QTL Analysis And Nitrogen Regulation Of Cd Accumulation In Wheat

Cadmium（Cd）is a heavy metal element that is toxic to all living organisms.Wheat（Triticum aestivum L.2n=6x=42,AABBDD）is the staple food for the world‘s population.To date,increasing agricultural lands in wheat-producing areas of China and the world have been contaminated by Cd,which caused Cd accumulation in wheat grains.As the main source of human intake of Cd,high Cd levels in wheat grains pose potential risks to human health.Thus,limitation of the Cd concentration in wheat grains is required to food security and genetic improvement of wheat.Cd accumulation in wheat grains is mainly controlled by three pathways,including Cd uptake by the roots,Cd translocation from roots to shoots,and Cd remobilization from shoots to grains.These processes are affected by genetic factors and environmental factors.Therefore,genetics and environmental measures can be used to reduce Cd accumulation in wheat grains.For genetic measures,limited information is available on genes or QTLs related to Cd accumulation in wheat grains and other tissues,which restricts the analysis of Cd accumulation mechanism and the genetic selection of low-Cd accumulators.For environmental mesureses,nitrogen（N）management is considered to be a cost-efficient,timesaving and effective strategy to increase crop yield and decrease Cd accumulation.However,the physiological and molecular effects of different N forms on the Cd absorption,translocation,remobilization and accumulation in wheat are unclear.In this study,dwarf Polish wheat（Triticum polonicum L.,2n=4x=28,AABB,DPW）,Tall Polish wheat（TPW）,and their recombinant inbred line（RIL）population,whose Cd concentrations are different in all tissues,were used to analysis quantitative trait loci（QTL）and predict the metal transporter genes in the candidate interval.Meanwhile,the physiological and molecular responses of different N forms regulating the Cd absorption,translocation and accumulation,and the effects of ammonium nitrogen（NH₄⁺-N）addition effects on the Cd/Zn interactions in DPW seedlings were analyzed.Additionally,the Cd concentration in the grains of 45 common wheat cultivars were evaluated,and the effects of NH₄⁺-N on the Cd transport route to grains in both high-and low-Cd accumulators were drawn.The main results obtained were:1.Cd concentration in grains of Polish wheat is significantly positively correlated with the Cd concentration of other tissue.In this study,DPW×TPW RILs were used to construct a genetic map using the wheat 55K single nucleotide polymorphism（SNP）array.A total of 14 novel and major QTLs were identified,which control the Cd concentration in grains,glumes,rachises,node 1,lower leaves,lower internodes,lower nodes,and roots of Polish wheat.Among them,three QTLs for grain Cd concentration were identified.A total of 30 metal transporter encoding genes were found in the 14 major QTL,belonging to the members of ZIP（ZIP transporter）,NRAMP（Natural resistance-associated macrophage protein）,HMA(P_1B-ATPase-Heavy metal associated protein),YSL（Yellow stripe-like transporter）,CCX（Cation/calcium exchanger）,HIPP（Heavy metal-associated isoprenylated plant protein）,MRS（Magnesium transporter protein）,MTP（Metal tolerance protein）,VIT（Vacuolar iron transporter）,ACA（Calcium-transporting ATPase）,and ABC（ATP-binding Cassette transporter）.2.Different nitrogen forms differentially affect Cd uptake,translocation and accumulation in DPW seedlings.In the absence of nitrogen,addition of ammonium nitrogen（NH₄⁺-N）reduced roots Cd concentration,roots F_CW（cell wall）,F_S（soluble fraction）and F_E（ethanol-extratable）Cd concentrations,and induced the expression of four genes encoding metal transporters,including ABCG42,ZIP5,ABCB11 and HAK4（Potassium transporter）in roots,while it promoted Cd translocation to shoots.In the presence of nitrate（NO₃^--N）,addition of