Supplemental Figure Legends

Supplemental figure 1. Binding of 3H-(+)-ABA to purified recombinant FCA protein. a, Binding of 3H-(+)-ABA is specific for FCA. Incubation mixtures contained 10 mg of either native FCA, heat-denatured FCA or bovine serum albumin (BSA) plus 50 nM 3H-(+)-ABA and all buffer components as described in methods. b, pH dependency. Assays contained appropriate buffer adjusted to the pH values shown. The 100% binding activity corresponds to approximately 0.52 mol ABA mol-1 protein. Each data point represents triplicate assays (error bars represent SD).

Supplemental figure 2. FCA protein purified on biotin-tagged (+)-ABA column. Commassie blue-stained SDS-PAGE showing the purified recombinant FCA protein. Lane 1, markers; Lane 2-4, first, second and third washes; Lane 5, elution with 3 mM (-)-ABA; Lane 6, elution with 5 mM (±)-ABA. The column’s matrix of Streptavidin linked to Sepharose had been modified by binding biotin-tagged (+)-ABA to the Streptavidin. The cleared lysate (approximately 11 mg protein in 2 ml lysis buffer) was passed through the column and the column washed, eluted first with (-)-ABA, then with (±)-ABA. All samples (Lane 2 - 6) were loaded at 5 ml from 1 ml concentrated volumes.

Supplemental figure 3. ABA binding disrupts FCA/FY interaction. GST:FCA-WW-FY interaction mixture was incubated for 90 min (0 time in the figure) before 1 mM 3H-(+)-ABA was added and the mixture pelleted at either 15 or 45 min after 3H-(+)-ABA addition. Dual activities of [35S]met-FY and 3H-(+)-ABA for the mixture were measured on a scintillation counter. The 100% binding activity represents the [35S]met-FY (approx. 2.7 x 103 DPM) or 3H-(+)-ABA (approx. 560 DPM, not shown) activity bound to FCA in the absence of the other binding component. Each data point represents triplicate assays (error bars represent SD).

Supplemental figure 4. ABA affects FCA autoregulation. Quantitative GUS activity in cell-free extracts from transgenic plants 2-3 and 4-6 day-old in the absence and presence of 10 mM ABA. Quantitative GUS activity assays were determined in cell-free extracts using 4-methylumbellyferyl β-glucuronide (4-MUG) as a substrate (see methods). Each data point represents triplicate assays (error bars represent SD).

Supplemental figure 5. ABA delays flowering. The plant on the left is untreated, while that on the right was treated every other day with 10 mM ABA, starting four days after germination for a period of 26 days.