Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Identification of the fludioxonil-responsive genes dependent on SskA, SrrA, HogA, and AtfA in Aspergillus nidulans

ABSTRACT: Microarray analysis was used to identify the fludioxonil-responsive genes dependent on SskA, SrrA, HogA, and AtfA in the filamentous fungus Aspergillus nidulans. In order to identify such genes, we conducted the several types of experiment. One was a comparison between wild type treated with fludioxonil and without the treatment (Exp.1). Others were comparison between wild type treated with fludioxonil and each mutant (sskA, Exp.2; srrA, Exp.3; hogA, Exp.4; atfA, Exp.5) treated with fludioxonil. Compared the result of Exp.1 with that of other experiments, we could identify the genes whose expression was induced or repressed in response to fludioxonil in a manner dependent on SskA, SrrA, HogA, or AtfA. KEY WORD; Aspergillus nidulans, fludioxonil, SskA, SrrA, HogA, AtfA Conidia of wild type or each mutant were cultured at 37C in 100ml CD medium containing 2% glucose for 18h and treated with fludioxonil (final concentration; 10 ug/ml) or DMSO (as a solvent control) for 15 min. The mycelia were harvested and frozen in liquid nitrogen, ground to powder, and used for RNA preparation. mRNA was purified and used for hybridization experiments. A total of 2 hybridizations were performed for each microarray experiment described in the summary. The following replicates were carried out: 1. In-slide replicates were carried out for each analysis. 2. Dye swap replicates were carried out for each experiment. The slides were scanned with an Axon GenePix 4000B scanner (Molecular Devices). The resulting TIFF images were imported into GenePix Pro and fluorescent intensity of spots were calculated for each of the Cy3 and Cy5 channels. Global normalization was applied to all analyses. Following normalization, spots whose Cy3 or Cy5 intensity was less than 0 were removed from the data set (the exceptional case was that intensity of the other channel was more than 100). The dye-swap replicates and in-slide replicates were subjected to all analyses. Finally, gene expression ratios (test/reference) were calculated for each replicates. Gene expression was considered to be significantly higher or lower whenever the spot intensity changed by at least 3-fold in all four replicates for each experiment.

ORGANISM(S): Emericella nidulans

SUBMITTER: Daisuke Hagiwara

PROVIDER: E-GEOD-16509 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Transcriptional profiling for Aspergillusnidulans HogA MAPK signaling pathway in response to fludioxonil and osmotic stress.

Hagiwara Daisuke D Asano Yoshihiro Y Marui Junichiro J Yoshimi Akira A Mizuno Takeshi T Abe Keietsu K

Fungal genetics and biology : FG & B 20090722 11

In filamentous fungi, the His-Asp phosphorelay signaling system and HOG pathway are involved in the action of the fungicides, fludioxonil, and iprodione, as well as osmotic and oxidative stress responses. Aspergillusnidulans response regulators (RRs), SskA and SrrA, and histidine kinase (HK), NikA, are involved in the growth inhibitory effects of these fungicides. To gain further insights into the molecular basis for these signaling systems, we performed DNA microarray analyses of fludioxonil an ...[more]

PMID: 19596074

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Project description:Microarray analysis was used to identify the osmotic stress-responsive genes dependent on HogA and AtfA in the filamentous fungus Aspergillus nidulans. In order to identify such genes, we conducted the several types of experiment. One was a comparison between wild type with and without osmotic shock (Exp.6). Others were comparison between wild type with osmotic shock and each mutant (hogA, Exp.7; atfA, Exp.8) with osmotic shock. Compared the result of Exp.6 with other experiments, we could identify the genes whose expression was induced or repressed in response to osmotic stress in a manner dependent on HogA and AtfA. KEY WORD; Aspergillus nidulans, osmotic stress, HogA, AtfA Conidia of wild type or each mutant were cultured at 37C in 100ml CD medium containing 2% glucose for 18h, and 50 ml of 3 M sorbitol was added (final concentration; 1 M) or equivalent volume of water (as a control) for 15 min. The mycelia were harvested and frozen in liquid nitrogen, ground to powder, and used for RNA preparation. mRNA was purified and used for hybridization experiments. A total of 2 hybridizations were performed for each microarray experiment described in the summary. The following replicates were carried out: 1. In-slide replicates were carried out for each analysis. 2. Dye swap replicates were carried out for each experiment. The slides were scanned with an Axon GenePix 4000B scanner (Molecular Devices). The resulting TIFF images were imported into GenePix Pro and fluorescent intensity of spots were calculated for each of the Cy3 and Cy5 channels. Global normalization was applied to all analyses. Following normalization, spots whose Cy3 or Cy5 intensity was less than 0 were removed from the data set (the exceptional case was that intensity of the other channel was more than 100). The dye-swap replicates and in-slide replicates were subjected to all analyses. Finally, gene expression ratios (channel 2/channel 1) were calculated for each replicates. Gene expression was considered to be significantly higher or lower whenever the spot intensity changed by at least 2-fold in all four replicates for each experiment.

