Wheat natural resistance-associated macrophage protein 3 (NRAMP3) are manganese (Mn) transporters that can also transport unwanted cadmium (Cd). However, their roles in regulating grain Cd concentration are unknown. Here, we functionally characterised TpNRAMP3-7A and TpNRAMP3-7B cloned from dwarf Polish wheat (Triticum polonicum L., AABB) in them of their expression patterns, transcript localisations, metal transport activities, and associated phenotypes. Both TpNRAMP3-7A and TpNRAMP3-7B were expressed in the epidermis, endodermis, and xylem parenchyma cells of roots, the xylem parenchyma cells and phloem region of nodes and leaf sheaths, and the phloem region of leaf blades. Knockout of TpNRAMP3-7A and/or TpNRAMP3-7B not only limited grain Cd concentration, but also reduced Cd uptake, root-to-shoot translocation, and shoot-to-grain distribution. They also limited grain Mn concentration by inhibiting shoot-to-grain Mn distribution when grown in the field (high-Mn concentration), and decreased Mn uptake and root-to-shoot translocation under low-Mn stress. Position 192F in TpNRAMP3 was the core amino acid determining Cd and Mn transport activity. These results provide a valuable guide and target gene for limiting Cd concentration in wheat grains.

Cadmium (Cd) accumulation in wheat grains occurs via three key physiological processes: root uptake, root-to-shoot translocation and/or shoot Cd distribution. Wheat ZIP transporters are hypothesized to involved in these pathways, their functional roles remain poorly characterized. Here, we identified and functionally characterized TpZIP3-2A from dwarf Polish wheat (DPW, Triticum polonicum L., AABB), which mediates Cd, Zn, Fe and Co transport. Expression pattern and subcellular localization analysis revealed that TpZIP3-2A encoding a plasma membrane protein, was mainly expressed in roots, flag leaves, peleas, lemmas and grains of DPW, which was growth stage-dependent. Its transcript in roots was up-regulated by Zn or Fe deficiency and Co or Cd supplementation. Expression of TpZIP3-2A in yeast increased the sensitivities and concentrations of Cd (by 104.84 %), Zn (by 36.12 %) and Co (by 81.87 %). The TpZIP3-2A-overexpressing lines of Arabidopsis showed sensitivity to Cd or Co stress and tolerance to Zn or Fe deficiency, and had higher root and shoot concentrations of Cd (61.17 % and 125.82 %), Co (27.65 % and 88.81 %), Zn (36.87 % and 70.04 %) and Fe (46.02 % in shoots), as well as root-to-shoot translocation factors (24.27 %-54.50 %). However, Tpzip3-2a (a mutation of TpZIP3-2A lacking the first transmembrane domain) had no ability to transport Cd, Zn and Fe. The study provides new insights into the molecular mechanisms regulating Cd uptake and translocation in wheat and offers potential applications for breeding low-Cd accumulating wheat cultivars.

BACKGROUND:Cadmium (Cd) is a toxic heavy metal for all organisms. Increasing of wheat grain accumulates Cd posing a serious risk to human health. Thus, reducing grain Cd concentration of wheat is urgently required for food security and human health. Here, we found a wheat yellow stripe-like protein 15 (YSL15-6B) governs grain Cd concentration. METHODS:The expression pattern, subcellular localization, Cd transport activity and Cd accumulation in mutant and overexpressing lines of wheat YSL15-6B were analyzed. RESULTS:TpYSL15-6B, cloned from Dwarf Polish wheat (Triticum polonicum L. 2n = 4x = 28, AABB), was mainly expressed in roots and leaves. Its protein was localized at the endoplasmic reticulum and plasma membrane in protoplast. Expression of TpYSL15 in yeast increased Cd concentration under Cd-NA stress. Loss-of-function of TtYSL15-6B in 'Kronos' increased Cd uptake, root-to-shoot Cd translocation, and grain Cd concentration. Meanwhile, Ttysl15-6B mutant line exhibited up-regulation of TtNRAMP5 and TtHMA2, and down-regulation of TtZIP1 when compared with the wide type. Overexpression of TpYSL15-6B in rice caused Cd exporting from roots, and limited root-to-shoot Cd translocation and grain Cd concentration. TpYSL15-6B-overexpressing lines showed up-regulation of OsZIP1 and OsABCG36, and down-regulation of OsIRT1 and OsNRAMP2 when compared with the wide type ZH11. CONCLUSION:wheat YSL15-6B governs Cd export from plant. These results provide a new gene and insight for limiting grain Cd concentration in wheat and the physiological pathway of Cd transport.

