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Characterization of Cercospora nicotianae Hypothetical Proteins in Cercosporin Resistance.

Beseli A, Noar R, Daub ME - PLoS ONE (2015)

Bottom Line: In previous research, the zinc cluster transcription factor CRG1 (cercosporin resistance gene 1) was found to be crucial for Cercospora species' resistance against cercosporin, and subtractive hybridization analysis identified 185 genes differentially expressed between Cercospora nicotianae wild type (wt) and a crg1 mutant.Gene disruption was used to generate C. nicotianae 71cR mutants; these mutants did not differ from wt C. nicotianae in cercosporin resistance or production.Analysis of 71cR conserved domains and secondary and tertiary structure identify the protein as having an NTF2-like superfamily DUF1348 domain with unknown function, to be intracellular and localized in the cytosol, and to have similarities to proteins in the steroid delta-isomerase family.

View Article: PubMed Central - PubMed

Affiliation: Department of Plant and Microbial Biology, North Carolina State University, Raleigh, North Carolina, United States of America.

ABSTRACT
The photoactivated toxin, cercosporin, produced by Cercospora species, plays an important role in pathogenesis of this fungus to host plants. Cercosporin has almost universal toxicity to cells due to its production of reactive oxygen species including singlet oxygen. For that reason, Cercospora species, which are highly resistant to their own toxin, are good candidates to identify genes for resistance to cercosporin and to the reactive oxygen species it produces. In previous research, the zinc cluster transcription factor CRG1 (cercosporin resistance gene 1) was found to be crucial for Cercospora species' resistance against cercosporin, and subtractive hybridization analysis identified 185 genes differentially expressed between Cercospora nicotianae wild type (wt) and a crg1 mutant. The focus of this work was to identify and characterize the hypothetical proteins that were identified in the Cercospora nicotianae subtractive library as potential resistance factors. Quantitative RT-PCR analysis of the 20 genes encoding hypothetical proteins showed that two, 24cF and 71cR, were induced under conditions of cercosporin toxicity, suggesting a role in resistance. Transformation and expression of 24cF and 71cR in the cercosporin-sensitive fungus, Neurospora crassa, showed that 71cR provided increased resistance to cercosporin toxicity, whereas no significant increase was observed in 24cF transformants. Gene disruption was used to generate C. nicotianae 71cR mutants; these mutants did not differ from wt C. nicotianae in cercosporin resistance or production. Quantitative RT-PCR analysis showed induction of other resistance genes in the 71cR mutant that may compensate for the loss of 71cR. Analysis of 71cR conserved domains and secondary and tertiary structure identify the protein as having an NTF2-like superfamily DUF1348 domain with unknown function, to be intracellular and localized in the cytosol, and to have similarities to proteins in the steroid delta-isomerase family.

No MeSH data available.


Related in: MedlinePlus

Distribution of 71cR orthologs among sequenced fungal species.For each genome sequence available in the JGI MycoCosm portal, tblastn was used to search the filtered model transcripts for 71cR orthologs. To the right of each taxon, the number of analyzed genome sequences is shown, along with the percent of genomes in the taxon for which a 71cR ortholog was identified.
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pone.0140676.g007: Distribution of 71cR orthologs among sequenced fungal species.For each genome sequence available in the JGI MycoCosm portal, tblastn was used to search the filtered model transcripts for 71cR orthologs. To the right of each taxon, the number of analyzed genome sequences is shown, along with the percent of genomes in the taxon for which a 71cR ortholog was identified.

Mentions: To analyze the distribution of 71cR among fungal species, orthologs of the 71cR protein sequence were identified from sequenced fungal genomes in the JGI MycoCosm portal [33,34]. With one exception, 71cR orthologs were found in fungal genomes in all classes within the Dikarya (Basidiomycota and Ascomycota), including both mycelial and yeast species (Fig 7). Distribution was not ubiquitous, however, ranging from less than 5% to 100% of sequenced genomes within a taxonomic group. 71cR orthologs were especially common within the Ascomycota sub-phylum Pezizomycotina and the Basidiomycota sub-phylum Agaricomycotina. Cercospora spp. are in class Dothideomycetes within sub-phylum Pezizomycotina, and 85% of the 96 sequenced species within this class contained 71cR orthologs. While the Cercospora canescens genome contained a close ortholog, we were unable to identify an ortholog within the Cercospora zeae-maydis genome sequence. Distribution was less or lacking outside of the Dikarya. The one sequenced genome from the Glomeromycota (Rhizophagus irregularis) also had a 71cR ortholog, but 71cR orthologs were not found from other clades traditionally grouped into Zygomycota (such as Mucoromycotina and Kickxellomycotina) or from basal fungal lineages such as Neocallimastigomycota, Chytridiomycota, and Microsporidia (Fig 7). Due to the low number of sequenced genomes from these basal fungal lineages, however, it is not possible to definitively determine whether 71cR orthologs are completely absent within these taxa.


