Comparative effectiveness of Se translocation between low-Se and high-Se rice cultivars under Se fertilization

Comparative effectiveness of Se translocation between low-Se and high-Se rice cultivars under Se fertilization

Ecotoxicology and Environmental Safety 205 (2020) 111372 Contents lists available at ScienceDirect Ecotoxicology and Environmental Safety journal ho...

1MB Sizes 0 Downloads 9 Views

Ecotoxicology and Environmental Safety 205 (2020) 111372

Contents lists available at ScienceDirect

Ecotoxicology and Environmental Safety journal homepage: www.elsevier.com/locate/ecoenv

Comparative effectiveness of Se translocation between low-Se and high-Se rice cultivars under Se fertilization Mu Zhang a, b, c, Yuwan Pang a, b, c, Qiong Yi a, b, c, Jianfeng Huang a, b, c, Xu Huang a, b, c, Qiaoyi Huang a, b, c, Peizhi Xu a, b, c, *, Shuanhu Tang a, b, c, ** a b c

Institute of Agricultural Resources and Environment, Guangdong Academy of Agricultural Sciences, Guangzhou, 510640, China Key Laboratory of Plant Nutrition and Fertiliser in South Region, Ministry of Agriculture, Guangzhou, 510640, China Guangdong Key Laboratory of Nutrient Cycling and Farmland Conservation, Guangzhou, 510640, China

A R T I C L E I N F O

A B S T R A C T

Keywords: Selenium Reutilization Distribution Transporter Rice (Oryza sativa L.)

The production of natural selenium (Se)-rich food by using a high-Se crop cultivar is beneficial to human health and environmental safety; however, the underlying mechanism of different Se-accumulation ability between high- and low-Se rice cultivars remains unclear. A low-grain-Se cultivar and high-grain-Se cultivar of rice were used as test materials, and two levels of Se (0 and 0.5 mg kg− 1) were arranged in a randomized design containing twelve replicates. The dynamic changes of shoot Se concentration and accumulation, xylem sap Se concentration, shoot and grain Se distribution, Se transporters genes (OsPT2, Sultr1;2, NRT1.1B) expression of the high- and lowSe rice cultivars were determined. The shoot Se concentration and accumulation of the high-Se rice showed a greater degree of reduction than those of the low-Se rice during grain filling stage, indicating that leaves of highSe rice served as a Se source and supplied more Se for the growth centre grain. The expression levels of OsPT2, NRT1.1B and Sultr1;2 in the high-Se rice cultivar were significantly higher than those in the low-Se rice cultivar, which indicated that the high-Se rice cultivar possessed better transport carriers. The distribution of Se in grain of the high-Se rice cultivar was more uniform, whereas the low-Se cultivar tended to accumulate Se in embryo end. The stronger reutilization of Se from shoots to grains promoted by increased transporters genes expression and optimized grain storage space may explain how the high-Se rice cultivar is able to accumulate more Se in grain.

1. Introduction

underlying Se accumulation in high- and low-efficiency rice cultivars is not yet well understood. Previous studies were conducted to examine the differential char­ acteristics of grain Se accumulation between high- and low-Se rice cul­ tivars. Du et al. (2009) compared the differences in the Se enrichment abilities of more than 60 rice cultivars and found that the Se concen­ tration of rice grains with high Se accumulation reached up to three times that of cultivars with low Se accumulation. Zhang et al. (2019a) found that the high-Se rice cultivar increased Se availability in the rhizosphere soil through root activities, thereby enhancing the absorp­ tion of Se. In addition to the difference in Se absorption, rice species also differ in the ability to translocate Se within plants. Zhou et al. (2014) suggested that the root xylem of a high-Se rice cultivar exhibited a greater Se carrying capacity, and that high-Se rice can transport more Se to the shoots at the same transpiration rate. Additionally, a study by Zhang et al. (2006) demonstrated that the shoots of rice served as source

Selenium (Se) is a beneficial element for plants, but the gap between beneficial and toxic levels of Se in plants is narrow (Silva et al., 2018). Excessive application of Se fertilizer may adversely affect the environ­ ment and threaten food safety, thus improving the ability of crops to obtain natural Se in soil is a very safe way to produce Se-rich food (Quinn et al., 2017). Different types of crops have different abilities to accumulate Se, and even different genotypes of the same crop show high and low Se efficiencies, e.g., significant differences are observed in Se absorption and accumulation by different rice cultivars (Zhou et al., 2016). The enhancement of Se in rice grain has been conducted since the late 1980s (Yoshida and Yasumoto, 1987), and that high-Se rice culti­ vars are often used as Se-enriching carriers to produce Se-enriched rice. Although cultivar screening can be applied to obtain high-Se rice cul­ tivars to produce Se-enriched rice, the differential mechanism

* Corresponding author. Institute of Agricultural Resources and Environment, Guangdong Academy of Agricultural Sciences, Guangzhou, 510640, China. ** Corresponding author. Institute of Agricultural Resources and Environment, Guangdong Academy of Agricultural Sciences, Guangzhou, 510640, China. E-mail addresses: [email protected] (P. Xu), [email protected] (S. Tang). https://doi.org/10.1016/j.ecoenv.2020.111372 Received 18 July 2020; Received in revised form 12 September 2020; Accepted 14 September 2020 Available online 22 September 2020 0147-6513/© 2020 Elsevier Inc. All rights reserved.