NH₄⁺-N increased roots Cd concentration,root F_CW and F_W（deionized water-extratable）Cd concentrations,and induced the expression of22 genes encoding metal transporters（ABC,COPT,HMA,CAX,MTP,and MRP）in roots.Thus,the effects of NH₄⁺-N addition on the uptake and accumulation of DPW seedlings were opposite in the absence of nitrogen or in the presence of NO₃^--N.Additionmally,no matter in the absence of nitrogen or in the presence of NH₄⁺-N,addition of NO₃^--N increased Cd uptake and accumulation in DPW seedlings.However,NO₃^--N greatly increased roots Cd concentration,roots F_CW,F_S,and F_WCd concentration,and lactose concentrations,and also induced the expression of more genes encoding metal transporters in roots in the presence of NH₄⁺-N,compared to the absence of nitrogen.Thus,NO₃^--N greatly increased Cd uptake and accumulation in the presence of NH₄⁺-N compared to the absence of nitrogen.3.In the presence of NO₃^--N,no matter under lack or supply of NH₄⁺-N,Zn significantly inhibited Cd uptake and accumulation in roots,but promoted Cd translocation from roots to shoots.NH₄⁺-N significantly reinforced the inhibition of Cd uptake and the promotion of Cd translocation caused by Zn.Under lack of NH₄⁺-N,Cd significantly inhibited Zn uptake and accumulation in roots,but promoted Zn translocation from roots to shoots.Under supply of NH₄⁺-N,Cd significantly inhibited Zn uptake and accumulation in roots and Zn translocation from roots to shoots.NH₄⁺-N significantly alleviated the inhibition of Zn uptake and partly reduced the promotion of Zn translocation stimulated by Cd.Under Cd treatment,supply of NH₄⁺-N significantly enhanced the Cd concentration in root F_CW and F_Sand shoot F_CW;while under Cd+Zn treatment,supply of NH₄⁺-N significantly enhanced the Cd concentration in above fractions,when compared with lack of NH₄⁺-N.Under Zn treatment,supply of NH₄⁺-N significantly enhanced the Cd concentration in all subcellular fractions of roots and shoots;while under Cd+Zn treatment,supply of NH₄⁺-N significantly enhanced the Cd concentration in root F_CW and F_COand shoot F_CW,F_CO and F_S,when compared with lack of NH₄⁺-N.Additionally,under Cd treatment,NH₄⁺-N significantly increased the Cd concentration in root F_E,F_W and F_{Na Cl},but decreased that in root F_HAc and F_HCl.Under Cd+Zn treatment,NH₄⁺-N significantly decreased the Cd concentration in root F_W and F_{Na Cl},and shoot F_{Na Cl},F_HAc and F_HCl,but increased that in shoot F_W and F_{Na Cl}.Therefore,NH₄⁺-N mediated the Cd/Zn interaction by affecting the subcellular distributions of Cd and Zn and the chemical forms of Cd.4.The Cd concentration of 45 common wheat cultivars were evaluated,and two different Cd-accumulating wheat cultivars,the high-Cd accumulator Zhoumai 18（ZM18）and the low-Cd accumulator Yunmai 51（YM51）were selected for further analysis.The effects of NH₄⁺-N on the Cd absorption,translocation,remobilization and accumulation in ZM18 and YM51 were investigated.In ZM18,NH₄⁺-N addition inhibited Cd remobilization from lower internodes,lower leaves,node 1,and internode 1 to grains during grain filling via the down-regulation of YSL,ZIP5,ZIP7,ZIP10,and HMA2,leading to decreased grain Cd content.In YM51,NH₄⁺-N addition promoted continuous Cd absorption during grain filling and direct transport to grains by up-regulated YSL,ZIP5,and COPT4,leading to increased grain Cd content.Parallelly,in ZM18,NH₄⁺-N addition reduced the grain biomass by a similar extent with the Cd content;in YM51,it increased the grain biomass by a significantly lower extent than Cd content.Thus,NH₄⁺-N addition unchanged the grain Cd concentration in ZM18,the high-Cd accumulator,and increased the grain Cd concentration in YM51,the low-Cd accumulator.These results further indicated that the addition of NH₄⁺-N in the field is not suitable for reducing the grain Cd concentration in common wheat.