1. A Peroxidase Contributes to ROS Production during Arabidopsis Root Response to Potassium Deficiency 
Kim Min Jung , Ciani Silvano , Schachtman Daniel P.  
Molecular Plant 
Volume 3, Issue 2, Pages (March 2010)
DOI: /mp/ssp121
Copyright © 2010 The Authors. All rights reserved. Terms and Conditions

2. Figure 1
Overexpression of the RCI3 Gene Activates the AtHAK5prom–LUC Reporter.
(A) The 8–1–24 mutant plants emit strong luminescence under K+-sufficient conditions. # 8–1–24, plants transformed with AtHAK5prom–LUC and tagged by pDKS2-7 activation vector. Negative control (NC), plants transformed with AtHAK5prom–LUC. Positive control (PC), plants transformed with 35Sprom–LUC (i.e. LUC under the control of 35S promoter). Left, bright-field image of all plants; right, luminescence image of the plants. The color scale on the right shows the luminescence intensity from dark blue (lowest) to white (highest).
(B) The T-DNA inserted upstream of RCI3 (At1g05260) gene in the 8–1–24 mutant. The GenomeWalker system was used to identify the T-DNA insertion site with left border-specific primers. The distance from the T-DNA insertion site to each gene is indicated in kilobases.
(C) RCI3 is overexpressed in the 8–1–24 mutant. Expression levels relative to the wild-type of two genes, At1g05260 and At1g05230, were obtained by quantitative real-time PCR in wild-type (WT), NC, and 8–1–24 mutant. Data represent the mean of three biological replicates.
Molecular Plant 2010 3, DOI: ( /mp/ssp121)
Copyright © 2010 The Authors. All rights reserved. Terms and Conditions

3. Figure 2
ROS Production Is Induced in Plants Overexpressing RCI3 under Normal Conditions.
(A) RT–PCR analysis of RCI3 expression in wild-type, four RCI3-ox, and rci3-1 plants.
(B) ROS fluorescence images are shown in wild-type, RCI3-ox, and rci3-1 roots grown on K+-sufficient (+ K, 1.75 mM K+) or K+-deprived (–K, 0 mM K+) medium. ROS was visualized by staining 3-day-old roots with 20 μM DFFDA. Yellow and red colors indicate higher ROS production as indicated in pseudocolor scale. Inset shows enlargement of RHDZ. White lines show the boundary of roots. Bar = 100 μm.
(C) Quantified data from images shown in (B). Roots were deprived K+ for 24 h. Data represent the mean ± standard deviation. Asterisks indicate a significant difference; n = 10 plants).
(D) Three-day-old RCI3-ox seedlings were incubated with NADPH oxidase inhibitor (DPI) for 1 h under K+-sufficient (+K, 1.75 mM K+) conditions.
Molecular Plant 2010 3, DOI: ( /mp/ssp121)
Copyright © 2010 The Authors. All rights reserved. Terms and Conditions

4. Figure 3
RCI3–YFP Protein Is Localized to ER and then Secreted to the Cell Wall.
(A) Sub-cellular localization of YFP control, RCI3–YFP, and ER–CFP in Arabidopsis protoplast. Signals were detected 24 h after incubation from bright-field (bright), YFP, or CFP.
(B) Sub-cellular co-localization of RCI3–YFP with ER–CFP in Arabidopsis protoplast. Expression of the genes was examined at 24 h after transformation. Signals from bright-field (bright), YFP (YFP; green), CFP (CFP: red), and merge (co-localization appears yellow).
(C) Fluorescence proteins were transiently expressed in onion epidermal cells: YFP (top) and RCI3–YFP (bottom). Signals were detected 48 h after incubation from bright-field (bright), YFP (YFP), and merged images.
(D) The localization of RCI3–YFP in plasmolyzed onion cells: YFP (top) and RCI3–YFP (bottom). Signals were detected 48 h after incubation from bright-field (bright), YFP (YFP), and merged images. White lines show the boundary of plasma membrane of the cell.
Molecular Plant 2010 3, DOI: ( /mp/ssp121)
Copyright © 2010 The Authors. All rights reserved. Terms and Conditions

5. Figure 4
RCI3 Expression Is Induced by K+ Deprivation and Affects AtHAK5 Expression.
(A) Quantitative real-time PCR analysis of RCI3 under nutrient-deprived conditions. Arabidopsis Col-0 roots were grown under N (–N), P (–P), or K+ deficiency (–K) for 6 or 30 h. Asterisks indicate a significant difference as compared to other means in each subfigure (* p < 0.05; Student's t-test).
(B) AtHAK5 expression in wild-type, RCI3-ox, and rci3-1 knockout plants under K+-sufficient or K+-deprived conditions. RNA was extracted from whole plants. Data represent the mean of three biological replicates. Different letters indicate a significant difference between means at p < 0.05 (TUKEY HSD).
Molecular Plant 2010 3, DOI: ( /mp/ssp121)
Copyright © 2010 The Authors. All rights reserved. Terms and Conditions

6. Figure 5
Altered RCI3 Expression Alters Primary Root Growth in K+-Sufficient but Not K+-Deprived Conditions.
(A, B) Plants were grown under complete nutrient conditions for 4 d and then transferred to either full nutrient (A, +K) or a medium with no potassium (B, –K) for 7 d.
(C) Primary root length of plants shown in (A) and (B) was measured. Data represent the mean ± standard deviation. Different letters indicate a significant difference between means at p < 0.05 (TUKEY HSD) (n = 70 plants).
(D) The primary root-length as A percentage of full nutrient medium to medium without potassium. Data represent the mean ± standard deviation. Different letters indicate a significant difference between means at p < 0.05 (TUKEY HSD) (n = 70 plants).
Molecular Plant 2010 3, DOI: ( /mp/ssp121)
Copyright © 2010 The Authors. All rights reserved. Terms and Conditions