M. Zhang et al.

Ecotoxicology and Environmental Safety 205 (2020) 111372

for storing Se, and the concentration of Se in grain was positively correlated with the biomass of shoot. Thus it can be seen that the loading of Se in grains of rice is related not only to root absorption process but also to the distribution of Se within tissues. According to previous studies, the Se concentration in the roots and shoot occurred in an ascending order for the Se hyperaccumulator Astragalus bisulcatus and in a descending order for the nonhyperaccumulator Hordeum vulgare (Barillas et al., 2012; Ilbas et al., 2012). Rice is not within the range of hyperaccumulator plants (Li et al., 2018), and the ability to transfer Se to aboveground tissue is limited. In term of Se application in rice for enhanced edible Se, the accumulation of Se in grain is the ultimate goal. The shoot serves as a “source” and the grain serves as a “reservoir” in rice, and reusable elements are contin­ uously transported from shoot to grain, especially in the late stage of growth. The reusable element is highly mobile in plant tissues and can migrate from the original accumulation site to newly growing organs (Yang et al., 2015; Mitani-Ueno et al., 2018). The growth centre of rice changes continuously at different growth stages, and the original growth centre will be replaced by a new growth centre. Se, as a non-essential nutrient element, can also be reused and redistributed to grain, which is the growth centre during the filling stage (Zhou et al., 2016). There­ fore, studying the difference in Se reutilization between high- and low-Se rice cultivars is of great importance to better understand the addition of Se to grain. Se in the functional leaves of rice is first transported to the apoplast, and then loaded into the phloem sieve tubes via active transport before being transported to the stem base, after which it enters the xylem vessels via transport cells and is transported near the grains (Taiz and Zeiger, 2010; Pilon-Smits, 2017). Earlier literature suggested that sele­ nite is transported by the phosphate transporter OsPT2 (Zhang et al., 2020) and selenate is transported by the sulphate transporters Sutr1;2 (Maruyama-Nakashita, 2016; Mehdawi et al., 2018). Furthermore, Zhang et al. (2019b) showed that a large proportion of the selenite absorbed by plants was transformed into organic Se compounds, and selenomethinone (SeMet) was the dominant Se form in rice tissue. The same study also demonstrated that the peptide transporter gene NRT1.1B overexpression significantly improved Se concentrations in rice grains by facilitating SeMet translocation. Studies are necessary to examine whether there are differences in the expression levels of Se transporters genes between high- and low-Se rice cultivars during grain filing stage as well as the effects on Se accumulation in grains. Grains serve as reserve for storing mineral elements, and the storage capacity of grains plays an important role in nutrients accumulation. Se in grains is distributed in the seed coat, embryo, and endosperm (Carey et al., 2012). The endosperm is the main structure in which nutrients are stored in rice. Hence, the Se concentration in the endosperm is the greatest contributor to the Se concentration of the entire grain, while the uniform distribution of Se in the endosperm indicates where the rice can fully utilize the storage capacity of the grains. Se localization and speciation characterization of rice grain were analysed by using X-ray fluorescence and X-ray absorption near edge structure techniques, and results revealed that Se concentration and speciation at different points were both different (Amos et al., 2012). It is necessary to investigate whether there is a difference in utilization of grain storage space be­ tween high- and low-Se rice cultivars, which may lead to different grain Se accumulation. In this study, we compared the migration process of Se from shoots to grains, and the distribution of Se in grains of high- and low-Se rice cultivars supplied with Se. We hypothesized that the difference in the accumulation of Se in grains between the high- and low-Se rice cultivars was due to the different redistribution of tissue Se in tissues and the utilization of grain storage space. Thus, a pot experiment with Se application was conducted, and the objectives of this study were to (1) investigate the dynamic changes of shoot Se concentration and accu­ mulation of the high- and low-Se rice cultivars during grain filling stage, (2) determine the transporters genes (OsPT2, Sultr1;2, NRT1.1B)

