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The role of node restriction on Cd accumulation in the brown rice of twelve Chinese rice (Oryza sativa L.) cultivars Gaoxiang Huang, Changfeng Ding, Fuyu Guo, Xiaogang Li, Zhigao Zhou, Taolin Zhang, and Xingxiang Wang J. Agric. Food Chem., Just Accepted Manuscript • DOI: 10.1021/acs.jafc.7b03333 • Publication Date (Web): 01 Nov 2017 Downloaded from http://pubs.acs.org on November 2, 2017

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Journal of Agricultural and Food Chemistry

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Title page

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1) Title: The role of node restriction on Cd accumulation in the brown rice of twelve

3 4 5 6

Chinese rice (Oryza sativa L.) cultivars 2) Author names: Gaoxiang Huang

a,c

, Changfeng Ding a, Fuyu Guo

a,c

, Xiaogang

Li a, Zhigao Zhou a, Taolin Zhang a, Xingxiang Wang a,b 3) Author affiliation:

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a. Key Laboratory of Soil Environment and Pollution Remediation, Institute of

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Soil Science, Chinese Academy of Sciences, Nanjing 210008, China

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b. Ecological Experimental Station of Red Soil, Chinese Academy of Sciences,

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Yingtan 335211, China

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c. University of Chinese Academy of Sciences, Beijing 100049, China

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4) Corresponding author:

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Professor Xingxiang Wang

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71 East Beijing Road, Institute of Soil Science, Chinese Academy of Sciences,

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Nanjing 210008, China

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Tel.: +86–25–86881200

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Fax: +86–25–86881000

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E–mail: [email protected]

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ACS Paragon Plus Environment


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ABSTRACT: For selection or breeding of rice (Oryza sativa L.) cultivars with low

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Cd–affinity, the role of node Cd–restriction on Cd accumulation in brown rice was

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studied. A pot experiment was conducted to investigate the concentration of Cd in

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different sections of twelve Chinese rice cultivars. The results indicated that the Cd

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accumulation in the brown rice was mainly dependent on the root or shoot Cd

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concentration. Among the cultivars with nearly equal shoot Cd concentrations, Cd

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accumulation in brown rice was mainly dependent on the transport of Cd in the shoot.

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However, the Cd transport in the shoot was significantly restricted by the nodes,

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especially by the first node. Furthermore, the area of the diffuse vascular bundle in the

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junctional region of the flag leaf and the first node was a key contributor to the

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variations in Cd–restriction by the nodes.

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KEYWORDS: Cadmium, rice, node–restriction, translocation, vascular bundles.

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INTRODUCTION Cadmium (Cd) is a highly toxic metal and is regarded as a human carcinogen by the

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National Toxicology Program

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Committee on Food Additives (JECFA), the tolerable weekly intake of Cd is 7 µg/kg

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for a human body 3. Long–term exposure to Cd causes many chronic diseases 4,

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including itai–itai disease, cancer and kidney disease

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approximately 90% of the Cd in the human body reportedly comes from food 8. As

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one of the main cereal types, rice (Oryza sativa L.) is widely consumed by half of the

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world’s population. However, rice has a higher Cd–bioaccumulation ability than most

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other cereals 9, and brown rice is considered to be a major source of the dietary intake

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of Cd for humans in rice–planting regions

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brown rice is vital to human health.

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1, 2

. According to the Joint FAO/WHO Expert

5-7

. For nonsmokers,

10

. Therefore, reducing the Cd level in

Cd accumulation in grain differs among different cultivars

11-15

, which is affected

46

by many factors, such as the transporter protein 16, tissue biomass 17 and translocation

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characters

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variability is helpful to identify cultivars with low Cd accumulation. Cd accumulation

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in brown rice is related to the Cd–uptake by the root from the soil. The ability of the

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root to uptake Cd differs among different cultivars

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attributed to the different Cd transporters in the root

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root, upward translocation plays an important role in the allocation of Cd to brown

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rice

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through several nodes where the elemental redistribution processes occur 24. A node is

18

. A better understanding of the mechanisms that contribute to this

13, 19

, and this variability is mainly

20, 21

. Once Cd is taken into the

22, 23

. The upward translocation of Cd from the shoots to brown rice passes

3



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the junction between the leaves or branches and the stem in rice, and there are

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generally 13–18 nodes on a stem. However, only a small number of aboveground

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nodes are elongated in the jointing stage, depending on the genotypes

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nodes, there are three main phases of vascular bundles (VBs) as follows: enlarged or

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large VBs (EVBs or LVBs), transit VBs (TVBs) and diffuse VBs (DVBs). An EVB is

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enlarged at the node by increasing the number of xylem vessels and sieve tubes; the

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TVB is a transit phase that occurs without extra enlargement in the node; the DVB

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surrounds the EVB and assembles just above the node 26. The VBs in nodes also play

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an important role in Cd translocation 27, 28, e.g., most Cd in the uppermost node of rice

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is accumulated on EVBs and DVBs, which inhibit Cd transportation to the internodes

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28, 29

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internodes on Cd translocation in rice and the role of different VB systems on Cd

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transport inside the nodes.

25

. Inside the

. However, few studies have investigated the role of different nodes and

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Therefore, the objectives of this work were 1) to investigate the variability of the

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node restriction of Cd among twelve Chinese rice cultivars and analyze its impacts on

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Cd allocation to the brown rice and 2) to test one hypothesis, which is that differences

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in node restriction rely on differences in node vasculatures.

