Project description:To understand the roles of DNA methylation and histone deacetylation in plant gene network that controls plant tolerance to freezing treatment, we used Affymetrix GeneChips ATH-121501 to analysis. For microarray analysis, Arabidopsis (ecotype Columbia) seedlings grew in four types of sterile growth medium: no any other modifying agents, added 7g/ml 0.5 M aza-dC in water (DNA methylation inhibitor), added TSA in methanol (histone deacetylation inhibitor), both aza-dC and TSA, all for 16 day in growth chamber (16-h day length at 70% relative humidity, 23 °C) , then all seedlings were cold treated at 0 °C for 24 h in growth chamber (16-h day length at 70% relative humidity). A total of 3305 genes expression were statistically analysized. Our study provides some clues that the transcript levels of some cold-responsive genes regulated by DNA methylation and histone deacetylation. This will be valuable for understanding gene regulation by epigenetic modifications under freezing stress. Keywords: Cold Stress response, DNA methylation, histone deacetylation
Project description:To understand the roles of DNA methylation and histone deacetylation in plant gene network that controls plant tolerance to freezing treatment, we used Affymetrix GeneChips ATH-121501 to analysis. For microarray analysis, Arabidopsis (ecotype Columbia) seedlings grew in four types of sterile growth medium: no any other modifying agents, added 7g/ml 0.5 M aza-dC in water (DNA methylation inhibitor), added TSA in methanol (histone deacetylation inhibitor), both aza-dC and TSA, all for 16 day in growth chamber (16-h day length at 70% relative humidity, 23 M-BM-0C) , then all seedlings were cold treated at 0 M-BM-0C for 24 h in growth chamber (16-h day length at 70% relative humidity). A total of 3305 genes expression were statistically analysized. Our study provides some clues that the transcript levels of some cold-responsive genes regulated by DNA methylation and histone deacetylation. This will be valuable for understanding gene regulation by epigenetic modifications under freezing stress. Keywords: Cold Stress response, DNA methylation, histone deacetylation Three replicates for aza-dc and TSA treatment, and two replicates for Mock and both aza-dc and TSA treatment. All samples were treated 0M-BM-0C 24h.
Project description:Blocking histone deacetylation with trichostatin A (TSA) or blocking cytosine methylation using 5-aza-2'-deoxycytosine (aza-dC) can derepress silenced genes in multicellular eukaryotes, including animals and plants. We questioned whether DNA methylation and histone deacetylation overlap in the regulation of endogenous plant genes by monitoring changes in expression of ~7800 Arabidopsis thaliana genes following treatment with azadC, TSA, or both chemicals together. RNA levels for ~4% of the genes were reproducibly changed 3-fold or more by at least one treatment. Distinct subsets of genes are up-regulated or down-regulated in response to aza-dC, TSA, or simultaneous treatment with both chemicals, with little overlap among subsets. Surprisingly, the microarray data indicate that TSA and aza-dC are often antagonistic rather than synergistic in their effects. Analysis of green fluorescent protein transgenic plants confirmed this finding, showing that TSA can block the up-regulation of silenced green fluorescent protein transgenes in response to aza-dC or a ddm1 (decrease in DNA methylation 1) mutation. Our results indicate that global inhibition of DNA methylation or histone deacetylation has complex, nonredundant effects for the majority of responsive genes and suggest that activation of some genes requires one or more TSA-sensitive deacetylation events in addition to cytosine demethylation.
Project description:Histone 3 lysine 4 and histone 3 lysine 9 methylation in wild type and ddm1 Arabidopsis thaliana seedlings. The purpose of the chromatin immunoprecipitation/microarray (ChIP/chip) experiment is to determine which regions of a genome are enriched for a particular histone modification in a single Arabidopsis thanliana genotype. Chromatin immunoprecipitation with antibodies raised against dimethyl histone-H3 lysine-9 (H3mK9) or dimethyl histone-H3 lysine-4 (H3mK4) is performed on a selected genotype. This purified DNA from each immunoprecipiation (mH3K9, mH3K4, no antibody control) is used for random amplification to increase the quantity of DNA for microarray hybridization. The amplified DNA from each experimental sample is then labeled with Cy5 and hybridized against total input DNA from the corresponding genotype, labeled in Cy3. In a single hybridization, the total input DNA serves as a baseline and is compared to the immunoprecipitated samples. Ratios of normalized signal intensities were calculated to identify enrichment of a particular sequence after immunoprecipitation, in comparison to the total input DNA. Dye swap analysis is carried out to take account of experimental variation by repeating the hybridization with identical samples labeled with Cy3 and Cy5, respectively. This SuperSeries is composed of the following subset Series: GSE1333: EV49+50, Histone 3 Lysine 4 methylation in wild-type Arabidopsis thaliana seedlings GSE1334: Histone 3 Lysine 4 methylation in ddm1 Arabidopsis thaliana seedlings GSE1335: EV104+105, Histone 3 Lysine 4 methylation in ddm1 Arabidopsis thaliana seedlings GSE1336: Ev106+107, Histone 3 Lysine 4 methylation in WT Arabidopsis thaliana seedlings GSE1337: EV51+52, Histone 3 Lysine 9 methylation in wild-type Arabidopsis thaliana seedlings GSE1338: EV59+60, Histone 3 Lysine 9 methylation in ddm1 Arabidopsis thaliana seedlings GSE1339: Histone 3 Lysine 9 methylation in wild-type Arabidopsis thaliana seedlings GSE1340: EV110+111, Histone 3 Lysine 9 methylation in ddm1 Arabidopsis thaliana seedlings Refer to individual Series