expression in shoots, (3) analyse the xylem sap Se concentration, and (4) evaluate the distribution of Se in grains. 2. Materials and methods 2.1. Materials and experimental treatments A low-grain-Se cultivar HFZ (L) and high-grain-Se cultivar FBZ (H) of rice (Oryza sativa L.) were used as test materials. Rice seeds of the two cultivars are bred by the Rice Research Institute of Guangdong Academy of Agricultural Sciences, China. A carefully managed pot experiment was conducted in the greenhouse of the Guangdong Academy of Agri­ cultural Sciences, China (23◦ 7′ N, 113◦ 15′ E) from 12 March to 25 July 2019. The meteorological conditions of the greenhouse were natural light and temperature. The basic agrochemical characteristics of the experimental soil were as follows (Table S1): clay soil, pH 5.0 (soil/ water ratio of 1:5), organic matter 41.1 g kg− 1, total Se 0.34 mg kg− 1, alkaline hydrolysable nitrogen 168.6 mg kg− 1, Olsen extractable phos­ phate 18.7 mg kg− 1 and exchangeable potassium 148.5 mg kg− 1. The soil was air-dried, ground and sieved through a 2 mm sieve, and the length, width and height of the experimental cuboid pot were 34 cm, 27 cm, and 15 cm, respectively. Each pot was filled with 6.0 kg of the soil and fertilized with the following macroelements (g kg− 1 soil): N 0.2, P2O5 0.15 and K2O 0.2 supplied as urea, ammonium phosphate and potassium chloride, respectively. The microelements supplied were as follows: MnCl2⋅4H2O 1.81 mg kg− 1, CuSO4⋅5H2O 0.08 mg kg− 1, ZnSO4⋅7H2O 0.22 mgkg− 1 and H3BO3 2.86 mgkg− 1. Treatments included two concentrations of Se (0 and 0.5 mg kg− 1) applied as sele­ nite (Na2SeO3), and each treatment contained 12 replicates. The fertil­ izers were dissolved into water and then irrigated to the soil before planting, and seedlings were thinned to 4 per pot 7 days after sowing. Experiment management was carried out daily in the greenhouse during rice growth period. Deionized water was used to irrigate plants during growth period, and make each pot hold the same amount of water after irrigation. The chlorpyrifos and avermectin were used for pest control. 2.2. Measurement of soil chemical properties The basic agrochemical characteristics of soil were analysed by the methods of Bao (2002). The pH was measured by a PB-10 pH meter (Sartorius, Germany). The organic matter was tested based on the method of volumetric with the potassium dichromate. Alkaline hydro­ lysable N was determined based on the method of alkali dissociation diffusion. Olsen-P was determined according to the method of colori­ metric with the molybdenum-antimony. Available K was tested by using a FP6410 flame photometer (Xingyi, China). For soil sample Se analysis, about 1.0 g of soil was digested with 6 mL ultrapurenitric acid and 2 mL perchloric acid at 160 ◦ C. After the soil turned gray, 10 mL of 6 M hy­ drochloric acid was then added, and the acid mixture was heated at 100 ◦ C for 10 min. The solution Se concentration was analysed by using an 8200 Atomic Fluorescence Spectrometer (Jitian, China) (NY/T 1101, 2006). The certified reference material (Soil, GBW07408/GSS-8) was used as the quality control sample. 2.3. Measurement of Se and other elements contents in shoots Rice samples for each treatment from four replicates were collected at early filling (10 June 2019) and maturation stages (25 July 2019), separated into leaf, stem and grain (husk included), washed three times with deionized water and then oven-dried at 40 ◦ C. The fresh and dry weights of leaves, stems and grains of the two rice cultivars were accurately recorded by analytical balance. The oven-dried plant mate­ rial was ground and filtered using a 1-mm nylon sieve. About 0.2 g of plant material was digested in pressurised perfluoroalkoxy vessel with 1 mL of ultra-pure water, 2 mL of ultra-pure nitric acid and 1 mL of hydrogen peroxide by microwave heating (Multiwave 3000, Anton Paar, 2

M. Zhang et al.

Ecotoxicology and Environmental Safety 205 (2020) 111372

Graz, Austria). Digested solution was diluted with 11 mL of ultra-pure water and stored in universal tube for elemental analysis. A further 10-fold dilution procedure was performed to finalize digestion solution for the ICP-MS matrix before multi-element analysis (Fig. S1) (Joy et al., 2016). The certified reference material, Bush Twigs and Leaves/GB W07603/GSV-2, was served as quality control standard sample.

comparison analysis by the LSD-test with a significance level of p < 0.05. Figures were drawn using Sigma Plot 10.0.

2.4. Measurement of Se contents in xylem sap and roots

Fig. 1 displays the grain yield of the high- and low-Se rice cultivars, cultivated in the presence of different concentration of Se. The appli­ cation of Se had a limited effect on the grain yield of the two rice cul­ tivars in this experiment. In addition, there were no significant differences in yield between the two rice cultivars in the absence and presence of Se application.

3. Results 3.1. Grain weight of two rice cultivars

At the early grain filling stage (10 June 2019), a sharp blade was used to cut along the stem base of 4 replicates for each treatment, and the height of the remaining stem base was 10 cm. Approximately 0.5 g of absorbent cotton that had been weighed was placed on the cross-section of the stem base, ensuring that the stem cross-section maintained full contact with the absorbent cotton, which was then covered with a plastic film to prevent water from evaporating. After 12 h, the absorbent cotton was weighed to calculate the amount of xylem sap collected, and then the xylem sap in the absorbent cotton was diluted to 50 mL using deionized water to measure the Se concentration in the solution. Se content in the xylem sap was determined by wet digestion in a 4:1 HNO3–HClO4 mixture, followed by atomic fluorescence spectrometry of Se in the digested solution. Immediately after extraction of the xylem sap, the soil was washed with a large amount of fresh water. The roots of the rice plants were collected, dried, crushed, and sieved, followed by the determination of Se content in the roots by ICP-MS.

3.2. Grain, leaf and stem Se concentrations and accumulations of two rice cultivars As shown in Fig. 2, in the absence of Se application, the differences in the grain, leaf and stem Se concentrations between the two rice cultivars were not significant at either the early grain filling or maturation stage. Following Se application, the grain Se concentrations of the high-Se rice cultivar were significantly higher than those of the low-Se cultivar at both the early grain filling and maturation stages. The grain Se con­ centration of the high-Se rice cultivar was 1.4-fold that of the low-Se cultivar at the early filling stage and was 1.7-fold at the maturation stage. From the early grain filling stage to the maturation stage, the grain Se concentration of the high-Se rice cultivar decreased by 38.3%, whereas that of the low-Se rice cultivar decreased by 49.3%. In the presence of Se application, the leaf Se concentration of the high-Se rice cultivar was significantly higher at the early grain filling stage and was 1.3-fold that of the low-Se cultivar, while there was no significant dif­ ference in the leaf Se concentration between the two rice cultivars at the maturation stage. The leaf Se concentration of the high-Se rice cultivar decreased by 57.6% during grain filling stage with Se application, whereas that of the low-Se rice cultivar only decreased by 48.0%. The application of Se also led to higher stem Se concentration in the high-Se rice cultivar than in the low-Se cultivar at the early filling stage. The stem Se concentration of the high-Se rice cultivar decreased by 55.7% during the grain filling stage with Se application, and that of the low-Se rice cultivar decreased by 53.2%. As shown in Fig. 3, the differences in the grain, leaf and stem Se accumulations between the high- and low-Se rice cultivars were not significant at the early grain filling and maturation stages without Se