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MATERIALS AND METHODS

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Soil Preparation and Pot Trial

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An acid paddy soil (0–20 cm) was collected from Xinghua county, Jiangsu

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province, China. The principal physical and chemical properties of the soil were as

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follows: pH (H2O), 4.90; organic matter (OM), 28.3 g kg−1; cation exchange capacity 4


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(CEC), 16.3 cmol kg–1; total nitrogen (TN), 1.59 g kg−1; available N (AN), 0.19 g kg−1;

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total phosphorus (TP), 0.94 g kg−1; available P (AP), 0.03 g kg−1; total potassium (TK),

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15.2 g kg−1; available K (AK), 0.28 g kg−1; total Cd, 0.14 mg kg–1; available Cd (0.01

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M CaCl2 extractable Cd), 0.024 mg kg−1; and soil texture as follows: sand (10.0%),

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silt (68.4%), clay (21.6%). After air–drying and passing through a 2-mm sieve, 0.6 mg

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kg-1 Cd (3CdSO4 ·8H2O) DWsoil was spiked to the soil (background Cd, 0.14 mg kg-1),

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and the soil was maintained at 80% of the field water holding capacity for three month

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to simulate long term Cd-contaminated field soil

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measured soil Cd concentration was 0.71 mg kg-1, and the soil was used for the pot

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experiment after air–drying and passing through a 2-mm sieve again.

30

. After the aging process, the

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The pot experiment was carried out in the greenhouse of the Institute of Soil

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Science Chinese Academy of Sciences, Nanjing, Jiangsu province, China. Each pot

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(25 cm diameter and 22 cm high) was filled with 6 kg of soil. The soil was mixed with

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base fertilizer composed as follows: 0.2 g N kg−1 (CO (NH2)2), 0.15 g P2O5 kg−1

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(CaH2PO4 H2O) and 0.2 g K2O kg−1 (KCl). Then, the soil was kept moist for seven

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days before planting the rice.

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Plant Materials and Culture

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Twelve widely cultivated Chinese rice (Oryza sativa L.) cultivars were used in the

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present study (Table 1). The seeds of all cultivars were surface sterilized in 15% H2O2

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solution for 15 min, fully rinsed with deionized water, and then soaked in deionized

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water in the dark for 24 h at 25°C. Thereafter, the seeds were left on moist gauze for

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24 h in the dark at 25°C. The germinated seeds of each cultivar were then pre–grown 5



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in the nursery tray for 20 days in the greenhouse, and uniform seedlings were

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prepared for the pot experiment on May 26th, 2016. Each cultivar had three pots as

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replicates, and each pot planted six uniform seedlings. All pots were arranged

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randomly in the greenhouse and kept submerged (2–3 cm depth of water above soil

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surface) during the entire growth stage.

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One week before harvesting at the mature stage of each rice cultivar, three similar

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plants of each cultivar from three of the replicate pots were selected to observe the VB

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structure in the first node (N1). The first node beneath the panicle of each plant was

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sampled using a razor blade and immediately stored in an MES (2–(N–morpholino)

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ethane sulfonic acid) buffer to observe its cellular structure 31. When the rice was ripe,

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the remaining five rice plants in each pot were harvested. The rice plants were

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sampled separately from top to bottom, including the brown rice (BR), flag leaf (FL),

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first internode (IN1), first node (N1), second leaf (L2), second internode (IN2),

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second node (N2), third leaf (L3), third internode (IN3), third node (N3), fourth leaf

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(L4), fourth internode (IN4), fourth node (N4) and roots (Figure 1). Each part was

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rinsed three times with tap water followed by deionized water then oven–dried to a

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constant weight at 75°C. After weighing the dry weight (DW), all the samples were

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milled to a fine powder using a blender (A11 basic, IKA, Germany) to prepare for the

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chemical analysis of Cd.

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Cellular Structure Observation

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The fresh first nodes in the MES buffer were cut into 0.4–0.5-mm thick sections by

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a razor blade, and the sections were then placed into a planchette coated with 6


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hexadecane, and another planchette was used to cover the sections. The sections were

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precooled by a high–pressure freezer (HPM100, Leica, Wetzlar, Germany) with a

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pressure of 210 MPa at –196°C for 30 s and then freeze–dried. Because the junction

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of the flag leaf and N1 (JFLN) is the most important region, which is directly

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connected to the panicle and flag leaf in the first node, 10-µm thick sections in this

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junctional region were sectioned by a Leica RM 2265 rotary microtome (Leica,

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Wetzlar, Germany) 31. These thick sections were stained with safranine and fast green

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to observe the cellular structure using a digital slide scanner (Pannoramic MIDI, 3D

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HISTECH, Hungary). The scanning images were analyzed using the professional

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software – Pannoramic viewer, and the images were enlarged at 1–400×

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magnification.

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Chemical Analysis of Plant and Soil

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All plant samples were digested with HNO3 and H2O2. Approximately 0.15 g of the

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samples were placed into polytetrafluoroethylene tubes, and then 4 mL of high–purity

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HNO3 was added for cold digestion overnight. Thereafter, 3 mL of high–purity H2O2

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was added to the tubes for digestion in high pressure sealed digestion vessels

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(HTLAB, HR-25, Shanghai) at 140°C for 4 h according to the Determination of

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Cadmium in Foods, National Food Safety Standard of China (GB/T 5009.15-2003).

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The tubes were removed when the system cooled. Finally, the digestive solution was

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transferred to centrifuge tubes after an acid evaporation and was passed through a

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0.45-µm filter membrane before the Cd measurement by graphite furnace atomic

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absorption spectroscopy (SpectrAA 220Z, Varian, USA) was performed. The 7



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detection limit is 0.001 µg kg–1, which equates to 0.1 µg kg–1 in the brown rice, and a

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standard reference material, green Chinese onion material (GBW10049, National

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Research Center for Certified Reference Materials, China) was used for the quality

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control. The recovery rate ranged from 95 to 106%.

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The soil pH was measured using a 1:2.5 soil-to-water ratio. The soil organic matter

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(K2CrO4−H2SO4 oil-bath-heating), cation exchange capacity (1 M ammonium acetate

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leaching method at pH 7.0), soil texture (Pipette method ), TN (N/C Soil Analyzer

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(Flash, EA, 1112 Series, Italy)), TP (H2SO4-H2O2 digestion, molybdenum-antimony

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colorimetric method), TK (H2SO4-H2O2 digestion, flame atomic absorption

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spectrophotometry), AN (Conway method), AP (Olsen method), and AK (1 M

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ammonium acetate extraction) were analyzed according to the routine analytical

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methods of agricultural chemistry in soil 32.