Wheat easily absorbs Cadmium (Cd) from soils and accumulates in its grains. Nitrogen (N) fertilizer management has been used to limit grain Cd concentrations in wheat, but the effects of compound fertilizers (NPK-N) are unclear. Here, three years’ of field experiments were conducted to evaluate the effects of NPK-N on grain Cd concentration and yield of 50 wheat cultivars. A high-Cd accumulating cultivar Chuanmai 68 (CM68) and a low-Cd accumulating Mianmai 902 (MM902) were further used to reveal the physiological and molecular mechanisms of the Cd transport pathway. The results showed that NPK-N application significantly reduced grain Cd accumulation in all 50 wheat cultivars compared to amide N fertilize (urea-N) application. MM902 showed lower Cd uptake before anthesis, higher Cd export from plant during grain filling, and lower grain Cd concentration than CM68. Meanwhile, NPK-N application significantly reduced grain Cd concentration in CM68 (39.70

Wheat (Triticum aestivum L.) is one of the most important crops worldwide and a major source of human cadmium (Cd) intake. Limiting grain Cd concentration (Gr_Cd_Conc) in wheat is necessary to ensure food safety. However, the genetic factors associated with Cd uptake, translocation and distribution and Gr_Cd_Conc in wheat are poorly understood. Here, we mapped quantitative trait loci (QTLs) for Gr_Cd_Conc and its related transport pathway using a recombinant inbred line (RIL) population derived from 2 Polish wheat varieties (RIL_DT; dwarf Polish wheat [DPW] and tall Polish wheat [TPW]). We identified 29 novel major QTLs for grain and tissue Cd concentration; 14 novel major QTLs for Cd uptake, translocation, and distribution; and 27 major QTLs for agronomic traits. We also analyzed the pleiotropy of these QTLs. Six novel QTLs (QGr_Cd_Conc-1A, QGr_Cd_Conc-3A, QGr_Cd_Conc-4B, QGr_Cd_Conc-5B, QGr_Cd_Conc-6A, and QGr_Cd_Conc-7A) for Gr_Cd_Conc explained 8.16% to 17.02% of the phenotypic variation. QGr_Cd_Conc-3A, QGr_Cd_Conc-6A, and QGr_Cd_Conc-7A pleiotropically regulated Cd transport; 3 other QTLs were organ-specific for Gr_Cd_Conc. We fine-mapped the locus of QGr_Cd_Conc-4B and identified the candidate gene as Cation/Ca exchanger 2 (TpCCX2-4B), which was differentially expressed in DPW and TPW. It encodes an endoplasmic reticulum membrane/plasma membrane-localized Cd efflux transporter in yeast. Overexpression of TpCCX2-4B reduced Gr_Cd_Conc in rice. The average Gr_Cd_Conc was significantly lower in TpCCX2-4BDPW genotypes than in TpCCX2-4BTPW genotypes of the RIL_DT population and 2 other natural populations, based on a Kompetitive allele-specific PCR marker derived from the different promoter sequences between TpCCX2-4BDPW and TpCCX2-4BTPW. Our study reveals the genetic mechanism of Cd accumulation in wheat and provides valuable resources for genetic improvement of low-Cd-accumulating wheat cultivars. Genetic mapping of Polish wheat reveals candidate genes associated with grain cadmium accumulation, a complex trait mainly controlled by root cadmium uptake and shoot cadmium sequestration.