2.5. Measurement of Se distribution in rice grains Grain samples for each treatment from four replicates were collected at the maturation stage (25 July 2019), separated into rice and husk. The grain (husk included) weight was also recorded. The rice (without husk) was cut into three segments at equal distance, namely embryonic end, intermediate section and non-embryonic end, and then oven-dried at 40 ◦ C for further Se analysis. The husk Se concentration was also determined. The Se concentrations in different parts of rice grains were measured by ICP-MS. 2.6. Semi-quantitative RT-PCR Leaf samples were collected for transporters genes expression anal­ ysis during early filling stage (10 June 2019). Extraction of total RNA from frozen leaf samples was implemented by the Trizolreagent (Invi­ trogen, USA). The RNA was analysed using spectrophotometry (A260/ A280), and 1% agarose gel electrophoresis was used to ensure the absence of nucleic acid degradation. First-strand cDNA was synthesized from 1 μg of total RNA using M-MLV Reverse Transcriptase according to the manufacturer’s instructions (Promega, USA). Real-time quantitative PCR for detecting transcript levels of OsPT2, Sultr1;2 and NRT1.1B genes was determined by the SYBR Green qPCR SuperMix (Vazyme, China) and the ABIPRISM® 7500 Sequence Detection System. For PCR re­ actions, 5.0 μl of the synthesized cDNA, 0.5 μl of each primer, 10 μl of the SYBR Green qPCR SuperMix Kit and 4.0 μl of dH2O were used in a 20 μl reaction mixture. The OsPT2, Sultr1;2 and NRT1.1B primer pairs designed for real-time quantitative PCR are demonstrated in Table S2 (Zhang et al., 2019b). 2.7. Data analysis The translocation factor (TF grain/leaf) between grain and leaf was defined as the ratio of grain Se concentration to the corresponding leaf Se concentration at the maturation stage. The translocation factor (TF leaf/stem) between leaf and stem was defined as the ratio of leaf Se concentration to the corresponding stem Se concentration at the matu­ ration stage (Elshamy et al., 2019). All the data were statistically analysed using SPSS 12.0 software, and the mean values of each treatment group were subjected to a multiple

Fig. 1. Yield of the two rice cultivars. The bars indicate the standard error of the mean. 3

M. Zhang et al.

Ecotoxicology and Environmental Safety 205 (2020) 111372

Fig. 2. Dynamic changes in grain, leaf and stem Se concentrations of the two rice cultivars. The bars indicate the standard error of the mean.

application. In the presence of Se application, the grain and leaf Se ac­ cumulations of the high-Se rice cultivar were significantly higher than those of the low-Se rice cultivar at both the early grain filling and maturation stages. During the grain filling stage, grain Se accumulation of the high-Se rice cultivar increased by 139.9%, whereas that of the low-Se rice cultivar increased by 87.1%. In contrast, the leaf and stem Se accumulations of the high-Se rice cultivar decreased by 38.5% and 24.6%, respectively, whereas those of the low-Se rice cultivar decreased by 25.7% and 23.2%.

accumulation in grain of the high-Se rice cultivar was higher than that of the low-Se rice cultivar during the whole grain filling stage. The Se translocation factors (TF grain/leaf) between grain and leaf of the high-Se rice cultivar were significantly higher than those of the low-Se rice at the two Se application levels. There were no significant differences in the Se translocation factors (TF leaf/stem) between the two rice cultivars in the absence and presence of Se application.

3.3. Distribution proportions and translocation factors of Se in shoots of two rice cultivars

As shown in Fig. 5, Se application had a limited effect on the total weight of xylem sap of the two rice cultivars. However, the xylem sap weight of the high-Se rice cultivar was significantly higher than that of the low-Se cultivar. Without Se application, the difference in the xylem sap Se concentration between the two rice cultivars was not significant, whereas following Se application, the xylem sap Se concentration of the high-Se rice cultivar was significantly higher and was 1.5-fold that of the low-Se cultivar. Se application significantly increased the xylem sap Se concentrations of the two rice cultivars; the xylem sap Se concentration of the high-Se rice cultivar increased by 79.3%, while that of the low-Se cultivar increased by 41.7%. In the presence and absence of Se

3.4. Root and xylem sap Se concentrations of two rice cultivars

The proportions of Se accumulations in different parts of shoots are shown in Fig. 4. At the early grain filling stage, the lines of Se proportion went from stem to leaf and then down to grain of the high- and low-Se rice cultivars in the absence and presence of Se application. At matu­ ration stage, the proportions of Se accumulation decreased from leaf to grain and then to stem of the two rice cultivars in the absence of Se application, whereas the proportion of Se in stem was highest, followed by grain and leaf in the presence of Se application. The proportion of Se 4

M. Zhang et al.

Ecotoxicology and Environmental Safety 205 (2020) 111372

Fig. 3. Dynamic changes in grain, leaf and stem Se accumulations of the two rice cultivars. The bars indicate the standard error of the mean.

application, the root Se concentrations of the high-Se rice cultivar were significantly higher than those of the low-Se cultivar. Specifically, without Se application, the root Se concentration of the high-Se rice cultivar was 1.5-fold that of the low-Se cultivar, whereas in the presence of Se application, the root Se concentration of the high-Se rice cultivar was 1.7-fold that of the low-Se cultivar. Se application increased the root Se concentrations of both the two rice cultivars, such that the root Se concentration of the high-Se rice cultivar increased by 209.7%, while that of the low-Se cultivar increased by 165.4%.