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Data Analysis

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The translocation factors (TFs, concentration ration between the upper parts and the

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lower parts) of Cd from the root to the shoot (the part between the root and the ear of

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rice) or brown rice (BR), from Nn to INn or Ln (n = 1, 2, 3, 4) and the translocation

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efficiency of Cd from IN4 to IN1 (TEIN4–IN1) were calculated based on the following

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equations 33: = ⁄ = ⁄

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where a represents the root, N1, N2, N3 or N4; b represents shoot, BR, IN1, IN2, IN3 8


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or IN4; Ca and Cb represent the Cd concentrations in a and b, respectively; and CIN1

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and CIN4 represent the Cd concentrations in IN1 and IN4, respectively.

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The total dry weight from N4 to the IN1 accounts for more than 90% of the dry

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weight of the shoot, so other parts in the shoot were not taken into consideration, and

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the weighted average concentration of Cd in the shoot was calculated based on the

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following equation: =

∑( × + × + × ) ∑( + + )

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where the value of n was 4; i represents 1, 2, 3, 4; CdNi, CdINi, and CdLi represent the

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concentration of Cd in Ni, INi, and Li, respectively; DWNi, DWINi, and DWLi represent

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the dry weight of Ni, INi, and Li, respectively.

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Statistical Analysis

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The statistical relationships among all test data were analyzed by one–way analysis

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of variance (ANOVA) through SPSS 19.0, and the least significant difference (LSD)

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was used to check the significant differences between mean values (P < 0.05). The

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data were expressed as the mean ± SD (n = 3). All figures were generated using the

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SigmaPlot 10.0 software.

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RESULTS

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Cd Accumulation in the Root, Shoot and Brown Rice of Different Rice Cultivars

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As shown in Figure 2, the Cd concentrations in the root significantly differed

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among the twelve rice cultivars, ranging on average from 0.27 to 5.22 mg kg–1 DW.

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ZXY146 and CLY4418 had the lowest and the highest root Cd concentrations among

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all cultivars, respectively. 9



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Cd concentration in the shoot also differed significantly among the twelve cultivars

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(Figure 2), ranging on average from 0.08 to 1.27 mg kg–1 DW. NXY8 and CLY4418

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had the lowest and the highest shoot Cd concentrations among all cultivars,

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respectively.

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In the BR, Cd concentration shows a significant (P < 0.05) difference among the

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twelve cultivars, ranging on average from 0.020 to 0.222 mg kg–1 DW (Figure 2).

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TYJ1 and NJ9108 had the lowest Cd concentrations in the BR, and CLY4418 had the

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highest Cd concentration in the BR, which exceeded the Chinese food safety standard

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(0.20 mg kg–1 DW) and was 11 times higher than that of TYJ1 and NJ9108 (Figure 2).

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In addition, the tendency of the concentrations of Cd in the BR was consistent with

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that of the concentrations of Cd in the root in the cultivars of YYJ2, SJ9, NJ4, ZD18,

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WYJ27 and CLY4418, while no positive relationship between BR Cd and root Cd

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concentration was found in the other cultivars.

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Cd Accumulation in the Nodes and Internodes of the Cultivars with Similar Cd

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Levels in the Shoots

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The concentrations of Cd differed in the nodes at different locations as well as in

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the internodes (Figure 3). From N4 to N1 in the cultivars with similar shoot Cd (TYJ1,

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NJ9108, YY2640, NJ52, and ZXY146), Cd concentrations in the nodes ranged from

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0.46 to 1.12 mg kg–1 DW, which were even higher than those of the root from the

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same cultivar. The concentration of Cd in N1 was the highest among all the nodes and

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approximately 1.5 to 2 times higher than that of the other nodes, but no obvious

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difference was found among the lower nodes (N2, N3, and N4). Cd concentrations in 10


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the internodes were approximately 3 to 10 times lower than those of nodes located at

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both sides of the internodes and gradually decreased as the location rose.

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Cd concentrations in the nodes and internodes significantly differed among the

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cultivars. TYJ1 had the highest Cd concentration in N1 and the lowest Cd

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concentration in IN1, with values of 1.12 and 0.03 mg kg–1 DW, respectively, whereas

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the lowest Cd concentrations in N1 and the highest Cd in IN1 were both in ZXY146,

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with values of 0.87 and 0.07 mg kg–1 DW, respectively (Figure 3).

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Internal Transport Characteristics of Cd in the Cultivars with Similar Cd Levels

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in the Shoots

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The translocation factors from the root to BR (TFroot–BR) were significantly different

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among the five rice cultivars with similar shoot Cd levels, with concentration ratios

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ranging from 0.036 to 0.153 (Figure 4). ZXY146 and TYJ1 had the highest and the

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lowest TFroot–BR, respectively. Among these cultivars, the translocation factors from

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the root to the shoot (TFroot–shoot) were similar (Figure 4). In the shoot, the

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translocation efficiency of Cd from the fourth internode to the first internode

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(TEIN4–IN1) ranged, on average, from 0.16 to 0.43 and differed significantly (P NJ52 > NJ9108, as well as

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the total area of VBs (Table 3), which was contrary to their ability to restrict the

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upward translocation of Cd. Interestingly, the variability of the DVB area was greater

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than that of the TVB area; the DVB area in ZXY146 was 2 times higher than that in

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NJ52 and 13 times higher than that in NJ9108. The variability in the TVB area was

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smaller, and the TVB areas in ZXY146 and NJ52 were similar and were approximately 12


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31.3% higher than that in NJ9108. The VB (TVB, DVB and total VB) area was

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significantly positively correlated with the BR Cd or TFN1-IN1 (Table 3).

