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doi: 10.1242/10.1242/jcs.00175

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Genome-wide gene expression profiling in Arabidopsis thaliana reveals new targets of abscisic acid and largely impaired gene regulation in the abi1-1 mutant

¹ Laboratory of Plant Molecular Biology, The Rockefeller University, 1230 York Avenue, New York, NY 10021, USA
² E. I. DuPont de Nemours, DuPont Agriculture & Nutrition — Molecular Genetics, PO Box 6104, Newark, Delaware 19714-6104, USA

View larger version (60K): [in a new window]   Fig. 1. ABA-responsive gene expression in Arabidopsis WT. The normalized abundance in ppm of each signature derived from the ABA-treated sample (y axis) was plotted on a logarithmic scale versus that derived from the untreated sample (x axis). The log scaling ends at 1 and signatures with abundance 0 are presented in the lower left corner. For each abundance (ppm) value, points are not placed on a line but allocated to a bar of a certain width that decreases with increasing abundance. Thus, data points are scattered in bi-dimensional space to better reflect density. This translates into a wide gap as seen at around 1 ppm because there were few signatures with both 1 or 2 ppm that would normally occupy a certain space either along the x- or y-axis. Up- or downregulated genes, as defined on the basis of a threefold ratio and/or a 0.0005% significance level, are shown in green. Genes unaffected by ABA are shown in red.

View larger version (52K): [in a new window]   Fig. 2. Confirmation of MPSS data using different techniques. (A) Induction or repression of seven genes was verified by RNA gel blot analyses. Total RNAs from an independent ABA (50 µM) treatment were isolated from WT plants after induction for 3 hours and 5 hours (+). Control plants were treated in parallel (-). The RNA gel blot was probed with DNA fragments of the indicated genes. (B) RT-PCR was used to confirm regulation of genes with low transcript abundance. 200-500 ng total RNA of samples from an independent 4 hours ABA (50 µM) treatment was used for each of the indicated RT-PCR reactions. The 25S RNA band stained with ethidium bromide served as a loading control. The names of genes shown in A and B are: KIN2 (At5g15970), putative NPK1-related MAP kinase (At1g05100), putative protein phosphatase 2C (At1g07430), ethylene responsive element binding factor 4 (At3g15210), citrate synthase-like protein (At3g58750), putative RNA-binding protein (At1g09340), MPK6 (At2g43790), ZFP2 (At5g57520), putative bHLH transcription factor (At2g46510), putative homeodomain transcription factor (At2g35940), potassium channel-like protein (At4g18160), and AAO2 (At3g43600).

View larger version (41K): [in a new window]   Fig. 3. Functional classification of ABA-induced genes. According to their known or suggested functions induced (A) and repressed genes (B) were grouped into the nine indicated categories. The segment sizes correspond to the number of genes in each category related to the total number of either up- or downregulated genes.

View larger version (38K): [in a new window]   Fig. 4. ABA-responsive gene expression in abi1-1. The normalized abundances in ppm for the treated abi1-1 sample were plotted against those for the untreated abi1-1 sample. Representation is as described for Fig. 1. Displayed signatures were selected on the basis of either being upregulated (left panel) or downregulated (right panel) in WT following ABA treatment (c.f. Fig. 1, signatures in green above the diagonal and below the diagonal, respectively).

View larger version (42K): [in a new window]   Fig. 5. (A) Kinetics of ABI1 and ABI2 expression. The time-dependent expression of ABI1 and ABI2 in the presence and absence of ABA in WT and abi1-1 was monitored at the indicated time points. The 25S RNA band stained with ethidium bromide served as a loading control of the RNA gels. (B) Subcellular localization of ABI1 and abi1. GFP fluorescence of ABI1-GFP (left), abi1-GFP (middle) and GFP (right) in onion epidermal cells was observed 16 hours after biolistic bombardment. Representative cells are shown. Bars, 50 µm.

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