Article Figures & Data
Figures
- Fig. 1.
Single and combined effects of VP1 and abi1-1 overexpression on HVA1 promoter-β-glucuronidase (GUS) expression in transiently transformed rice protoplasts. The non-ABA-inducible ubiquitin promoter-luciferase cDNA reporter construct (Christensen and Quail, 1996) was cotransformed as an internal control for transcription activity. Treatments were performed in triplicate; variance bars are ±se.
- Fig. 2.
Specific synergistic effect of lanthanide ions on the ABA-inducible Em promoter, but not the non-ABA-inducible rice actin (Ra) promoter. Numbers in parentheses show fold induction over untreated controls. A, Effects of lanthanum with or without ABA cotreatment on the actin and Em promoters, measured by GUS/LUC reporter enzyme assays. B, Effects of terbium on the actin and Em promoters, cotransformed and measured in the same samples by reporter enzyme activities and GFP flow cytometry, respectively. Results are the average of three to nine replicates (±se). a, Significantly higher than control treatment (paired t test,P < 0.02). b, Significantly higher than ABA (P < 0.05).
- Fig. 3.
Scatter plot (cell size versus GFP fluorescence) of Em-GFP-transformed protoplasts treated for 20 h with 10 μm ABA (A), 5 mm LaCl3 (B), or both ABA plus LaCl3 (C). Ten-thousand protoplasts were measured by flow cytometry, and those with fluorescence above an arbitrary background (horizontal line) were gated for quantitation of fluorescence intensity.
- Fig. 4.
Overexpression of abi1-1 inhibits lanthanum-induced Em expression to a similar extent as ABA-induced Em expression. Rice protoplasts were transformed with Em-GFP in the presence of overexpressed abi1-1 (pG2) or a nullABI1 control (pG1; Sheen, 1998). Numbers above theabi1-1-treated samples (white bars) indicate the percentageEm expression relative to control. Transformations were performed in triplicate and flow cytometry measurements in duplicate; variance bars are ±se.
Tables
- Table I.
Specificity of the lanthanide effect on the ABA-inducible Em and HVA1 promoters in transiently transformed protoplasts
Treatments Promoter Fold-Induction (±se) ABA Salt 35S Em HVA1 μm mm 0 0 1.0 1.0 1.0 100 0 0.8 (0.1) 16.6 (2.0) 13.7 (1.0) 0 1 LaCl3 1.1 (0.0) 2.2 (0.6) 3.21-a (0.5) 0 2 TbCl3 1.1 (0.1) 4.61-a (0.2) n/a1-c 0 5 LaCl3 1.4 (0.3) 2.7 (0.8) 3.81-a (0.4) 100 1 LaCl3 1.1 (0.1) 32.61-b (2.9) 29.61-b (5.6) 100 2 TbCl3 1.2 (0.1) 31.91-b (1.5) n/a1-c 100 5 LaCl3 1.1 (0.4) 38.81-b (9.9) 24.1 (5.5) Induction was calculated as the average ratio of relative reporter gene activities (promoter-GUS/Ubi-LUC) between treatment and control (set to unity) from three- or four-paired replicates in two independent experiments.
- Table II.
ABA and lanthanide salt effects on viability of transformed protoplasts after an 18- to 20-h treatment
Treatment Viability Percentage (±se) ABA2-a Salt2-b Experiment I Experiment II Experiment III μm mm 0 0 36.2 (6.7) 15.0 (0.6) 45.4 (0.6) + 0 31.7 (4.0) 15.5 (0.7) 43.8 (0.6) 0 1 14.9 (1.2) 15.5 (0.6) 49.6 (1.7) + 1 9.4 (4.6) 17.7 (0.7) 48.5 (0.7) 0 5 12.8 (11.0) 17.4 (0.3) 48.5 (1.4) + 5 3.4 (1.8) 16.5 (0.4) 47.5 (1.3) Samples were assayed by flow cytometry of protoplasts treated with 0.01% (w/v) fluorescein diacetate for 5 min.
-
↵F2-a Ten micromolar ABA in experiments I and II; 100 μm ABA in experiment III.
-
↵F2-b Experiment I, LaCl3 (two independent transformations); experiment II, TbCl3(five–nine independent transformations); experiment III, average of parallel LaCl3 and TbCl3 treatments (three–nine independent transformations).
-
- Table III.
Trivalent, but not divalent, ions stimulate Em promoter activity in transiently transformed protoplasts
Treatment Em Promoter Fold-Induction (±se) PValue3-a Concentration Salt mm 5.0 LiCl 0.9 (0.1) 0.29 5.0 MgCl2 1.0 (0.1) 0.44 5.0 MgSO4 1.0 (0.1) 0.48 10.0 MnCl2 1.0 (0.3) 0.44 0.13 AlCl3 3.7 (0.6) 0.03 0.13 LaCl3 2.4 (0.4) 0.05 0.13 TbCl3 1.6 (0.2) 0.09 Induction was calculated as the average ratio of relative reporter gene activities (Em-GUS/Ubi-LUC) between treatment and control (set to unity) from three paired replicates for the LiCl, MgCl2, MgSO4, and MnCl2 treatments. For the AlCl3, LaCl3, and TbCl3treatments, induction was calculated as the average ratio of relative weighted Em-GFP expression between treatment and control from three paired replicates.
-
↵F3-a One-sided paired t test.
-
