), where Aeo stands for 2-amino-9,10-epoxi-8-oxodecanoic acid. It is a determinant of specificity and virulence in the interaction between the producing fungus, Cochliobolus carbonum, and its host, maize. HC-toxin qualifies as one of the few microbial secondary metabolites whose ecological function in nature is understood. Reaction to C. carbonum and to HC-toxin is controlled in maize by the Hm1 and Hm2 loci. These loci encode HC-toxin reductase, which detoxifies HC-toxin by reducing the 8-carbonyl group of Aeo. HC-toxin is an inhibitor of histone deacetylases (HDACs) in many organisms, including plants, insects, and mammals, but why inhibition of HDACs during infection by C. carbonum leads to disease is not understood. The genes for HC-toxin biosynthesis (collectively known as the TOX2 locus) are loosely clustered over >500 kb in C. carbonum. All of the known TOX2 genes are present in multiple, functional copies and are absent from natural toxin non-producing isolates. The central enzyme in HC-toxin biosynthesis is a 570-kDa non-ribosomal synthetase encoded by a 15.7-kb open reading frame. Other genes known to be required for HC-toxin encode alpha and beta subunits of fatty acid synthase, which are presumed to contribute to the synthesis of Aeo; a pathway-specific transcription factor; an efflux carrier; a predicted branched-chain amino acid aminotransferase; and an alanine racemase.

Animals↗
PubMed2003

Isolation of the carbon catabolite repressor (CREA) gene from the plant-pathogenic fungus Cochliobolus carbonum.

The CREA gene has been implicated in glucose repression in several fungi. The product of this gene, CreA, binds to the promoter region of several enzymes and down-regulates gene expression. An ortholog of CREA was isolated and characterized from the maize pathogenic fungus, Cochliobolus carbonum. The deduced amino acid sequence of the C. carbonum CREA gene is very similar to the CreA proteins of Aspergillus niger, Gibberella fujikuroi, Sclerotinia sclerotiorum and Trichoderma reesei, as well as the Mig1 protein of Saccharomyces cerevisiae. And like the other fungal proteins, C. carbonum CreA has two zinc finger regions and a nuclear localization signal. Putative CreA binding sequences were also identified in the 5' region of three C. carbonum cell wall degrading enzyme genes suggesting that the protein may play a role in the regulatory process that controls these enzymes expression.

Amino Acid Sequence↗
PubMed2003

Quantitative trait loci and comparative genomics of cereal cell wall composition.

Quantitative trait loci (QTLs) affecting sugar composition of the cell walls of maize (Zea mays) pericarp were mapped as an approach to the identification of genes involved in cereal wall biosynthesis. Mapping was performed using the IBM (B73 x Mo17) recombinant inbred line population. There were statistically significant differences between B73 and Mo17 in content of xylose (Xyl), arabinose (Ara), galactose (Gal), and glucose. Thirteen QTLs were found, affecting the content of Xyl (two QTLs), Ara (two QTLs), Gal (five QTLs), Glc (two QTLs), Ara + Gal (one QTL), and Xyl + Glc (one QTL). The chromosomal regions corresponding to two of these, affecting Ara + Gal and Ara on maize chromosome 3, could be aligned with a syntenic region on rice (Oryza sativa) chromosome 1, which has been completely sequenced and annotated. The contiguous P1-derived artificial chromosome rice clones covering the QTLs were predicted to encode 117 and 125 proteins, respectively. Two of these genes encode putative glycosyltransferases, displaying similarity to carbohydrate-active enzyme database family GT4 (galactosyltransferases) or to family GT64 (C-terminal domain of animal heparan synthases). The results illustrate the potential of using natural variation, emerging genomic resources, and homeology within the Poaceae to identify candidate genes involved in the essential process of cell wall biosynthesis.

Carbohydrate Metabolism↗
PubMed2002

An extended physical map of the TOX2 locus of Cochliobolus carbonum required for biosynthesis of HC-toxin.

In genetic crosses, HC-toxin production in the filamentous fungus Cochliobolus carbonum appears to be controlled by a single locus, TOX2. At the molecular level, TOX2 is composed of at least seven duplicated and coregulated genes involved in HC-toxin biosynthesis, export, and regulation. All copies of four of the TOX2 genes were previously mapped within a 540-kb stretch of DNA in strain SB111. Subsequently, an additional three TOX2 genes, TOXE, TOXF, and TOXG, have been discovered. In this paper we have mapped all copies of the new genes, a total of seven, and show that except for one of the two copies of TOXE, which was previously shown to be on a chromosome of 0.7 Mb in strain SB111, they are all linked to the previously known TOX2 genes within approximately 600 kb of each other on a chromosome of 3.5 Mb. We show here that this chromosome also contains at least one non-TOX2 gene, EXG2, which encodes an exo-beta1,3-glucanase. EXG2 is still present in strains that have undergone spontaneous deletion of up to approximately 1.4 Mb of the 3.5-Mb chromosome. The results contribute to our understanding of the complex organization of the genes involved in HC-toxin biosynthesis and are consistent with the hypothesis that a reciprocal chromosomal translocation accounts for the pattern of distribution of the TOX2 genes in different C. carbonum isolates.

Ascomycota↗
PubMed2002

Characterization of inhibitor-resistant histone deacetylase activity in plant-pathogenic fungi.