Grain copper (Cu) concentrations represent a qualitative trait mainly controlled by genetic factors, which may differ between wheat varieties from the Sichuan Basin of China and other areas. However, the differences are poorly understood. Here, we investigated the grain Cu concentration in a remaining heterozygous line population derived from a multiparental recombinant inbred line. The grain Cu concentration varied from 4.25 to 13.44 mg/kg and 3.32 to 7.74 mg/kg over a two-year investigation, and the broad-sense heritability was 0.67. Bulked-segregation analysis revealed three quantitative trait loci on chromosomes 2A (QGr_Cu_Conc-2A), 2B (QGr_Cu_Conc-2B), and 4D (QGr_Cu_Conc-4D). QGr_Cu_Conc-2B is a novel locus, which was further narrowed between KASP-52.32 and KASP-56.57 with an interval of 52.32–56.57 Mb, explaining 17.10

Wheat is one of major sources of human cadmium (Cd) intake. Reducing the grain Cd concentrations in wheat is urgently required to ensure food security and human health. In this study, we performed a field experiment at Wenjiang experimental field of Sichuan Agricultural University (Chengdu, China) to reveal the effects of FeCl3 and Fe2(SO4)3 on reducing grain Cd concentrations in dwarf Polish wheat (Triticum polonicum L., 2n = 4x = 28, AABB). Soil application of FeCl3 and Fe2(SO4)3 (0.04 M Fe3+/m2) significantly reduced grain Cd concentration in DPW at maturity by 19.04

Wheat grown in cadmium (Cd)-contaminated soils easily accumulates more Cd in edible parts than the Chinese safety limit (0.1 mg/kg). FeCl3 and Fe2(SO4)3 have been used to extract Cd from Cd-contaminated soils. Thus, we hypothesized that FeCl3 and Fe2(SO4)3, used as iron (Fe) fertilizers, can reduce Cd uptake and accumulation in wheat. Here, a hydroponic experiment was performed with three FeCl3 and Fe2(SO4)3 concentrations under 80 μM CdCl2 stress on dwarf Polish wheat (Triticum polonicum L., 2n = 4x = 28, AABB) seedlings. Compared with Fe deficiency, FeCl3 and Fe2(SO4)3 additions competitively reduced Cd concentrations. The reductions were not associated with changes in dry weight and root morphological parameters. FeCl3 and Fe2(SO4)3 additions reduced Cd concentrations in the following order from smallest to largest reduction: 25 μM Fe2(SO4)3 < 200 μM FeCl3 < 50 μM FeCl3 < 100 μM Fe2(SO4)3. Investigation of subcellular distributions showed that the four Fe fertilizers differentially reduced Cd binding in the root cell walls and enhanced root sucrose and trehalose. Cd chemical form analysis revealed that Fe fertilizer addition also differentially reduced root FE, FW, and FNaCl. Transcriptomic analysis revealed that addition of FeCl3 and Fe2(SO4)3 differentially up-regulated several genes that hydrolyze cell wall polysaccharides and metal transporter genes for Cd uptake (IRT1 and CAX19) and export (ZIP1, ABCG11, ABCG14, ABCG28, ABCG37, ABCG44, and ABCG48) reducing Cd uptake and accumulation. Our results demonstrated that FeCl3 and Fe2(SO4)3 can reduce Cd accumulation in wheat, and 50 μM FeCl3 is the most effective treatment.