Characterization of Cercospora nicotianae Hypothetical Proteins in Cercosporin Resistance.

Beseli A, Noar R, Daub ME - PLoS ONE (2015)

Distribution of 71cR orthologs among sequenced fungal species.For each genome sequence available in the JGI MycoCosm portal, tblastn was used to search the filtered model transcripts for 71cR orthologs. To the right of each taxon, the number of analyzed genome sequences is shown, along with the percent of genomes in the taxon for which a 71cR ortholog was identified.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4608573&req=5

pone.0140676.g007: Distribution of 71cR orthologs among sequenced fungal species.For each genome sequence available in the JGI MycoCosm portal, tblastn was used to search the filtered model transcripts for 71cR orthologs. To the right of each taxon, the number of analyzed genome sequences is shown, along with the percent of genomes in the taxon for which a 71cR ortholog was identified.
Mentions: To analyze the distribution of 71cR among fungal species, orthologs of the 71cR protein sequence were identified from sequenced fungal genomes in the JGI MycoCosm portal [33,34]. With one exception, 71cR orthologs were found in fungal genomes in all classes within the Dikarya (Basidiomycota and Ascomycota), including both mycelial and yeast species (Fig 7). Distribution was not ubiquitous, however, ranging from less than 5% to 100% of sequenced genomes within a taxonomic group. 71cR orthologs were especially common within the Ascomycota sub-phylum Pezizomycotina and the Basidiomycota sub-phylum Agaricomycotina. Cercospora spp. are in class Dothideomycetes within sub-phylum Pezizomycotina, and 85% of the 96 sequenced species within this class contained 71cR orthologs. While the Cercospora canescens genome contained a close ortholog, we were unable to identify an ortholog within the Cercospora zeae-maydis genome sequence. Distribution was less or lacking outside of the Dikarya. The one sequenced genome from the Glomeromycota (Rhizophagus irregularis) also had a 71cR ortholog, but 71cR orthologs were not found from other clades traditionally grouped into Zygomycota (such as Mucoromycotina and Kickxellomycotina) or from basal fungal lineages such as Neocallimastigomycota, Chytridiomycota, and Microsporidia (Fig 7). Due to the low number of sequenced genomes from these basal fungal lineages, however, it is not possible to definitively determine whether 71cR orthologs are completely absent within these taxa.

Bottom Line: In previous research, the zinc cluster transcription factor CRG1 (cercosporin resistance gene 1) was found to be crucial for Cercospora species' resistance against cercosporin, and subtractive hybridization analysis identified 185 genes differentially expressed between Cercospora nicotianae wild type (wt) and a crg1 mutant.Gene disruption was used to generate C. nicotianae 71cR mutants; these mutants did not differ from wt C. nicotianae in cercosporin resistance or production.Analysis of 71cR conserved domains and secondary and tertiary structure identify the protein as having an NTF2-like superfamily DUF1348 domain with unknown function, to be intracellular and localized in the cytosol, and to have similarities to proteins in the steroid delta-isomerase family.

View Article: PubMed Central - PubMed

Affiliation: Department of Plant and Microbial Biology, North Carolina State University, Raleigh, North Carolina, United States of America.

ABSTRACT
The photoactivated toxin, cercosporin, produced by Cercospora species, plays an important role in pathogenesis of this fungus to host plants. Cercosporin has almost universal toxicity to cells due to its production of reactive oxygen species including singlet oxygen. For that reason, Cercospora species, which are highly resistant to their own toxin, are good candidates to identify genes for resistance to cercosporin and to the reactive oxygen species it produces. In previous research, the zinc cluster transcription factor CRG1 (cercosporin resistance gene 1) was found to be crucial for Cercospora species' resistance against cercosporin, and subtractive hybridization analysis identified 185 genes differentially expressed between Cercospora nicotianae wild type (wt) and a crg1 mutant. The focus of this work was to identify and characterize the hypothetical proteins that were identified in the Cercospora nicotianae subtractive library as potential resistance factors. Quantitative RT-PCR analysis of the 20 genes encoding hypothetical proteins showed that two, 24cF and 71cR, were induced under conditions of cercosporin toxicity, suggesting a role in resistance. Transformation and expression of 24cF and 71cR in the cercosporin-sensitive fungus, Neurospora crassa, showed that 71cR provided increased resistance to cercosporin toxicity, whereas no significant increase was observed in 24cF transformants. Gene disruption was used to generate C. nicotianae 71cR mutants; these mutants did not differ from wt C. nicotianae in cercosporin resistance or production. Quantitative RT-PCR analysis showed induction of other resistance genes in the 71cR mutant that may compensate for the loss of 71cR. Analysis of 71cR conserved domains and secondary and tertiary structure identify the protein as having an NTF2-like superfamily DUF1348 domain with unknown function, to be intracellular and localized in the cytosol, and to have similarities to proteins in the steroid delta-isomerase family.

No MeSH data available.


Related in: MedlinePlus