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DISCUSSION

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Cd transportation to brown rice from the soil will pass through three main processes,

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including the root uptake from soil, transport from the root to the shoot, and transport

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from the shoot to the brown rice. Root Cd concentration differed significantly among

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different cultivars in this work, which mainly resulted from the interaction of the three

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physiological processes. First, Cd uptake by the roots differed among the different

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cultivars

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Resistance–Associated Macrophage Protein) transporter protein

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sequestration in iron plaque depends on the ability to produce radial oxygen loss

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(ROL) involved in the formation of iron plaque on the root surface

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sequestration in root cells depends on the expression of the transporter protein (like

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OsHMA3) involved in the sequestration of Cd in the vacuolar membrane 37. Third, Cd

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loading in the xylem depends on the expression of the transporter protein (like

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OsHMA2) 38. The present work found a significant positive linear correlation between

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brown rice and root Cd concentration (R2 = 0.96, P < 0.01, n = 36) (Figure 6a), and

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this correlation was still significantly positive (R2 = 0.67, P < 0.01, n = 33), excluding

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CLY4418 (Figure S5), suggesting that the Cd concentration in brown rice was

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significantly affected by the root Cd concentration. However, the Cd accumulation in

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the brown rice of the cultivars with similar root Cd (i.e., TYJ1, NJ9108, NXY8,

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YY2640, NJ52, and ZXY146) mainly depended on the transport of Cd from the root to

because

of

the

different

expression

of

13


the

Nramp5

(Natural

20, 34

. Second, Cd

35, 36

, and Cd


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the brown rice, including root–shoot transport and shoot–brown rice transport. In the

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processes of upward Cd transport, xylem loading and phloem transport are the two

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main pathways

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shoot and is considered to be the main and most common physiological factor

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accounting for the different allocations of Cd to different rice parts

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translocation factors from the root to the shoot were similar in the cultivars with

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similar root Cd levels, except for NXY8, in this work. Therefore, Cd accumulation in

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the brown rice of the five cultivars with similar shoot Cd amounts mainly depended

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on the shoot–brown rice transport (Figure 6b) mediated mainly by phloem transport.

281

39-41

. Xylem loading mediates Cd translocation from the root to the

Nodes are the central organ where xylem–phloem transfer occurs

40, 42

. The

43, 44

, and the

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totally organized vascular systems in the node includes the EVB, TVB and DVB 25.

283

An axial vascular system includes three nodes and a leaf (Nn+2, Nn+1, Nn and Ln),

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elements (including Cd) transport from the DVB of Nn+2 to the TVB of Nn+1, then to

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the EVB of Nn, and finally to the DVB of Nn via nodal vascular anastomosis or to Ln

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via the xylem of EVB 26. In this way, Cd was finally transferred to the panicle and the

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brown rice through several circular translocations. The nodes had strong Cd

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accumulation ability in the shoot and controlled the translocation efficiency of Cd

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(Table 2 and Figure 4). Moreover, the TFN1–IN1 was the lowest among all nodes (Table

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2), indicating that the first node played the most important role in restricting the

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transport of Cd to the brown rice. This phenomenon most likely occurred because Cd

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accumulation in the node mainly depends on the chelation of metallothionein 28, and

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the transcript levels of cDNA that encode metallothionein–like proteins are enhanced 14


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in the first node 45, and, thus, more Cd will be chelated in the first node. These results

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suggested that the upward translocation of Cd in the shoots was mainly restricted by

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the nodes, especially by the first node, and, thus, Cd accumulation in the brown rice

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was much lower than in the other parts of the rice plant. However, because Cd

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accumulated in the node would be readily transported to the leaves via the EVB 28, the

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nodes showed no significant obstruction of Cd transport to the leaves.

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The Cd restriction in the nodes differed among the different rice cultivars, which

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was associated with the difference in the vascular bundles in the node

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morphological variation in the VB in the junction of the flag leaf and the first nodes

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(JFLN) was considered to contribute greatly to this variability in the present work.

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There are three phases of VB in a node. At the bottom of the node, only EVB and

305

TVB exist in the VB system, and they are connected by nodal vascular anastomosis

306

(NVA). Cd in the node mainly accumulated on the EVB surface due to the

307

metallothionein 28, and it can be partly transported to the TVB by transporters via the

308

NVA or to the EVB in the upper region 16. In the middle section of the node, the NVA

309

was changed to an axial DVB 46, and thus, three VB phases – EVB, TVB and DVB –

310

constitute the basic structure of each node. Cd is also sequestrated on the EVB and

311

transported to the DVB or the upper EVB. At the upper part of the node, where the

312

leaf sheath is located (the junction of the flag leaf and the first nodes, JFLN), the EVB

313

is located in the leaf sheath, and only TVB and DVB exist in the central organ (Figure

314

5). In the JFLN region, Cd can be directly transported to the flag leaf via the EVB or

315

transported to the first internode and panicle via the DVB, indicating that Cd transport 15


26

. The


316

in this region plays an important role in Cd allocation to the brown rice, which was

317

confirmed by the significant positive correlation (R2 = 0.96, P < 0.01) between the Cd

318

concentration in the first internode and the concentration of Cd in brown rice (Figure

319

7a). Therefore, the morphological difference of the DVB in the JFLN region may

320

contribute to the variability of the brown rice Cd concentration in cultivars with

321

similar shoot Cd concentrations. In this work, we found that the Cd transport to the

322

brown rice was easier in the cultivars with larger DVB areas in the JFLN region. This

323

was because the phloem transport of Cd through the DVB is the main pathway to the

324

panicle or brown rice

325

transport of Cd, so more Cd will be transferred to the brown rice.

29

, and a larger DVB area may provide more space for the

326

In addition, Cd in the first node was mainly sequestrated on the surface of EVB 28,

327

which would be partly transported to the DVB via nodal vascular anastomosis, then to

328

the panicle via the phloem of DVB; another portion would be transported to the flag

329

leaf via the xylem of EVB 26. When the DVB area was too small to load sufficient Cd,

330

such as in NJ9108, only a small proportion of Cd might be transferred to the first

331

internode, and more Cd might be transferred to the flag leaf compared with the

332

cultivars with larger DVB areas (such as in ZXY146). The significant negative

333

correlation between TFN1-IN1 and TFN1-FL (Figure 7b) further confirmed this

334

assumption. As a result, Cd transport to the flag leaf seemed to play a negative role in

335

Cd accumulation in brown rice (Figure 7c).

336

Therefore, selecting or breeding cultivars with lower root Cd or a smaller DVB area

337

in the first node is promising to reduce Cd accumulation in brown rice, therefore 16


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lowering the health risks associated with the consumption of rice. However, the

339

factors controlling the DVB area of the node require further study.

