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Nat Plants
2016 Jan 01;21:15202. doi: 10.1038/nplants.2015.202.
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Inositol transporters AtINT2 and AtINT4 regulate arsenic accumulation in Arabidopsis seeds.
Duan GL
,
Hu Y
,
Schneider S
,
McDermott J
,
Chen J
,
Sauer N
,
Rosen BP
,
Daus B
,
Liu Z
,
Zhu YG
.
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Arsenic contamination of groundwater and soils threatens the health of tens of millions of people worldwide. Understanding the way in which arsenic is taken up by crops such as rice, which serve as a significant source of arsenic in the human diet, is therefore important. Membrane transport proteins that catalyse arsenic uptake by roots, and translocation through the xylem to shoots, have been characterized in a number of plants, including rice. The transporters responsible for loading arsenic from the xylem into the phloem and on into the seeds, however, are yet to be identified. Here, we show that transporters responsible for inositol uptake in the phloem in Arabidopsis also transport arsenic. Transformation of Saccharomyces cerevisiae with AtINT2 or AtINT4 led to increased arsenic accumulation and increased sensitivity to arsenite. Expression of AtINT2 in Xenopus laevis oocytes also induced arsenite import. Disruption of AtINT2 or AtINT4 in Arabidopsis thaliana led to a reduction in phloem, silique and seed arsenic concentrations in plants fed with arsenite through the roots, relative to wild-type plants. These plants also exhibited a large drop in silique and seed arsenic concentrations when fed with arsenite through the leaves. We conclude that in Arabidopsis, inositol transporters are responsible for arsenite loading into the phloem, the key source of arsenic in seeds.
Figure 2. Arsenite transportation with expression of AtINT2 and AtINT4 in yeast and oocytesa: Arsenic concentration in yeast cells D458-1B after grown at 30°C for 24 h in liquid minimal medium supplemented with 2 μg mL−1
myo-inositol and different concentrations of As(III).b: Oocytes from X. laevis were incubated in ND96 complete buffer supplemented with 1 mM of sodium As(III) at room temperature for 30 min. Oocytes injected with water were used as controls.Asterisk indicates significance at P<0.05, and double asterisk indicates significance at P<0.01 compared to controls. Averages and standard errors are shown; n = 4.
Figure 3. Arsenite uptake inhibition by Myo-inositolArsenic concentration in yeast cells D458-1B expressing AtINT2 or AtINT4 and containing empty vector after grown at 30 °C for 24 h in liquid minimal medium supplemented with 250 μM As(III) and the indicated concentration of myo-inositol. Double asterisk indicates significance at P<0.01 compared to cells with vector only. Averages and standard errors are shown; n = 4.
Figure 4. Kinetic properties of AtINT2 and AtINT4 for As(III)Cultures of yeast strain D458-1B expressing either AtINT2 or AtINT4 were incubated in liquid minimal medium supplemented with 2 μg mL−1
myo-inositol and the indicated concentration of As(III) for 30 min. Kinetic data were fitted using a least-squares analysis with SigmaPlot 12.0. Averages and standard errors are shown; n = 4.
Figure 5. Arsenic concentration in phloem exudates, xylem sap and plant tissuesa: Plants were grown in hydroponic solution, after 3 d treatment with 50-μM As(III), rosette leaves were harvested and phloem exudates were rapidly collected by an EDTA-facilitated method.b: Plants were grown in hydroponic solution, one week before harvesting, rosette leaves were brushed with a solution containing 50 μM As(III) and 0.1% Tween using a painting brush. Each plant was brushed with a 2 ml solution daily. After harvesting, the plants were separated into shoots (including upper stem and leaves that had not been brushed) and siliques (including seeds).c: Plants were grown in hydroponic solution, from flowering stage to harvest, As(III) was added to the nutrient solution to a final concentration of 5 μM. After harvesting, plants were separated into roots, shoots, empty siliques and seeds.d: Plants were grown in soil, after 3 d treatment with 5-μM As(III), xylem sap was collected.* indicates significance at P<0.05, and ** indicates significance at P<0.01 compared to WT. Data are shown as average ± SE; n = 4.
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