3.6. Grain Se distribution of two rice cultivars As shown in Fig. 7, the distribution of Se in grain (without husk) was generally uniform in the high-Se rice cultivar in the presence and absence of Se application. There was no significant difference in grain (without husk) Se concentration between the embryo end, the inter­ mediate section, and non-embryo end of the high-Se rice cultivar. However, the distribution of Se in rice grain (without husk) of the low-Se rice cultivar was uniform in the absence of Se application but not uni­ form in the presence of Se application. The Se concentration in grain embryo end was significantly higher than in the intermediate section and non-embryo end of the low-Se rice cultivar. There was no significant difference in husk Se concentration between the high- and low-Se rice cultivar in the absence of Se application. Following Se application, the husk Se concentration of the high-Se rice cultivar was significantly higher than that of the low-Se cultivar.

3.5. Transcript levels of NRT1.1 B, OsPT2 and Sultr1; 2 in leaves of two rice cultivars As shown in Fig. 6, in the absence of Se application, the expression levels of NRT1.1B and OsPT2 genes in leaves of the high-Se rice cultivar were significantly lower than those in the low-Se cultivar. In the pres­ ence of Se application, the expression levels of NRT1.1B and OsPT2 were both significantly increased in the high-Se rice cultivar, and were significantly higher than those in the low-Se cultivars. However, in both the presence and absence of Se application, the expression levels of Sultr1;2 were significantly higher than those of the low-Se cultivar.

4. Discussion Selenium is a beneficial element that can improve photosynthesis (Feng et al., 2015) and stress defense of plants (Ashraf et al., 2017). 5

M. Zhang et al.

Ecotoxicology and Environmental Safety 205 (2020) 111372

Fig. 4. Proportion of shoot Se accumulation and Se translocation factors of the two rice cultivars. The bars indicate the standard error of the mean.

Previous studies showed that applying low concentrations of Se can promote the growth of wheat (Boldrin et al., 2016), winter jujube (Jing et al., 2017), sunflower (Garousi et al., 2018) and so on. However, Se demonstrated no obvious effect on grain yield of the two rice cultivars in this experiment. The discrepancy was likely to be caused by different experimental conditions, especially the concentration of Se applied, as well as the availability of Se in soil. The two rice cultivars showed no significant differences in yield, while the grain Se concentration of the high-Se rice cultivar was significantly higher than that of the low-Se cultivar. The difference in grain Se concentration between the highand low-Se rice cultivar was not due to the diluting effect of different yield. In terms of Se supplementation of human diets for enhanced nutrition, the accumulation of Se in grain should not exceed 0.3 mg kg− 1 (GB/T 22499, 2008). In the presence of Se application, the grain Se contents of the high- and low-Se rice cultivars were 1.75 mg kg− 1 and 1.05 mg kg− 1, respectively (Fig. 2b). Therefore, Se fertilizer application should be strictly controlled in the production of Se-rich rice. Previous study demonstrated that high-Se rice cultivar could obtain more external Se source by a stronger mass flow process (Zhang et al., 2019) and uptake more Se through better root architecture (Zhang et al., 2020). A similar result was also demonstrated in the present research. The root Se concentration of the high-Se rice cultivar was significantly higher than that of the low-Se rice cultivar, indicating that the former

had a stronger Se absorption capacity than the latter. However, the start of the grain filling process indicates that the rice plants are entering the late reproductive phase of growth, where there is a gradual decline in root activity and continuous weakening of the plant’s absorption ca­ pacity for soil nutrients (Yang et al., 2013). The supply of Se from roots to shoots is greatly reduced during grain filling stage due to root aging. Xylem sap plays a major role in long-distance transport of water, nu­ trients and metabolites (Sung et al., 2015), and xylem sap Se status can reflect the Se supply capacity of root system. Although the xylem sap Se concentration of the high-Se rice cultivar was higher than that of the low-Se rice cultivar, the Se concentration of xylem sap was extremely low during grain filling stage. Reutilization of mineral elements within tissues is also an important way for the efficient use of nutrients by plants (Lei et al., 2014). During the grain filling stage, rice plants reactivate the mineral elements accumulated in other tissues, which are reutilized and transported to the grains, such as N, P, K and so on (Zhang et al., 2018). After the reusable elements enter the aboveground parts of plants, they are still in the ionic stage or unstable compounds, which can be continuously decomposed and transferred to other organs (Cang and Li, 2017), and Se also has this reusable property. From the early grain filling stage to the maturation stage, the leaf Se concentration of the high-Se rice cultivar decreased by 57.6% and its accumulation decreased by 38.5%, whereas the flag leaf 6

M. Zhang et al.

Ecotoxicology and Environmental Safety 205 (2020) 111372

Fig. 5. Root and bleeding sap Se concentrations of the two rice cultivars. The bars indicate the standard error of the mean.