This study investigated the effects of manganese (Mn) and copper (Cu) on dwarf Polish wheat under cadmium (Cd) stress by evaluating plant growth, Cd uptake, translocation, accumulation, subcellular distribution, and chemical forms, and the expression of genes participating in cell wall synthesis, metal chelation, and metal transport. Compared with the control, Mn deficiency and Cu deficiency increased Cd uptake and accumulation in roots, and Cd levels in root cell wall and soluble fractions, but inhibited Cd translocation to shoots. Mn addition reduced Cd uptake and accumulation in roots, and Cd level in root soluble fraction. Cu addition did not affect Cd uptake and accumulation in roots, while it caused a decrease and an increase of Cd levels in root cell wall and soluble fractions, respectively. The main Cd chemical forms (water-soluble Cd, pectates and protein integrated Cd, and undissolved Cd phosphate) in roots were differently changed. Furthermore, all treatments distinctly regulated several core genes that control the main component of root cell walls. Several Cd absorber (COPT, HIPP, NRAMP, and IRT) and exporter genes (ABCB, ABCG, ZIP, CAX, OPT, and YSL) were differently regulated to mediate Cd uptake, translocation, and accumulation. Overall, Mn and Cu differently influenced Cd uptake and accumulation; Mn addition is an effective treatment for reducing Cd accumulation in wheat.

Abstract As the valuable germplasm resources of wheat genetic improvement, tetraploid wheat has several reduced height (Rht) genes that reduced plant height and enhanced harvest index. Rht22 discovered from Jianyangailanmai (JAM, Ttiticum turgidum L., 2n = 4x = 28, AABB), significantly increases in spikelet number per spike, while its accurate position was still unknown. In this study, a high-density genetic map was constructed by specific-length amplified fragment sequencing (SLAF-seq) in F7 RIL_DJ population derived from a cross between dwarf Polish wheat (DPW, Ttiticum polonicum L., 2n = 4x = 28, AABB) and JAM. Two plant height loci, Qph.sicau-4B and Qph.sicau-7A, were mapped on 4BS and 7AS, respectively. Qph.sicau-7A on 7AS with the interval of 0.33-4.46Mb was recommended as the candidate region of Rht22. Fine mapping confirmed and narrowed Rht22 on 7AS between xbag295.s53 and xb295.191 in three different populations. The physical region ranged from 0.53 to 1.48 Mb and included 18 candidate genes. Transcriptome analysis of two pairs of NILs found 135 differentially expressed genes (DEGs) associated with semi-dwarfism. Out of 135 DEGs, 83 annotated DEGs participated in hormones synthesis and signal transduction, cell wall composition, DNA replication, microtubules and phragmoplast arrays were significant down-regulation in semi-dwarf line. Thus, Rht22 disturbs these processes to impair the cell proliferation, and then reduces cell number of internode to cause semi-dwarfism in JAM.

Kernel size, one of the traits that determine wheat yield, is controlled by multiple quantitative trait loci. Polish wheat (Triticum polonicum) has elongated and plump kernel and is a valuable material for breeding high-yielding wheat cultivars. However, genes or loci determining kernel length (KL) in Polish wheat are unknown. We identified and validated a major KL gene, KL-PW, at the P1 locus in Polish wheat. KL-PW is VRT-A2, which encodes a MIKC-type MADS-box protein (MADS55). An insertion/deletion mutation in intron 1 of VRT-A2PW led to an alternatively spliced transcript, VRT-A2PW2. Quantitative PCR analysis showed that VRT-A2PW was more highly expressed in developing seeds than was VRT-A2Ailanmai. Brassinosteroid (BR) sensitivity experiment and the expression of BR-related genes indicated that VRT-A2PW functions as a positive regulator of BR responses. VRT-A2PW significantly increased KL of wheat. These findings not only reveal the molecular basis of KL-PW in controlling KL, but also provide a valuable genetic resource for increasing kernel size in wheat.

