Modification of nitrate uptake pathway in plants affects the cadmium uptake by roots

Abstract

NRT1.1 is a dual-affinity nitrate (NO₃⁻) transporter involved in both high- and low-affinity NO₃⁻ uptake in Arabidopsis plants. In a recent study, we showed that, under cadmium (Cd) exposure, blocking the NRT1.1-mediated NO₃⁻ uptake reduces Cd entry into roots, thus lowing Cd levels in plants and improving plant growth. In addition, we also found that the Cd levels in edible parts of 11 Chinese cabbage (Brassica rapa L. ssp. pekinensis) cultivars correlated well with the NO₃⁻ uptake rates of their roots. These results suggested that the NO₃⁻ uptake of roots negatively regulate Cd uptake. Modification of NO₃⁻ uptake in crops by modulating NO₃⁻ uptake pathway might provide a biological engineering approach to reducing Cd accumulation in edible organs, thus improving food safety.

Keywords:

• cadmium
• nitrate uptake
• nitrate transporter
• nitrogen form

Cadmium (Cd) is highly toxic to humans. A survey showed that vegetables and rice account for almost 80% of total Cd exposure in residents of Shanghai, China's largest city.¹ Several strategies have been developed to minimize Cd uptake by crops from Cd concentration soil. These strategies mainly include: (1) ‘dig-and-dump’ or encapsulation of the contaminated soil, (2) chemical immobilization or extraction of Cd, and (3) phytoremediation by Cd-hyperaccumulating plants.² In addition, researchers also throw the lights to agronomic measures to minimize the Cd entry into crops, as several of the plant nutrients have many effects on Cd's availability and Cd uptake,³ such as phosphate precipitating Cd²⁺ in soil,⁴ Fe²⁺ competing with Cd²⁺ for IRON-REGULATED TRANSPORTER 1(IRT1).^5,6 Recently, we found that, in the hydroponical growth condition, the NO₃⁻ treatment resulted in higher Cd concentration and less Cd tolerance in tomato plants (Solanum lycopersicum) compared with ammonium (NH₄⁺) treatment.⁷ A similar result was also observed in hydroponically grown Arabidopsis (Arabidopsis thaliana) plants (Fig. 1). Therefore, appropriate use of nitrogen (N) fertilizers might provide a relatively inexpensive, time-saving, and effective strategy for reducing Cd entry into, and accumulation in crop. However, in another study, we found that the actual effect of the N form on Cd accumulation of the plants grown with soil was strongly associated with the pH-buffering capacity of the soil. In soil with higher pH-buffering capacity, the effect of N-form on Cd accumulation in plants was similar to that observed in the hydroponically grown plants, but in soil with lower pH-buffering capacity, application of NH₄⁺ resulted in higher Cd levels in plants than application of NO₃⁻, probably as a result of soil acidification by rapid nitrification of NH₄⁺ (S.K. Fan, S.T. Du, C.W. Jin; unpublished data). This finding suggests that it could pose higher risk and difficulty to employ N-fertilization management to prevent Cd entry into crops since the pH-buffering capacity of soil varies greatly in different regions.

Figure 1. Effect of N-form on growth and Cd concentration in Arabidopsis Col-0 plants grown with solution medium. Effect of N-form on growth and Cd concentration in Arabidopsis Col-0 plants grown with solution medium. (A) and (B) The relative biomass. (C) and (D) The Cd concentration in plants from Cd treatment. At the age of 5 weeks, the plant weere transferred to Cd-free or 10 μM Cd-added growth solutions with either NO₃⁻ or NH₄⁺ as the sole nitrogen source. The shoots and roots of plants after 7 d of treatments were harvested for biomass and Cd concentration measurements. The relative biomass was calculated as the mean of fresh weight relative to the control treatment. Error bars represent standard deviations (n = 5–8). n refers to number of biological replicates

Most recently, we found that, in Arabidopsis plants, exposure of Cd reduces NO₃⁻ uptake of roots by inhibiting the activity of NITRATE TRANSPORTER 1.1 (NRT1.1), but this process, in turn, reduces Cd entry into the root cells. As a result, Cd level in plants is lower and plant growth is improved.⁸ Therefore, inhibition of NRT1.1-mediated NO₃⁻ uptake could be recognized as a mechanism of plants for resisting Cd toxicity. This mechanism may provide a new strategy for minimizing Cd accumulation in crops grown in contaminated soil using biotechnological pathways to decrease NO₃⁻ uptake. Interestingly, in 11 Chinese cabbage (Brassica rapa L. ssp. pekinensis) cultivars, we found that their Cd levels in edible parts (shoots) correlated well with the NO₃⁻ uptake rates of roots (Fig. 2). The result indicates that, in addition to engineering or breeding of crops, screening the cultivar with low NO₃⁻ uptake rate in its roots may be also an available strategy for decreasing Cd accumulation in crops.

Figure 2. Correlation between NO₃⁻ uptake rate and Cd concentration in edible parts in 11 Chinese cabbage cultivars. For measurement of NO₃⁻ uptake rate, roots of hydroponically grown plants were immersed in 100 mL of a constantly agitated and aerated solution containing 0.5 mM CaSO₄ and 2 mM KNO₃, and the net uptake of NO₃ ⁻ was measured as NO₃ ⁻ depletion from the solution per unit of time. The Cd concentration in edible parts was measured in plants grown with soil containing Cd at rate of 10 mg kg⁻¹ soil.

Because Cd is a non-essential element for plants, Cd²⁺ entry into root cell may rely on the transporters/channels used for acquiring bivalent cation.^9-11 Although knockout of those bivalent nutrient transporters, such as Fe²⁺ transporter AtIRT1 in Arabidopsis and Mn²⁺ transporter osNRAMP5 in rice, could also decrease Cd accumulation in plant tissues, but they often strongly inhibited biomass production, because these transporters are the primary determinants in uptake of relative bivalent nutrients from growth medium.^6,12 Therefore, biotechnological modification of these bivalent nutrient transporters to decrease Cd accumulation in crops seems not to be a wise strategy. However, the N acquisition of plants is controlled by several transporters. For example, in Arabidopsis plant, 5 NRTs and 4 NH₄⁺ transporters (AMTs) have been currently indentified to be involved in N uptake of roots from growth medium.^13-16 This concept allows us to assume that the N acquisition in the plants with inhibited/blocked a certain N uptake process, may be complemented by the other N uptake processes. This is probably the reason why blocking the NRT1.1-controlled NO₃⁻ uptake only slightly affected on the plant growth in the growth condition or our previous study.⁸ Therefore, biotechnological modifying of NO₃⁻ uptake pathway to decrease Cd entry into crop, not only can circumvent both the risk and the difficulty arisen by the N fertilizer management but also may be interfere less with crop production.

Disclosure of Potential Conflicts of Interest

No potential conflicts of interest were disclosed.

Funding

This work was financially supported by the Zhejiang Province Natural Science Foundation (LR13C130001 and LY14C130001), the Natural Science Foundation of China (31270041) and the Fundamental Research Funds for the Central Universities (2014QNA6006).