340

Supporting Information

341

Cd concentrations of the nodes and internodes in other cultivars NXY8, YYJ2, SJ9,

342

NJ4, ZD18, WYJ27, and CLY4418 (Figure S1); Cd concentrations of leaves in all

343

cultivars (Figure S2); Cd translocation characters in other cultivars NXY8, YYJ2, SJ9,

344

NJ4, ZD18, WYJ27, and CLY4418 (Figure S3 and Table S1); dry weight of rice root,

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shoot and brown rice of all cultivars (Figure S4); Linear correlation between the Cd

346

concentration in the brown rice and in the root of all rice cultivars excluded CLY4418

347

(Figure S5).

348

Funding

349

This work was jointly sponsored by the National Key Technology Research and

350

Development Program of China (2015BAD05B04), the National Natural Science

351

Foundation of China (41501347), the Natural Science Foundation of Jiangsu Province,

352

China (BK20151054), and the Knowledge Innovation Program of the Chinese

353

Academy of Sciences (ISSASIP1612).

354

Notes

355

The authors declare no competing financial interest.

17



356

REFERENCES

357

1.

358

Mori, S.; Wu, J. Z.; Handa, H.; Itoh, T.; Matsumoto, T., Genome-wide transcriptome

359

analysis reveals that cadmium stress signaling controls the expression of genes in

360

drought stress signal pathways in rice. Plos One 2014, 9, e96946.

361

2.

362

Health and Human Services: Research Triangle Park, NC, 2000; pp III-42-III-44.

363

3.

364

Health Organization Technical Report 2000, 62, 165–166.

365

4. Aziz, R.; Rafiq, M. T.; Li, T. Q.; Liu, D.; He, Z. L.; Stoffella, P. J.; Sun, K.; Xiaoe,

366

Y., Uptake of cadmium by rice grown on contaminated soils and its

367

bioavailability/toxicity in human cell lines (Caco-2/HL-7702). J Agr Food Chem 2015,

368

63, 3599-3608.

369

5.

370

Nomoto, K.; Hayashi, S.; Miwa, S.; Nakajima, T.; Nakanishi, Y.; Aoshima, K.; Imura,

371

J., The liver in itai-itai disease (chronic cadmium poisoning): pathological features

372

and metallothionein expression. Modern Pathol 2013, 26, 1228-1234.

373

6. Skroder, H.; Hawkesworth, S.; Kippler, M.; El Arifeen, S.; Wagatsuma, Y.; Moore,

374

S. E.; Vahter, M., Kidney function and blood pressure in preschool-aged children

375

exposed to cadmium and arsenic - potential alleviation by selenium. Environ Res 2015,

376

140, 205-213.

377

7.

Oono, Y.; Yazawa, T.; Kawahara, Y.; Kanamori, H.; Kobayashi, F.; Sasaki, H.;

NTP National toxicology program, tenth report on carcinogens; Department of

JECFA, Safety evaluation of certain food additives and contaminants. World

Baba, H.; Tsuneyama, K.; Yazaki, M.; Nagata, K.; Minamisaka, T.; Tsuda, T.;

Moulis, J. M.; Thevenod, F., New perspectives in cadmium toxicity: an 18


Page 18 of 37

Page 19 of 37


378

introduction. Biometals 2010, 23, 763-768.

379

8.

380

report of the Joint FAO/WHO Expert Committee on Food Additives. World Health

381

Organization: Geneva, 2004; p 1-128.

382

9.

383

Zhu, Y. G.; Li, G.; Zhao, F. J.; McGrath, S.; Villada, A.; Sommella, A.; De Silva, P. M.

384

C. S.; Brammer, H.; Dasgupta, T.; Islam, M. R., Variation in rice cadmium related to

385

human exposure. Environ Sci Technol 2013, 47, 5613-5618.

386

10. Arao, T.; Maejima, Y.; Baba, K., Uptake of aromatic arsenicals from soil

387

contaminated with diphenylarsinic acid by rice. Environ Sci Technol 2009, 43,

388

1097-101.

389

11. Cao, F. B.; Ahmed, I. M.; Zheng, W. T.; Zhang, G. P.; Wu, F. B., Genotypic and

390

environmental variation of heavy metal concentrations in rice grains. J Food Agric

391

Environ 2013, 11, 718-724.

392

12. Ye, X. X.; Ma, Y. B.; Sun, B., Influence of soil type and genotype on Cd

393

bioavailability and uptake by rice and implications for food safety. J Environ

394

Sci-China 2012, 24, 1647-1654.

395

13. Zhou, H.; Zeng, M.; Zhou, X.; Liao, B. H.; Peng, P. Q.; Hu, M.; Zhu, W.; Wu, Y.

396

J.; Zou, Z. J., Heavy metal translocation and accumulation in iron plaques and plant

397

tissues for 32 hybrid rice (Oryza sativa L.) cultivars. Plant Soil 2015, 386, 317-329.

398

14. Chen, D.; Guo, H.; Li, R. Y.; Li, L. Q.; Pan, G. X.; Chang, A.; Joseph, S., Low

399

uptake affinity cultivars with biochar to tackle Cd-tainted rice - A field study over four

FAO/WHO, Evaluation of certain food additives and contaminants: sixty-first

Meharg, A. A.; Norton, G.; Deacon, C.; Williams, P.; Adomako, E. E.; Price, A.;

19



400

rice seasons in Hunan, China. Sci Total Environ 2016, 541, 1489-1498.

401

15. Huang, Y.; Sun, C. X.; Min, J.; Chen, Y. L.; Tong, C.; Bao, J. S., Association

402

mapping of quantitative trait loci for mineral element contents in whole grain rice

403

(Oryza sativa L.). J Agr Food Chem 2015, 63, 10885-10892.

404

16. Uraguchi, S.; Kamiya, T.; Clemens, S.; Fujiwara, T., Characterization of OsLCT1,

405

a cadmium transporter from indica rice (Oryza sativa L.). Physiol Plantarum 2014,

406

151, 339-347.

