Fig. 7.
Overexpression of CAR1-DADA leads to a dominant-negative phenotype with respect to ABA sensitivity. (A) CAR1-DADA OE lines show reduced sensitivity to ABA compared with Col WT (Upper Left). Expression of HA-tagged CAR1-DADA was verified by immunoblot analysis of the transgenic lines (Upper Right). Photographs are shown of representative seedlings from Col WT and two CAR1-DADA OE lines grown for 12 d on MS medium either lacking or supplemented with 0.5 μM ABA. Seedlings were rearranged on agar plates, and longitudinal or zenital photographs were taken to measure root length or the maximum rosette radius. The histograms show the quantification of root length (Lower Left) or the maximum rosette radius (Lower Right). (B) Quantification of ABA-mediated inhibition of seedling establishment of Col WT compared with hab1-1abi1-2 double mutant, CAR1 OE lines, and HAB1 OE and CAR1D22 D27A OE lines. Approximately 100 seeds of each genotype were sown on MS plates lacking or supplemented with either 0.5 or 1 mM ABA and scored for the presence of green expanded cotyledons 7 d later. *P < 0.05 (Student’s t test) when comparing data of each genetic background to Col WT plants in the same assay conditions. (C) The CAR1-DADA mutation abolishes the interaction of CAR1 with PYR1 in the plasma membrane. Confocal images of transiently transformed tobacco epidermal cells coexpressing the indicated constructs and increasing amounts of CAR1-DADA. The ratio of the relative concentration of agrobacteria in the different coinfiltrations is indicated by numbers (1×, 2×, or 4×).