HC-toxin, a cyclic peptide made by the filamentous fungus Cochliobolus carbonum, is an inhibitor of histone deacetylase (HDAC) from many organisms. It was shown earlier that the HDAC activity in crude extracts of C. carbonum is relatively insensitive to HC-toxin as well as to the chemically unrelated HDAC inhibitors trichostatin and D85, whereas the HDAC activity of Aspergillus nidulans is sensitive (G. Brosch et al., Biochemistry 40:12855-12863, 2001). Here we report that HC-toxin-resistant HDAC activity was present in other, but not all, plant-pathogenic Cochliobolus species but not in any of the saprophytic species tested. The HDAC activities of the fungi Alternaria brassicicola and Diheterospora chlamydosporia, which also make HDAC inhibitors, were resistant. The HDAC activities of all C. carbonum isolates tested, except one non-toxin-producing isolate, were resistant. In a cross between a sensitive isolate and a resistant isolate, resistance genetically cosegregated with HC-toxin production. When fractionated by anion-exchange chromatography, extracts of resistant and sensitive isolates and species had two peaks of HDAC activity, one that was fully HC-toxin resistant and a second that was larger and sensitive. The first peak was consistently smaller in extracts of sensitive fungi than in resistant fungi, but the difference appeared to be insufficiently large to explain the differential sensitivities of the crude extracts. Differences in mRNA expression levels of the four known HDAC genes of C. carbonum did not account for the observed differences in HDAC activity profiles. When mixed together, resistant extracts protected extracts of sensitive C. carbonum but did not protect other sensitive Cochlibolus species or Neurospora crassa. Production of this extrinsic protection factor was dependent on TOXE, the transcription factor that regulates the HC-toxin biosynthetic genes. The results suggest that C. carbonum has multiple mechanisms of self-protection against HC-toxin.

Ascomycota↗
PubMed1999

A putative branched-chain-amino-acid transaminase gene required for HC-toxin biosynthesis and pathogenicity in Cochliobolus carbonum.

The cyclic tetrapeptide HC-toxin is required for pathogenicity of the filamentous fungus Cochliobolus carbonum on maize. HC-toxin production is controlled by a complex locus, TOX2. The isolation and characterization of a new gene of the TOX2 locus, TOXF, is reported. It is shown that TOXF is specifically required for HC-toxin production and pathogenicity. It is present as two or three copies in all HC-toxin-producing (Tox2+) isolates and is absent in toxin-non-producing strains. The deduced amino acid sequence of TOXF has moderate homology to many known or putative branched-chain-amino-acid transaminases from various species. A strain of C. carbonum with all copies of TOXF disrupted grew normally but lost HC-toxin production and pathogenicity. It is proposed that TOXF has a biosynthetic role in HC-toxin synthesis, perhaps to aminate a precursor of Aeo (2-amino-9,10-epoxi-8-oxodecanoic acid).

Amino Acid Sequence↗
PubMed1999

A unique eukaryotic beta-xylosidase gene from the phytopathogenic fungus Cochliobolus carbonum.

The plant-pathogenic fungus Cochliobolus carbonum secretes one major beta-xylosidase (Xyp1) when grown on xylan or maize cell walls. cDNA and genomic DNA encoding Xyp1 were isolated using PCR primers based on peptide sequences from the purified protein. XYP1 contains three introns, has 5' and 3' untranslated regions of 74 and 145 bp, respectively, and is predicted to encode a protein of 328 amino acids (Mr 36700) with four N-glycosylation sites. Although it is secreted, Xyp1 has no predicted signal peptide. Furthermore, Xyp1 appears not to be processed at the N-terminus because one of the peptides isolated from the mature protein is located only six amino acids downstream of the translational start methionine. The primary sequence of Xyp1 is unrelated to any known eukaryotic beta-xylosidase but has 35% overall identity to two bacterial bifunctional beta-xylosidase/alpha-arabinosidases. Mutation of XYP1 by targeted gene replacement resulted in the loss of the major beta-xylosidase activity corresponding to the product of XYP1, but a significant amount of secreted beta-xylosidase activity (25% of wild-type) remained in the culture filtrates. The xyp1 mutant was still fully pathogenic on maize.

Amino Acid Sequence↗
PubMed1996

A putative cyclic peptide efflux pump encoded by the TOXA gene of the plant-pathogenic fungus Cochliobolus carbonum.

Race 1 isolates of Cochliobolus carbonum are pathogenic on certain maize lines due to production of a host-selective cyclic tetrapeptide, HC-toxin. Flanking HTS1, which encodes the central enzyme in HC-toxin biosynthesis, a gene was identified and named TOXA. Like HTS1, TOXA occurred only in isolates of the fungus that make HC-toxin and was present as two linked copies in most toxin-producing isolates. HTS1 and TOXA were transcribed in the opposite orientation and their transcriptional start sites were 386 bp apart. The predicted product of TOXA was a 58 kDa hydrophobic protein with 10-13 membrane-spanning regions. The sequence was highly similar to several members of the major facilitator superfamily that confer resistance to tetracycline, methylenomycin, and other antibiotics. Although it was possible to mutate one copy or the other of TOXA by targeted gene disruption, numerous attempts to disrupt both copies in a single strain were unsuccessful, suggesting that TOXA is an essential gene in strains that synthesize HC-toxin. On the basis of its presence only in HC-toxin-producing strains, its proximity to HTS1 and its predicted amino acid sequence, we propose that TOXA encodes an HC-toxin efflux pump which contributes to self-protection against HC-toxin and/or the secretion of HC-toxin into the extracellular milieu.

Amino Acid Sequence↗