407

17. Perrier, F.; Yan, B.; Candaudap, F.; Pokrovsky, O. S.; Gourdain, E.; Meleard, B.;

408

Bussière, S.; Coriou, C.; Robert, T.; Nguyen, C., Variability in grain cadmium

409

concentration among durum wheat cultivars: impact of aboveground biomass

410

partitioning. Plant Soil 2016, 404, 307-320.

411

18. Vergine, M.; Aprile, A.; Sabella, E.; Genga, A.; Siciliano, M.; Rampino, P.;

412

Lenucci, M. S.; Luvisi, A.; De, B. L., Cadmium concentration in grains of durum

413

wheat (Triticum turgidum L. subsp. durum). J Agr Food Chem 2017, 65, 6240-6246.

414

19. Liu, J. G.; Qian, M.; Cai, G. L.; Yang, J. C.; Zhu, Q. S., Uptake and translocation

415

of Cd in different rice cultivars and the relation with Cd accumulation in rice grain. J

416

Hazard Mater 2007, 143, 443-447.

417

20. Sasaki, A.; Yamaji, N.; Yokosho, K.; Ma, J. F., Nramp5 is a major transporter

418

responsible for manganese and cadmium uptake in rice. Plant Cell 2012, 24,

419

2155-2167.

420

21. Yang, M.; Zhang, Y. Y.; Zhang, L. J.; Hu, J. T.; Zhang, X.; Lu, K.; Dong, H. X.;

421

Wang, D. J.; Zhao, F. J.; Huang, C. F.; Lian, X. M., OsNRAMP5 contributes to 20


Page 20 of 37

Page 21 of 37


422

manganese translocation and distribution in rice shoots. J Exp Bot 2014, 65,

423

4849-4861.

424

22. Miyadate, H.; Adachi, S.; Hiraizumi, A.; Tezuka, K.; Nakazawa, N.; Kawamoto,

425

T.; Katou, K.; Kodama, I.; Sakurai, K.; Takahashi, H.; Satoh-Nagasawa, N.; Watanabe,

426

A.; Fujimura, T.; Akagi, H., OsHMA3, a P-1B-type of ATPase affects root-to-shoot

427

cadmium translocation in rice by mediating efflux into vacuoles. New Phytol 2011,

428

189, 190-199.

429

23. Takahashi, R.; Ishimaru, Y.; Shimo, H.; Ogo, Y.; Senoura, T.; Nishizawa, N. K.;

430

Nakanishi, H., The OsHMA2 transporter is involved in root-to-shoot translocation of

431

Zn and Cd in rice. Plant Cell Environ 2012, 35, 1948-1957.

432

24. Feng, X. M.; Han, L.; Chao, D. Y.; Liu, Y.; Zhang, Y. J.; Wang, R. G.; Guo, J. K.;

433

Feng, R. W.; Xu, Y. M.; Ding, Y. Z.; Huang, B. Y.; Zhang, G. L., Ionomic and

434

transcriptomic analysis provides new insight into the distribution and transport of

435

cadmium and arsenic in rice. J Hazard Mater 2017, 331, 246-256.

436

25. Hoshikawa, B. K., The growing rice plant : an anatomical monograph. Nosan

437

Gyoson Bunka Kyokai: Tokyo, 1989.

438

26. Yamaji, N.; Ma, J. F., The node, a hub for mineral nutrient distribution in

439

graminaceous plants. Trends Plant Sci 2014, 19, 556-563.

440

27. Fujimaki, S.; Suzui, N.; Ishioka, N. S.; Kawachi, N.; Ito, S.; Chino, M.;

441

Nakamura, S., Tracing cadmium from culture to cpikelet: noninvasive imaging and

442

quantitative characterization of absorption, transport, and accumulation of cadmium in

443

an intact rice plant. Plant Physiol 2010, 152, 1796-1806. 21



444

28. Yamaguchi, N.; Ishikawa, S.; Abe, T.; Baba, K.; Arao, T.; Terada, Y., Role of the

445

node in controlling traffic of cadmium, zinc, and manganese in rice. J Exp Bot 2012,

446

63, 2729-2737.

447

29. Uraguchi, S.; Kamiya, T.; Sakamoto, T.; Kasai, K.; Sato, Y.; Nagamura, Y.;

448

Yoshida, A.; Kyozuka, J.; Ishikawa, S.; Fujiwara, T., Low-affinity cation transporter

449

(OsLCT1) regulates cadmium transport into rice grains. P Natl Acad Sci USA 2011,

450

108, 20959-20964.

451

30. Alexander, P. D.; Alloway, B. J.; Dourado, A. M., Genotypic variations in the

452

accumulation of Cd, Cu, Pb and Zn exhibited by six commonly grown vegetables.

453

Environ Pollut 2006, 144, 736-745.

454

31. Chen, Y.; Moore, K. L.; Miller, A. J.; McGrath, S. P.; Ma, J. F.; Zhao, F. J., The

455

role of nodes in arsenic storage and distribution in rice. J Exp Bot 2015, 66,

456

3717-3724.

457

32. Lu, R. K., Analytical methods of agricultural chemistry in soil. China

458

Agricultural Scientech Press: Beijing, China, 2000 (in Chinese).

459

33. Liu, J. G.; Qu, P.; Zhang, W.; Dong, Y.; Li, L.; Wang, M. X., Variations among

460

rice cultivars in subcellular distribution of Cd: The relationship between translocation

461

and grain accumulation. Environ Exp Bot 2014, 107, 25-31.

462

34. Ishimaru, Y.; Takahashi, R.; Bashir, K.; Shimo, H.; Senoura, T.; Sugimoto, K.;

463

Ono, K.; Yano, M.; Ishikawa, S.; Arao, T.; Nakanishi, H.; Nishizawa, N. K.,

464

Characterizing the role of rice NRAMP5 in Manganese, Iron and Cadmium Transport.