Project description:Microarray analysis was used to identify the calcium-responsive genes dependent on CrzA in the filamentous fungus Aspergillus nidulans. In order to identify such genes, we conducted the two types of experiment. One was a comparison between wild type with calcium treatment and wild type without calcium treatment. Another was a comparison between wild type with calcium treatment and crzA mutant with calcium treatment. From a comparison between the results of these experiments, we could identify the genes whose expression was induced or repressed in response to calcium in a manner dependent on CrzA. KEY WORD; Aspergillus nidulans, calcium response, crzA Conidia of wild type or crzA mutant strains were incubated at 37C in CD medium for 18h and treated with or without calcium (final concentration; 50 mM) for 15 min. The mycelia were harvested and frozen in liquid nitrogen, ground to a powder, and used for RNA preparation. mRNA was purified and used for hybridization experiments. A total of 2 hybridizations were performed for each microarray experiment described in the summary. The following replicates were carried out: 1. In-slide replicates were carried out for each analyses. 2. Dye swap replicates were carried out for each analyses. The slides were scanned with an Axon GenePix 4000B scanner (Molecular Devices). The resulting TIFF images were imported into GenePix Pro and fluorescent intensity of spots were calculated for each of the Cy3 and Cy5 channels. Global normalization was applied to all analyses. Following normalization, spots whose Cy3 or Cy5 intensity was less than 0 were removed from the data set (the exceptional case was that intensity of the other channel was more than 100). The dye-swap replicates and in-slide replicates were subjected to all analyses. Finally, gene expression ratios (channel 2/channel 1) were calculated for each replicates. Gene expression was considered to be significantly higher or lower whenever the spot intensity changed by at least 2-fold in all four replicates.

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Project description:Aflatoxins are carcinogenic fungal secondary metabolites. Levels of aflatoxins in agricultural commodities are stringently regulated by many countries. A cluster of genes is responsible for aflatoxin biosynthesis by Aspergillus flavus and other closely related species. Expression of the clustered aflatoxin genes is governed by a complex network of regulatory mechanisms. To better understand the molecular events that are associated with aflatoxin production, transcription profiling by microarray analyses which compared three independent aflatoxigenic A. flavus strains to individual isogenic progenies that no longer produced aflatoxins after serial transfers was carried out. Twenty-two significantly differentially expressed features were identified. After physical mapping using the A. oryzae genome sequence as the reference, the number of unique genes was reduced to 16. Compared to the parental strains, changes in the aflatoxin gene expression levels in the progenies were not significant, which suggests that the inability to produce aflatoxins is not caused by decreased expression. The only gene showing higher expression levels in the progenies is homologous to glutathione S-transferease genes. Overexpression of this gene, named hcc, at six- to nine-fold in an aflatoxigenic A. flavus did not cause discernible changes in colony morphology or aflatoxin production. Loss of aflatoxin production after serial transfers may not result from a single event but caused by multiple factors. Keywords: Compartiave hybridization toxigenic and atoxigenic lines of Aspergillus Aspergillus flavus NRRL 29459, NRRL 29474, and NRRL 29490 are aflatoxigenic strains originated from soil collection in a peanut field (Terrell Co., Georgia, USA). Strains 459B-20-2, 474A-20, and 499A-20 were nonaflatoxigenic isolates obtained after 20 serial transfers of the parental strains on potato dextrose agar slants (Horn and Dorner 2002). Comparsions in each experiment consisted of one aflatoxigenic parental strain and one nonaflatoxigenic progeny, compared after 48- or 72-hr growth. Each comparison was repeated with duplicate dye-flip.

Dataset Information

Identification of the fludioxonil-responsive genes dependent on SskA, SrrA, HogA, and AtfA in Aspergillus nidulans

Publications

Transcriptional profiling for Aspergillusnidulans HogA MAPK signaling pathway in response to fludioxonil and osmotic stress.

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