第73回千葉医学会総会記事

Microarray data of genes with lower expression levels (<0.74, p < 0.05) in 35S:ROXY19 plants as compared to control plants and their response to TIBA in the Col-0 background. The table contains the gene identity (AGI), description, the mean expression values of four independent biological replicates of the genotypes Col-0 and 35S:ROXY19#8, the ratios (FC, fold changes, log2) of the transcript levels in the transgenic line with respect to Col-0 and the corresponding p-values, the ratios between Col-0 treated with 0.1 % DMSO and Col-0 treated with 0.1 mM TIBA/0.1 % DMSO and the corresponding p-values. Since the microarray analysis of the TIBA-treated plants was performed with the Affimetrix ATH1 gene chip, the list contains 301 and not 321 genes. Genes that are not induced by TIBA are shown in light grey. FC, fold changes. For TIBA induction, plants were grown for six to seven weeks on steamed soil (Archut, Fruhstorfer Erde, T25, Str1fein) in growth chambers with light intensity at 37 to 45 μmol photons m−2 s−1 at 22 °C and 60 % humidity. Eight plants were sprayed with either 0.1 mM TIBA/0.1 % DMSO or with 0.1 % DMSO and leaves were harvested after eight h. The experiment was repeated three times. (XLSX 205 kb

Additional file 2: Table S2. of Ectopically expressed glutaredoxin ROXY19 negatively regulates the detoxification pathway in Arabidopsis thaliana

Microarray data of genes with lower expression levels (FC < 0.74, p < 0.05) in 35S:ROXY19 plants as compared to control plants. The table contains the gene identity (AGI), description, the mean expression values of four independent biological replicates of the genotypes Col-0, 35S:GRXC2, 35S:ROXY19 SSMS , 35S:ROXY19#8 and 35S:ROXY19#12, and the ratios (FC, fold changes, log2) of the transcript levels in the transgenic lines with respect to Col-0 and the corresponding p-values. (XLSX 305 kb

Glucocorticoids regulate a large part of the diurnal transcriptome.

<p>(A) <i>rx3</i> mutant phenotypes. Blue: <i>pomc</i> expression in larval head. sib: wild-type sibling. (B) Sampling schedules of the transcriptomics studies. Zebrafish embryos/larvae were raised under 12 h light (yellow) / 12 h dark (black) cycles. dpf: days post fertilization. ZT: <i>Zeitgeber</i> Time in hours after lights on (ZT0); lights off: ZT12. (C,D) Heatmaps of normalized mRNA expression levels of model 11 (C) and model 5-6-14-15 (D) genes. Red, high expression. Green, low expression. Expression levels normalized by the mean of each gene within each phenotype combination. (E) Overview of gene expression profiles in the different models. Black curved line: oscillation with same phase and amplitude as wild-type, red or green curved lines: oscillation with a phase or amplitude different from wild-type, black straight line: no oscillation. The green curved line indicates an oscillation with an amplitude and/or phase that is both different from wild-type (black) and from the other changed oscillation (red). (F) Model distribution among genes of different functional categories. (G) Tukey boxplots depicting amplitudes of gene expression oscillations in the indicated functional categories. Red stars, significant amplitude change (adjusted <i>p</i>-value = 2.2e-16).</p

Diurnal metabolite patterns show differential dependence on glucocorticoids.

<p>(A, B) Principal component analysis (PCA) score plots of the NMR spectra of untreated samples (A) and of DEX treated together with control samples (B). (C-E) Left: Temporal profiles of metabolite levels determined by UPLC-FLR for the indicated phenotype/treatment combinations. Lines fitted by linear regression if statistical models indicate rhythmicity for wild-type (black) and <i>rx3 strong</i> mutant samples (white). Rhythmic model fits are indicated as continuous lines, non-rhythmic fits as broken lines. Error bars represent SEM. Right: Tukey boxplots depicting metabolite levels at all examined time points in wild-type siblings (WT, white) and <i>rx3 strong</i> mutants (RX3, grey) in the absence or presence of DEX.</p

Glucocorticoid dependent alterations of diurnal TCA cycle function.

<p>(A) Schematic depicting glycolysis, gluconeogenesis, the TCA cycle, and glutaminolysis, indicating the steps where genes with dysregulated temporal mRNA expression in <i>rx3 strong</i> mutants are involved. Gene names are color-coded according to the models in C. Check marks in (A) and (C) indicate rescue of dysregulated expression in <i>rx3 strong</i> by DEX treatment. (B) Tukey boxplots showing fold changes of TCA cycle metabolites when comparing the indicated conditions. Statistically significant changes are shown in green for up-regulation and in red for down-regulation. (C) Heatmap of normalized mRNA expression levels of the genes shown in (A), classified according to their expression models. Red, high expression. Green, low expression. (D, E) Gene expression levels of <i>pck1</i> (D) and <i>gls2</i> (E) mRNA levels in 5 dpf larvae in the indicated conditions. Lines fitted by linear regression if statistical models indicate rhythmicity for wild-type (continuous) and <i>rx3 strong</i> mutant samples (broken).</p

Metabolites quantification and supplementation.