465

Sci Rep-Uk 2012, 2, 286. 22


Page 22 of 37

Page 23 of 37


466

35. Cheng, H.; Wang, M. Y.; Wong, M. H.; Ye, Z. H., Does radial oxygen loss and

467

iron plaque formation on roots alter Cd and Pb uptake and distribution in rice plant

468

tissues? Plant Soil 2014, 375, 137-148.

469

36. Wang, M. Y.; Chen, A. K.; Wong, M. H.; Qiu, R. L.; Cheng, H.; Ye, Z. H.,

470

Cadmium accumulation in and tolerance of rice (Oryza sativa L.) varieties with

471

different rates of radial oxygen loss. Environ Pollut 2011, 159, 1730-1736.

472

37. Sasaki, A.; Yamaji, N.; Ma, J. F., Overexpression of OsHMA3 enhances Cd

473

tolerance and expression of Zn transporter genes in rice. J Exp Bot 2014, 65,

474

6013-6021.

475

38. Satoh-Nagasawa, N.; Mori, M.; Nakazawa, N.; Kawamoto, T.; Nagato, Y.;

476

Sakurai, K.; Takahashi, H.; Watanabe, A.; Akagi, H., Mutations in rice (Oryza sativa

477

L.) heavy metal ATPase 2 (OsHMA2) restrict the translocation of zinc and cadmium.

478

Plant Cell Physiol 2012, 53, 213-224.

479

39. Ishikawa, S.; Ishimaru, Y.; Igura, M.; Kuramata, M.; Abe, T.; Senoura, T.; Hase,

480

Y.; Arao, T.; Nishizawa, N. K.; Nakanishi, H., Ion-beam irradiation, gene

481

identification, and marker-assisted breeding in the development of low-cadmium rice.

482

P Natl Acad Sci USA 2012, 109, 19166-19171.

483

40. Uraguchi, S.; Mori, S.; Kuramata, M.; Kawasaki, A.; Arao, T.; Ishikawa, S.,

484

Root-to-shoot Cd translocation via the xylem is the major process determining shoot

485

and grain cadmium accumulation in rice. J Exp Bot 2009, 60, 2677-2688.

486

41. Tanaka, K.; Fujimaki, S.; Fujiwara, T.; Yoneyama, T.; Hayashi, H., Quantitative

487

estimation of the contribution of the phloem in cadmium transport to grains in rice 23



488

plants (Oryza sativa L.). Soil Sci Plant Nutr 2007, 53, 72-77.

489

42. Ishikawa, S.; Suzui, N.; Ito-Tanabata, S.; Ishii, S.; Igura, M.; Abe, T.; Kuramata,

490

M.; Kawachi, N.; Fujimaki, S., Real-time imaging and analysis of differences in

491

cadmium dynamics in rice cultivars (Oryza sativa L.) using positron-emitting Cd-107

492

tracer. Bmc Plant Biol 2011, 11, 172.

493

43. Rodda, M. S.; Li, G.; Reid, R. J., The timing of grain Cd accumulation in rice

494

plants: the relative importance of remobilisation within the plant and root Cd uptake

495

post-flowering. Plant Soil 2011, 347, 105-114.

496

44. Yoneyama, T.; Gosho, T.; Kato, M.; Goto, S.; Hayashi, H., Xylem and phloem

497

transport of Cd, Zn and Fe into the grains of rice plants (Oryza sativa L.) grown in

498

continuously flooded Cd-contaminated soil. Soil Sci Plant Nutr 2010, 56, 445-453.

499

45. Yu, L. H.; Umeda, M.; Liu, J. Y.; Zhao, N. M.; Uchimiya, H., A novel MT gene of

500

rice plants is strongly expressed in the node portion of the stem. Gene 1998, 206,

501

29-35.

502

46. Kobayashi, N. I.; Tanoi, K.; Hirose, A.; Nakanishi, T. M., Characterization of

503

rapid intervascular transport of cadmium in rice stem by radioisotope imaging. J Exp

504

Bot 2013, 64, 507-517.

505

24


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Figure Captions Figure 1. Image showing the location of each sampled section in a rice plant Figure 2. Concentrations of Cd in the root, shoot, and brown rice of twelve rice cultivars. Values are given as the means with standard deviations, as shown by the vertical bars (n = 3). Bars with the same letter(s) indicate no significant differences among the different cultivars at P < 0.05. Figure 3. Concentrations of Cd in the nodes and internodes of rice cultivars TYJ1, NJ9108, YY2640, NJ52, and ZXY146. The values are given as the means with standard deviations, as shown by the vertical bars (n = 3). Figure 4. Translocation characteristics of Cd in rice. TFroot–brown

rice:

translocation

factor from the root to the brown rice, TFroot–shoot: translocation factor from the root to the shoot, TEIN4–IN1: translocation efficiency from the fourth internode to the first internode. Values are given as the means with standard deviations, as shown by the vertical bars (n = 3). Bars with the same letter(s) indicate no significant differences among the different cultivars at P < 0.05. Figure 5. Vascular bundle (VB) systems in the junction of the flag leaf and the first node (JFLN) of NJ9108, NJ52 and ZXY146; scale bars = 500 µm (a). Selected area by the dotted line, scale bars = 100 µm (b). Three different VB phases are shown as follows: the enlarged VB (EVB, yellow), transit VB (TVB, red) and diffuse VB (DVB, purple). Figure 6. Linear correlation between the Cd concentration in the brown rice and in the root of the twelve rice cultivars (a). Linear correlation between the Cd 25



concentration in brown rice and the translocation efficiency of Cd from internode 4 to internode 1 in the cultivars TYJ1, NJ9108, YY2640, NJ52, and ZXY146 (b). The dashed lines indicate 95% confidence intervals. Figure 7. Linear relationship between the concentration of Cd in brown rice and the concentration of Cd in the first internode (a), TFN1–IN1 and TFN1–FL (b) in the twelve cultivars. Linear relationship between the concentration of Cd in brown rice and the concentration of Cd in the flag leaf in the cultivars with similar shoot Cd values (c). The dashed lines indicate 95% confidence intervals.