Rht22 was fine mapped in the interval of 0.53–1.48 Mb on 7AS, which reduces cell number of internode to cause semi-dwarfism in Jianyangailanmai. As a valuable germplasm resource for wheat genetic improvement, tetraploid wheat has several reduced height (Rht) and enhanced harvest index genes. Rht22, discovered in Jianyangailanmai (JAM, Triticum turgidum L., 2n = 4x = 28, AABB), significantly increases the spikelet number per spike, but its accurate chromosomal position is still unknown. In this study, a high-density genetic map was constructed using specific-length amplified fragment sequencing in an F7 RIL_DJ population, which was derived from a cross between dwarf Polish wheat (T. polonicum L., 2n = 4x = 28, AABB) and JAM. Two plant height loci, Qph.sicau-4B and Qph.sicau-7A, were mapped on chromosomes 4BS and 7AS, respectively. Qph.sicau-7A was mapped to the 0.33-4.46 Mb interval on 7AS and likely represents the candidate region of Rht22. Fine mapping confirmed and narrowed Rht22 on chromosome arm 7AS between Xbag295.s53 and Xb295.191 in three different populations. The physical region ranged from 0.53 to 1.48 Mb and included 18 candidate genes. Transcriptome analysis of two pairs of near-isogenic lines revealed that 135 differentially expressed genes (DEGs) were associated with semi-dwarfism. Of these, the expression of 83 annotated DEGs involved in hormones synthesis and signal transduction, cell wall composition, DNA replication, microtubule and phragmoplast arrays was significantly down-regulated in the semi-dwarf line. Therefore, Rht22 causes semi-dwarfism in JAM by disrupting these cellular processes, which impairs cell proliferation and reduces internode cell number.

Ethylene regulates plant abiotic stress responses and tolerances, and ethylene-insensitive3 (EIN3)/EIN3-like (EIL) proteins are the key components of ethylene signal transduction.Although the functions of EIN3/EIL proteins in response to abiotic stresses have been investigated in model plants, little is known in non-model plants, including mulberry (Morus L.), which is an economically important perennial woody plant.We functionally characterized a gene encoding an EIN3-like protein from mulberry, designated as MnEIL3.A quantitative real-time PCR analysis demonstrated that the expression of MnEIL3 could be induced in roots and shoot by salt and drought stresses.Arabidopsis overexpressing MnEIL3 exhibited an enhanced tolerance to salt and drought stresses.MnEIL3 overexpression in Arabidopsis significantly upregulated the transcript abundances of ethylene biosynthetic genes.Furthermore, MnEIL3 enhanced the activities of the MnACO1 and MnACS1 promoters, which respond to salt and drought stresses.Thus, MnEIL3 may play important roles in tolerance to abiotic stresses and the expression of ethylene biosynthetic genes.

The uptake and translocation of non-essential heavy metals in plant are always through metal transporters for essential micronutrient transport, such as NRAMP (Natural Resistance-Associated Macrophage Protein). NRAMPs from different species exhibit different biological functions, although their sequences are highly identical. In the present study, a NRAMP6 was isolated from Ailanmai (Triticum turgidum L. ssp. turgidum). TtNRAMP6, localized on chromosome 3B, was mainly expressed in roots, followed by other tissues varied with different growth stages. At the seedling stage, TtNRAMP6 was significantly regulated by Cd stress in roots, but not by the deficiency of Zn, Fe or Mg. Subcellular localization analysis indicated that TtNRAMP6 encoded a plasma membrane protein. Expressing-TtNRAMP6 significantly enhanced the Cd concentration in yeast, and increased the Cd sensitivity. Meanwhile, overexpression of TtNRAMP6 also increased the Cd concentration in roots, stems, leaves and the whole plant of Arabidopsis, which indicated that overexpression of TtNRAMP6 enhanced the Cd accumulation. Thus, genetic manipulation of TtNRAMP6 may reduce the uptake of Cd from external solution to wheat, finally protecting the safety of wheat food.

Amphidiploids generated by distant hybridization are commonly used as genetic bridge to transfer desirable genes from wild wheat species into cultivated wheat. This method is typically used to enhance the resistance of wheat to biotic or abiotic stresses, and to increase crop yield and quality. Tetraploid Thinopyrum elongatum exhibits strong adaptability, resistance to stripe rust and Fusarium head blight, and tolerance to salt, drought, and cold.