Figure 3.

Regulation of MFT by ABA Is Mediated by ABI3 and ABI5.

(A) MFT expression determined by quantitative real-time PCR in wild-type (WT) and various abi mutants mock treated or treated with 10 μM ABA. Because abi3-1 seeds germinated around 14 h after stratification, we collected all germinating seeds 12 h after stratification for comparing MFT expression. Error bars denote sd.

(B) Germination phenotype of the wild type, 35S:ABI3-6HA, and 35S:ABI5-6HA treated with 1 or 5 μM ABA. Error bars denote sd.

(C) Expression of ABI3, ABI5, and MFT determined by quantitative real-time PCR in germinating seeds of the wild type, 35S:ABI3-6HA, and 35S:ABI5-6HA mock treated or treated with 1 or 5 μM ABA. All germinating seeds were collected 16 h after stratification. Error bars denote sd.

(D) ChIP enrichment test showing the binding of ABI3-6HA and ABI5-6HA to the MFT promoter. The upstream region and the first intron of MFT are represented by white boxes, while the first exon is represented by a black box. The arrowheads in the top panel indicate the sites containing putative ABREs on the MFT promoter. Hatched boxes represent the DNA fragments amplified in ChIP assays. ChIP assay results of 35S:ABI3-6HA and 35S:ABI5-6HA are shown in the bottom panels. Seeds were sown on Murashige and Skoog (MS) medium supplemented with 10 μM ABA and harvested 16 h after stratification for ChIP assays. Em6, which has been identified as a direct target of ABI5 (Lopez-Molina et al., 2002), is used as a positive control for ABI5-6HA ChIP assay. Significant differences in comparison with the enrichment of a TUB2 fragment are indicated with asterisks (P < 0.05, Student's t test). Error bars denote sd.