Fig. 6. Relative gene expressions (leaf transcript accumulation) of the peptide transporter NRT1.1B, phosphate transporter OsPT2 and sulphate transporter Sutr1;2 of the two rice cultivars. The bars indicate the standard error of the mean.

Se concentration of the low-Se rice cultivar decreased by 48.0% and its accumulation decreased by 25.7%. Selenite is absorbed and transported in plants by the phosphate transporter OsPT2 (Zhang et al., 2020), but upon entering the plants, selenite is readily converted to organic forms, including selenomethionine (SeMet), methyl-selenocysteine (MeSeCys) and secystine (SeCys) (Williams et al., 2009). Sun et al. (2010) investi­ gated organic Se species of 40 rice samples from four villages and results showed that SeMet was the major Se species, comprising 82.9% of organic Se. NRT1.1 B, a member of the PTR family, encodes a protein containing a peptide transporter domain, and Zhang et al. (2019b) showed that NRT1.1B overexpression significantly improved Se con­ centrations not only in shoots but also in rice grains by facilitating SeMet translocation. In the present research, the transcript levels of OsPT2, NRT1.1B and Sultr1;2 genes in the leaves of the high-Se rice cultivar

were all significantly higher than those of the low-Se rice cultivar by selenite application, which indicated that the high-Se rice cultivar possessed better transport carriers for Se redistribution. In fact, there is a competitive relationship between “source” and “reservoir” in plants, and reproductive organs served as reservoirs have the strongest nutrient competitiveness (Lei et al., 2014). Se accumulation in the shoots of the two rice cultivars decreased sharply in the grain filling stage, which implied that it gave the priority to the supply of grains. Inevitably, the Se translocation factor between the grain and leaf of the high-Se rice cultivar was significantly higher than that of the low-Se rice (Fig. 4b). These results demonstrated that the reutilization rate of Se in the tissues 7

M. Zhang et al.

Ecotoxicology and Environmental Safety 205 (2020) 111372

Fig. 7. Grain Se distribution and husk Se concentration of the two rice cultivars. The bars indicate the standard error of the mean.

of the high-Se rice cultivar was higher than that of the low-Se rice cultivar. During the grain filling stage, mineral elements enter the grains via the embryonic end and then transported towards the non-embryonic end within the grains. Previous studies showed that the distributions of different elements in the grain of the same rice cultivar were different (De Brier et al., 2015), and the distributions of the same elements in the grain of different rice cultivars were also different (Wang et al., 2016). Freeman et al. (2015) showed that in seeds of Se hyperaccumulator plants Stanleyapinnata and Astragalusbisulcatus, Se was mainly concen­ trated in the embryo. However, in the present research, Se distribution in the grains of the high-Se rice cultivar was relatively more uniform, whereas that of the low-Se rice cultivar demonstrated a significantly higher Se concentration in the embryonic end than in the other parts. The accumulation of Se at the embryonic end was not conducive to making full use of the storage capacity in the whole grain, and the low Se concentration in the non-embryonic end and intermediate section of the low-Se rice cultivar resulted in the low total Se concentration in the grain. A similar result was also reported in previous study by Chen et al. (2012), who analysed the distribution of mineral elements in grains of different Zn-efficient rice cultivars with μXRF and showed that the dis­ tribution of Zn in grains between high- and low-Zn rice cultivars was also quite different. This study indicated that the grain Se storage ca­ pacity of the high-Se rice cultivar is higher than that of the low-Se cultivar.

given their written permission to be named. Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. Acknowledgements This work was supported by the National Natural Science Funds of China (Grant No.31872176; 31501835), the Science and Technology Project of Guangzhou (Grant No. 201804010341) and the President’s Special Fund of the Guangdong Academy of Agricultural Sciences (Grant No. 201938). Appendix A. Supplementary data Supplementary data to this article can be found online at https://doi. org/10.1016/j.ecoenv.2020.111372. References Amos, W., Webb, S., Liu, Y., Andrews, J.C., Leduc, D.L., 2012. Imaging translocation and transformation of bioavailable selenium by stanleya pinnata, with x-ray microscopy. Anal. Bioanal. Chem. 404 (5), 1277–1285. Ashraf, M.A., Akbar, A., Parveen, A., Rasheed, R., Hussain, I., Iqbal, M., 2017. Phenological application of selenium differentially improves growth, oxidative defense and ion homeostasis in maize under salinity stress. Plant Physiol. Biochem. (Paris) 123, 268–280. Bao, S.D., 2002. Soil and Agricultural Chemistry Analysis, third ed. China Agriculture Press, Beijing. Barillas, J.R.V., Quinn, C.F., Freeman, J.L., Lindblom, S.D., Fakra, S.C., Marcus, M.A., Gilligan, T.M., Alford, E.R., Wangeline, A.L., Pilon-Smits, E.A.H., 2012. Selenium distribution and speciation in the hyperaccumulator Astragalus bisulcatus andassociated ecological partners. Plant Physiol 159, 1834–1844. Boldrin, P.F., De Figueiredo, M.A., Yang, Y., Luo, H., Giri, S., Hart, J.J., Faquin, V., Guilherme, L.R., Thannhauser, T.W., Li, L., 2016. Selenium promotes sulfur accumulation and plant growth in wheat (triticum aestivum). Physiol. Plantarum 158 (1), 80–91. Cang, J., Li, W., 2017. Plant Physiology. Higher Education Press, Beijing. Carey, A.M., Scheckel, K.G., Lombi, E., Newville, M., Choi, Y., Norton, G.J., Price, A.H., Meharg, A.A., 2012. Grain accumulation of selenium species in rice (oryza sativa l.). Environ. Sci. Technol. 46 (10), 5557–5564. Chen, L., Wu, C., Liao, H., Guo, W., Chen, W., Tian, S., 2012. In situ micro-distributions of mineral elements in rice grains with different zinc efficiency. Chin. J. Rice Sci. 26 (6), 706–714. De Brier, N., Gomand, S.V., Donner, E., Paterson, D., Delcour, J.A., Lombi, E., Smolders, E., 2015. Distribution of minerals in wheat grains (Triticum aestivum L.) and in roller milling fractions affected by pearling. J. Agric. Food Chem. 63 (4), 1276–1285. Du, Q.J., Zhang, Y.F., Zeng, B., Li, G.Q., Tang, S.M., 2009. Screening of rich selenium of rice variety from Hainan regions rich in selenium. Soil and Fertilizer Sciences in China 1, 46–49.