<p>(A) Schematic representation of sulphur assimilation in <i>A</i>. <i>thaliana</i>. Colour squares above the metabolites represent the log2 value of the <i>msa1-1</i>/WT Col-0 ratio of the concentration of each metabolite. APR: APS reductase; APS: adenosine 5’-phosphosulfate; ATPS: ATP sulfurylase; CBL: cystathionine β-lyase; CGS: cystathionine γ-synthase; Cyst: cystathionine; γ-ECS: γ-glutamylcysteine synthetase; γ-GluCys: γ-glutamylcysteine; GSHS: glutathione synthetase; Hcy: homocysteine; MS: methionine synthase; OAS: O-acetylserine; OAS-TL: OAS(thiol)lyase; SAT: serine acetyltransferase; SAMS, S-adenosylmethionine synthetase; SiR: sulphite reductase; SHM: serine hydroxymethyltransferase. (B-G) Measurement of sulphur-related metabolites. Plants were grown on agar solidified MGRL media under S sufficient (S1500) or S deficient (S0) conditions. Metabolites were extracted from shoots and roots and quantified by HPLC. Data are presented as means ± SD (<i>n</i> = 3). *, <i>P</i> ≤ 0.05; **, <i>P</i> ≤ 0.01, Student’s <i>t</i> test. (H-I) The concentrations of SAM and MTA in the shoots and roots of WT Col-0 and <i>msa1-1</i> grown under S sufficient condition. (J) Total S in the shoots of WT Col-0 and <i>msa1-1</i> grown under S sufficient condition without (CK) or with SAM added to the growth medium. Data in (B-J) are presented as means ± SD (<i>n</i> = 3 in (B-G), <i>n</i> = 5 in (H-I), and <i>n</i> = 6 in (J)). * and ** in (B-J) indicate values significantly different between WT Col-0 and <i>msa1-1</i> mutant at <i>P</i> ≤ 0.05 and <i>P</i> ≤ 0.01, respectively (Student’s <i>t</i> test). DW, dry weight. CK, control.</p

Mineral content of Col-0 and <i>fou8</i>.

<p>Col-0 and <i>fou8</i> plants were grown for 30 days in soil in controlled environment room. Whole rosettes were harvested and the mineral levels were determined by X-ray fluorescence spectrophotometry as % of dry weight. Results from one of two independent experiments are presented as means ± SD from three individual plants. Values substantially different between the two genotypes (P<0.05) are marked by asterisks.</p

<i>fou8</i> is affected in glucosinolate synthesis.

<p>Col-0, <i>fou8</i>, <i>apk1 apk2</i>, and <i>fou8 apk1 apk2</i> plants were grown for 5 weeks in controlled environment room. The total content of <b>A</b> glucosinolates and <b>B</b> desulfo-glucosinolates was measured in leaves. <b>C</b> Total RNA was isolated from leaves and the transcript levels of six genes involved in glucosinolate synthesis was determined by quantitative RT-PCR. The qRT-PCR reactions were performed in triplicate for each biological sample. The values in Col-0 were set to 1 for all genes. Results are presented as means ± SE from six pools of three individual plants grown in two independent experiments. Different letters mark values significantly different at P<0.05; asterisks mark values significantly different from Col-0 at P<0.05.</p

Cytosine methylation levels at CG, CHG, and CHH and total cytosine sites in WT and <i>msa1-1</i>.

<p>Cytosine methylation levels at CG, CHG, and CHH and total cytosine sites in WT and <i>msa1-1</i>.</p

Sulfate uptake and flux through sulfate assimilation in <i>fou8</i> and related mutants.

<p>WT Col-0 and mutants <i>fou8</i>, <i>fou2</i>, and <i>aos</i> were grown for 3 weeks on MS-phytagel vertical plates in controlled environment room. The seedlings were incubated for four hours with their roots submerged in nutrient solution adjusted to sulfate concentration of 0.2 mM and supplemented with 6.7 μCi [³⁵S]sulfate. Shoot and root material was harvested separately, and the flux was determined as incorporation of ³⁵S from [³⁵S] sulfate to thiols and proteins. <b>A</b> sulfate uptake, <b>B</b> Percentage of ³⁵S transported to leaves from the [³⁵S]sulfate taken up, <b>C</b> relative flux through the sulfate assimilation in the leaves calculated as % of incorporation in thiols and proteins from total [³⁵S]sulfate taken up. Results are presented as means ± SE from six independent pools of 8 seedlings grown in two independent experiments. Values marked with an asterisk show significant (P≤0.05) difference from Col-0.</p