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Supplemental Figure 1. The cell death phenotype of fhy3 far1 double mutants. A. The cell death phenotype of fhy3-4 far1-2 mutant plants under LD conditions.

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Presentation on theme: "Supplemental Figure 1. The cell death phenotype of fhy3 far1 double mutants. A. The cell death phenotype of fhy3-4 far1-2 mutant plants under LD conditions."— Presentation transcript:

1 Supplemental Figure 1. The cell death phenotype of fhy3 far1 double mutants. A. The cell death phenotype of fhy3-4 far1-2 mutant plants under LD conditions. The leaf rugosity and patches of cell death are presented in fhy3 far1 mutants while not in No-0 wild-type plants. Bars, 5 mm. B. Morphological phenotypes of wild-type No-0, fhy3-4, far1-2, fhy3-4 far1-2 and fhy3-1 far1-2 mutant plants under SD conditions. Photographs were taken when plants were 14, 18, 21 and 48 days after seed germination in soil. Bars, 5 mm in the upper three panels and 1 cm in the lower panel. C. Induced expression of FHY3-GR in fhy3-4 far1-2 mutant plants background rescued the cell death phenotype in the young leaves. Left panel, mock treatment; right panel, 5  M DEX treatment for 1 week under SD conditions. 4-week-old plants grown under SD conditions were used for the DEX treatment. Red arrows indicate the patches of cell death; leaves labeled with 1-4 indicate areas where no severe cell death occurred on the newly generated leaves of fhy3 far1 plants after DEX treatment. Bars, 1 cm. D. Night break treatment largely suppress darkness induced severe cell death and stunted growth of fhy3-4 far1-2 under SD conditions. One-hour light were given in the middle of the night at ZT16 (ZT0 is the start of light phase, and ZT8 is the beginning of the dark phase) when No-0, fhy3-4 and fhy3-4 far1-2 were grown under SD conditions. Photographs were taken when plants were four-week old. Right panel, without night break control. Arrows indicates the leaves with severe cell death. Bars, 5 mm. Supplemental Figures

2 Supplemental Figure 2. Expression of oxidative stress-related genes in fhy3 far1 mutants using an RT-PCR assay. Gene names in red indicate the genes with altered mRNA expression levels in fhy3-4 far1-2 mutants, compared with No-0 wild type. RNA from three-week-old plants grown in SD conditions was used to perform RT-PCR assays.

3 Supplemental Figure 3. Microarray expression profiling of fhy3 far1 mutants. A-D. Heat map analysis showing the altered gene expression in fhy3 far1 mutants. E-F. Expression of genes related to plant defense or SA (E), and reactive oxygen species or oxidative stress (F) are highly expressed in the fhy3 far1 mutant. The abundance of the constitutively expressed histone 3 (H3.3, At4g40040) and ribosomal protein S5 (At2g09990) are used as negative controls in E.

4 Supplemental Figure 4. FHY3 and FAR1 regulate the transcript levels of MIPS2. A. RT-qPCR measurement of transcript levels of MIPS2 after transfer of plants from dark to white light. Left, the expression of MIPS2 is induced by light in Col, Ler, and C24 ecotypes, but not in the No-0 ecotype. Right, the basal transcript levels of MIPS2 decreased in the fhy3, far1, and fhy3 far1 mutants, compared with wild type (No-0). B. RT-qPCR measurement of transcript levels of MIPS2 after transfer of plants from dark to FR (left), R (middle), and B (right) light conditions. High expression of MIPS2 induced by red (R, middle), or blue (B, right) light decreased in the fhy3 and fhy3 far1 mutants, compared with wild type (No-0). R0-R3, F0-F3, and B0-B3 indicate red, far-red, and blue light treatment for 0, 1 or 3 hours, respectively. Data are shown as mean ± SD; n = 3.

5 fhy3 far1 MIPS1-OE No-0 L27 L45  -FLAG  -Actin Supplemental Figure 5. FHY3 binds directly to the MIPS1/2 promoter region. A. ChIP-PCR showing that FHY3 binds to the promoter regions (P, promoter; E, exon) of MIPS1/2, but not MIPS3. 7-day-old 3FLAG-FHY3-3HA/fhy3 transgenic seedlings grown under LD conditions (ZT4) were used to perform ChIP-PCR assays. B. The expression of MIPS1/2 under long-day (LD), short day (SD), and continuous darkness (DD_DDHC) conditions. The expression data for MIPS1/2 were extracted from Diurnal (http://diurnal.mocklerlab.org).http://diurnal.mocklerlab.org Supplemental Figure 6. Western blots showing MIPS1-FLAG protein levels in MIPS1-OE transgenic plants. Seven-day-old No-0, fhy3-4 far1-2, and MIPS1-OE/fhy3-4 far1-2 seedlings grown under LD conditions were used to detect the protein levels of MIPS1-FLAG using anti-FLAG antibodies (sigma). Anti-Actin was used as a sample loading control.


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