5. Conclusion The present study has provided strong physiological evidence that increasing tissue Se reutilization and optimising grain storage facilitates grain selenium accumulation in the high-Se rice cultivar. The paper expounds systematically that the combined action of enhanced shoot Se migration, upregulated Se transporter gene expression, and optimized grain storage space results in the improvement of grain Se loading in the high-Se rice cultivar, which also presents evidences for the high-Se crop cultivar as a potential carrier to produce Se-rich food in low-Se soil. Credit author statement I have made substantial contributions to the conception or design of the work; or the acquisition, analysis, or interpretation of data for the work. I have drafted the work or revised it critically for important in­ tellectual content. I have approved the final version to be published. I agree to be accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved. All persons who have made substantial contributions to the work reported in the manuscript, including those who provided editing and writing assistance and have 8

M. Zhang et al.

Ecotoxicology and Environmental Safety 205 (2020) 111372 Ecophysiology, vol. 11. Springer, Cham. https://doi.org/10.1007/978-3-319-562490_11. Silva, V.M., Boleta, E.H.M., Lanza, M.G.D.B., Lavres, J., Martins, J.T., Santos, E.F., Santos, F.L.M.S., Putti, F.F., Junior, E.F., White, P.J., Broadley, M.B., Carvahlo, H.W. P., Reis, A.R., 2018. Physiological, biochemical, and ultrastructural characterization of selenium toxicity in cowpea plants. Environ. Exp. Bot. 150 (6), 172–182. Sun, G.X., Liu, X., Williams, P.N., Zhu, Y.G., 2010. Distribution and translocation of selenium from soil to grain and its speciation in paddy rice (oryza sativa l.). Environ. Sci. Technol. 44 (17), 6706–6711. Sung, J., Sonn, Y., Lee, Y., Kang, S., Ha, S., Krishnan, H.B., Oh, T., 2015. Compositional changes of selected amino acids, organic acids, and soluble sugars in the xylem sap of n, p, or k-deficient tomato plants. J. Plant Nutr. Soil Sci. 178 (5), 792–797. Taiz, L., Zeiger, E., 2010. Plant Physiology, fifth ed. Sinauer Associates, Sun-derland. Wang, J.L., Li, L.H., Duan, Z.M., Li, R.W., Pan, Q.L., Chen, L., 2016. Trace elements anslysis in Chinese rice of different brands as studied by X-ray fluorescence. J. Beijing Normal Univ. (Nat. Sci.) 52 (2), 156–160. Williams, P.N., Lombi, E., Sun, G.X., Scheckel, K., Zhu, Y.G., Feng, X., Zhu, J., Carey, A. M., Adomake, E., Lawgali, Y., Deacon, C., Meharg, A.A., 2009. Selenium characterization in the global rice supply chain. Environ. Sci. Technol. 43 (15), 6024–6030. Yang, J.R., Xie, W.L., Jian, L.I., 2015. The rank of the mobility and reusability of mineral elements in naval orange plant. Southeast Horticulture 3 (2), 1–11. Yang, Z.Y., Sun, Y.J., Xu, H., Qin, J., Jia, X.W., Ma, J., 2013. Influence of cultivation methods and no-tillage on root senescence at filling stage and grain-filling properties of Eryou 498. Sci. Agric. Sin. 46 (7), 1347–1358. Yoshida, M., Yasumoto, K., 1987. Selenium contents of rice grown at various sites in Japan. J. Food Compos. Anal. 1 (1), 71–75. Zhang, L., Hu, B., Deng, K., Gao, X., Sun, G., Zhang, Z., Li, P., Wang, W., Li, H., Zhang, Z., Fu, Z., Yang, J., Gao, S., Li, L., Yu, F., Li, Y., Ling, H., Chu, C., 2019a. NRT1.1B improves selenium concentrations in rice grains by facilitating selenomethinone translocation. Plant Biotechnol. J. 17 (6), 1058–1068. Zhang, L., Shi, W., Wang, X., Zhou, X., 2006. Genotypic differences in selenium accumulation in rice seedlings at early growth stage and analysis of dominant factors influencing selenium content in rice seeds. J. Plant Nutr. 29 (9), 1601–1618. Zhang, M., Tang, S.H., Huang, Q.Y., Pang, Y.W., Yi, Q., Huang, X., Li, P., Fu, H.T., 2018. The nutrient supply characteristics of co-application of slow-release urea and common urea in double-cropping rice. Sci. Agric. Sin. 51 (20), 3985–3995. Zhang, M., Wilson, L., Xing, G., Jiang, L., Tang, S., 2020. Optimizing root architecture and increasing transporter gene expression are strategies to promote selenium uptake by high-se accumulating rice cultivar. Plant Soil 447 (1), 319–332. Zhang, M., Xing, G.F., Tang, S.H., Pang, Y.W., Yi, Q., Huang, Q.Y., Huang, X., Huang, J. F., Li, P., Fu, H.T., 2019b. Improving soil selenium availability as a strategy to promote selenium uptake by high-Se rice cultivar. Environ. Exp. Bot. 163, 45–54. Zhou, X.B., Yu, S.H., Lai, F., 2014. Mechanisms of differences in selenium absorption and transport between rice plants different in cultivar. Acta Pedol. Sin. 3, 594–599. Zhou, X.B., Zhang, C.M., Wang, Y.K., Xu, W.H., 2016. Differences in selenium accumulation in grains of two rice cultivars. Bangladesh J. Bot. 45 (4), 811–818.