26


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Page 27 of 37


Table 1 The name, code, subspecies, growth time, culm length and 1000-grain weight of rice cultivars in the present study

Cultivar name

Code

Growth time

Culm length

1000-grain

(day)

(cm)

weight (g)

Subspecies

Nanjing52

NJ52

Japonica inbred

156

101.1

28.1

Nanjing9108

NJ9108

Japonica inbred

153

96.4

26.4

Yangyujing2

YYJ2

Japonica inbred

156

99.8

28.2

Ningjing4

NJ4

Japonica inbred

155

99.1

25

Sujing9

SJ9

Japonica inbred

163

99.1

26.4

Zhendao18

ZD18

Japonica inbred

161

98.6

26.3

Wuyunjing27

WYJ27

Japonica inbred

145

92.4

26.4

Yongyou2640

YY2640

Japonica hybrid

135

103.0

24.3

Tongyoujing1

TYJ1

Japonica hybrid

160

120.5

26.0

Cliangyou4418

CLY4418

Indica hybrid

135

116.5

23.5

Ningxianyou8

NXY8

Indica hybrid

138

123.4

25.9

Zhenxianyou146

ZXY146

Indica hybrid

146

122.0

28.1

Source: China Rice Data Center (http://www.ricedata.cn/index.htm)

27



Page 28 of 37

Table 2 TFNn–INna and TFNn–Lnb in the cultivars with similar shoot Cd values TYJ1

NJ9108

YY2640

NJ52

ZXY146

TFN4–IN4

0.41±0.02 a

0.31±0.03 b

0.32±0.03 b

0.32±0.03 b

0.38±0.03 a

TFN3–IN3

0.21±0.01 c

0.25±0.02 b

0.28±0.02 a

0.24±0.02 b

0.28±0.02 a

TFN2–IN2

0.15±0.01 b

0.15±0.01 b

0.15±0.01 b

0.20±0.02 a

0.20±0.01 a

TFN1–IN1

0.03±0.01 e

0.04±0.01 d

0.05±0.01 c

0.06±0.01 b

0.08±0.01 a

TFN4–L4

0.23±0.02 d

0.26±0.02 c

0.49±0.06 a

0.16±0.02 e

0.30±0.02 b

TFN3–L3

0.10±0.01 e

0.21±0.02 c

0.62±0.04 a

0.18±0.01 d

0.35±0.03 b

TFN2–L2

0.08±0.01 c

0.18±0.02 d

0.28±0.02 b

0.21±0.02 d

0.31±0.01 a

TFN1-FL

0.15±0.02 a

0.15±0.01 a

0.12±0.01 b

0.11±0.01 b

0.09±0.01 c

a

TFNn–INn: translocation factor of Cd from Nn to INn (n = 1, 2, 3 or 4)

b

TFNn–Ln: translocation factor of Cd from Nn to Ln (n = 1, 2, 3 or 4)

28


Page 29 of 37


Table 3 Area of VBs in the central part of the junction of the flag leaf and the first node, and its correlation coefficients with TFN1-IN1 TVBa (mm2)

DVBb (mm2)

Total VBsc (mm2)

NJ9108

0.65±0.02 b

0.04±0.01 c

0.69±0.03c

NJ52

0.84±0.03 a

0.29±0.03 b

1.13±0.06b

ZXY146

0.84±0.02 a

0.53±0.02 a

1.38±0.04a

r between TFN1-IN1 and each VBd

0.775*

0.934**

0.909**

r between BR-Cd and each VB

0.705*

0.888**

0.934**

a

TVB: transit vascular bundle;

bundles;

d

b

DVB: diffuse vascular bundle;

c

VBs: vascular

r between TFN1-IN1 and each VB: Correlation coefficients (r) between the

translocation factor (the first node to the first internode) and each VB (n = 9, *P < 0.05 and **P < 0.01)

29



For Table of Contents Only

30


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Figure 1. Image showing the location of each sampled section in a rice plant 160x203mm (300 x 300 DPI)



Concentrations of Cd in the root, shoot, and brown rice of twelve rice cultivars. Values are given as the means with standard deviations, as shown by the vertical bars (n = 3). Bars with the same letter(s) indicate no significant differences among the different cultivars at P < 0.05. 128x106mm (300 x 300 DPI)


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Concentrations of Cd in the nodes and internodes of rice cultivars TYJ1, NJ9108, YY2640, NJ52, and ZXY146. The values are given as the means with standard deviations, as shown by the vertical bars (n = 3). 121x85mm (300 x 300 DPI)



Translocation characteristics of Cd in rice. TFroot–brown rice: translocation factor from the root to the brown rice, TFroot–shoot: translocation factor from the root to the shoot, TEIN4–IN1: translocation efficiency from the fourth internode to the first internode. Values are given as the means with standard deviations, as shown by the vertical bars (n = 3). Bars with the same letter(s) indicate no significant differences among the different cultivars at P < 0.05. 121x100mm (300 x 300 DPI)


Page 34 of 37

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Vascular bundle (VB) systems in the junction of the flag leaf and the first node (JFLN) of NJ9108, NJ52 and ZXY146; scale bars = 500 µm (a). Selected area by the dotted line, scale bars = 100 µm (b). Three different VB phases are shown as follows: the enlarged VB (EVB, yellow), transit VB (TVB, red) and diffuse VB (DVB, purple). 202x206mm (300 x 300 DPI)



Linear correlation between the Cd concentration in the brown rice and in the root of the twelve rice cultivars (a). Linear correlation between the Cd concentration in brown rice and the translocation efficiency of Cd from internode 4 to internode 1 in the cultivars TYJ1, NJ9108, YY2640, NJ52, and ZXY146 (b). The dashed lines indicate 95% confidence intervals. 123x49mm (300 x 300 DPI)


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Linear relationship between the concentration of Cd in brown rice and the concentration of Cd in the first internode (a), TFN1–IN1 and TFN1–FL (b) in the twelve cultivars. Linear relationship between the concentration of Cd in brown rice and the concentration of Cd in the flag leaf in the cultivars with similar shoot Cd values (c). The dashed lines indicate 95% confidence intervals. 124x38mm (300 x 300 DPI)


(Oryza sativa L.) Cultivars - ACS Publications - American Chemical

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