Elshamy, M.M., Heikal, Y.M., Bonanomi, G., 2019. Phytoremediation efficiency of Portulaca oleracea L naturally growing in some industrial sites, Dakahlia District, Egypt. Chemosphere 225, 678–687. Feng, T., Chen, S.S., Gao, D.Q., Liu, G.Q., Bai, H.X., Li, A., Peng, L.X., Ren, Z.Y., 2015. Selenium improves photosynthesis and protects photosystem ii in pear (pyrus bretschneideri), grape (vitis vinifera), and peach (prunus persica). Photosynthetica 53 (4), 609–612. Freeman, J.L., Marcus, M.A., Fakra, S.C., Devonshire, J., McGrath, S.P., Quinn, C.F., Pilon-Smits, E.A.H., 2012. Selenium hyperaccumulator plants Stanleya pinnata and Astragalus bisulcatus are colonized by Se-resistant, Se-excluding wasp and beetle seed herbivores. PloS One 12 (7), e50516. Garousi, F., Kov´ acs, B., Veres, S., 2018. Sunflower seedlings hyperaccumulate selenium. Acta Biol. Hung. 69 (2), 197–209. GB/T 22499, 2008. Rich Selenium Paddy. Standardization Administration of China, Beijing. Ilbas, A.I., Yilmaz, S., Akbulut, M., Bogdevich, O., 2012. Uptake and distribution of selenium, nitrogen and sulfur in three barley cultivars subjected to selenium applications. J. Plant Nutr. 35, 442–452. Jing, D.W., Du, Z.Y., Ma, H.L., Ma, B.Y., Liu, F.C., Song, Y.G., Xu, Y.F., Li, L., 2017. Selenium enrichment, fruit quality and yield of winter jujube as affected by addition of sodium selenite. Sci. Scientia Horticulturae 225, 1–5. Joy, E.J.M., Ander, E.L., Broadley, M.R., Young, S.D., Chilimba, A.D.C., Hamilton, E.M., Watts, M.J., 2016. Elemental composition of malawian rice. Environ. Geochem. Hlth. 39 (4), 1–11. Lei, G.J., Zhu, X.F., Wang, Z.W., Dong, F., Zheng, S.J., 2014. Abscisic acid alleviates iron deficiency by promoting root iron reutilization and transport from root to shoot in arabidopsis. Plant Cell Environ. 37 (4), 852–863. Li, J.T., Gurajala, H.K., Wu, L., Antony, V.D.E., Qiu, R.L., Baker, A.J.M., Tang, Y.T., Yang, X.E., Shu, W.S., 2018. Hyperaccumulator plants from China: a synthesis of the current state of knowledge. Environ. Sci. Technol. 52 (21), 11980–11994. Maruyama-Nakashita, A., 2016. Combinatorial use of sulfur-responsive regions of sulfate transporters provides a highly sensitive plant-based system for detecting selenate and chromate in the environment. Soil Sci. Plant Nutr. 62 (4), 1–6. Mehdawi, A.E.F., Jiang, Y., Guignardi, Z.S., Esmat, A., Pilon, M., Pilon-Smits, E.A.H., Schiavon, M., 2018. Influence of sulfate supply on selenium uptake dynamics and expression of sulfate/selenate transporters in selenium hyperaccumulator and nonhyperaccumulators brassicaceae. New Phytol. 217 (1), 194–205. Mitani-Ueno, N., Yamaji, N., Ma, J.F., 2018. Transport system of mineral elements in rice. In: Sasaki, T., Ashikari, M. (Eds.), Rice Genomics, Genetics and Breeding. Springer, Singapore. https://doi.org/10.1007/978-981-10-7461-5_13. NY/T 1101, 2006. Determination of Selenium in Soils. Ministry of Agriculture of the People’s Republic of China, Beijing. Pilon-Smits, E.A.H., 2017. Selenium in Plants. Progress in Botany. Springer International Publishing. Quinn, C.F., El Mehdawi, A.F., Pilon-Smits, E.A.H., 2017. Ecology of selenium in plants. In: Pilon-Smits, E., Winkel, L., Lin, Z.Q. (Eds.), Selenium in Plants. Plant

9