Effects Of Nitrate Transporters NRT1.1 And NRT1.2 On Zinc And Cadmium Accumulation In Plants

Zinc(Zn)and cadmium(Cd)contaminations in soil are serious problems for the safety of agricultural production.Scientific management of nitrate nitrogen fertilizer to minimize the entry of Zn or Cd into the crops are recognized as important strategies for ensuring the safety of food crops produced in contaminated soils.NRT1.1 and NRT1.2 are the major nitrate transporters in plant roots.However,the effects of NRT1.1 and NRT1.2 on regulating Zn and Cd accumulation in plants are still unclear.Therefore,ecotype Columbia-0 of Arabidopsis and nrtl.1 mutants(nrtl.1 and chll.5)were used in the present study for revealing the effect of NRT1.1 on Zn accumulation in plant by plate and hydroponic culture methods.On the other hand,studies have shown that exogenous abscisic acid(ABA)could inhibit Cd accumulation in plants.Considering that NRT1.2 is an ABA-importing transporter,wild-type Arabidopsis(Col-0)and NRT1.2-knockout mutant nrtl.2 were also employed to investigate the role of NRT1.2 in synergistic transport of ABA in inhibiting Cd accumulation.The main research includes two parts as follows:(1)By scanning ion-selective electrode technique SIET,it was found that 250 ?M Zn treatment significantly promoted NO3-uptake rate in the meristematic zone,the elongation zone and the mature zone of Col-0 plant roots by 218%,253%and 38%,respectively,compared with the Zn-free control;Zn stress also significantly induced the enhancement of GFP fluorescence at the root tip of the transgenic line pNRT1.1::NRT1.1-GFP,and the enhancement of GUS staining at the root tip of the transgenic line pNRTl.1::NRT1.1-GUS.These findings indicated that the Zn-induced NO3-uptake was significantly related to NRT1.1.Then we measured the Zn concentrations of roots and shoots in Zn-stressed Col-0,nrtl.l and chll.5.In plate culture,Zn concentration in Col-0 plants were,respectively,82%and 83%higher(in roots),and 61%and 55%(in shoots)higher than nrtl.1 and chll.5;In aqueous culture,the Zn concentration in Col-0 plants were,respectively,31%and 64%higher(in roots),and 24%and 43%(in shoots)higher than nrtl.1 and chll.5.The above results suggested that inhibiting NRT1.1 activity could significantly reduce Zn concentration in plants.In addition,biomass and photosynthetic indexes(chlorophyll content and Y(II))within two culture methods showed that inhibiting NRT 1.1 activity could alleviate Zn induced-growth stress and photosynthetic inhibition.NO3--Zn coexistence rotation and interval rotation schemes were designed to test whether the NRT 1.1 regulated Zn accumulation in plants depending on NO3-or not.The results showed that the difference of Zn concentration between Col-0 and nrtl.1 mutants(nrtl.1 and chll.5)in NO3--Zn coexistence treatment was significantly higher than that in NO3--Zn interval treatment,indicating that NRT 1.1 regulated Zn accumulation in plants related to the co-transport of nitrate.In conclusion,Zn stress may induce the expression of NRT1.1 in a certain way and promote NO3-uptake and co-transport Zn2+ into plants.Inhibiting NRT1.1 activity could reduce Zn concentration and alleviate Zn-induced growth stress and photosynthetic inhibition.(2)In hydroponic culture,ABA application could significantly alleviate Cd-induced(10 ?M Cd)growth inhibition and photosynthetic damage in Co1-0 arabidopsis,while its effect on nrtl.2 mutant(nrtl.2)was obviously weakened.Furthermore,ABA application could increase ABA concentrations in the shoots and roots of Cd stressed Col-0 by 12%and 28%,respectively,whereas no significant changes were observed in nrtl.2 plants.The results indicated that NRT1.2 may be involved in the co-transport of exogenous ABA to alleviate Cd-induced growth inhibition and photosynthetic damage in plants.Based on the analysis of Cd concentration in plants,we found that the ABA application reduced the Cd concentrations of NRT1.2 overexpression transgenic plants NRT1.2ox-1 and NRT1.2ox-2,Col-0 and nrtl.2 by 77%,64%,38%and 27%(shoots)and 39%,29%,27%and 8%(roots),respectively.These results suggested that NRT1.2 may participate in the synergistic ABA transport and thus promote the inhibition of exogenous ABA in Cd accumulation in plants.In addition,ABA could inhibit the transcript levels of Cd-uptake related genes IRT1,ZIP1,ZIP4 and Nrampl in Col-0 roots.Furthermore,by compared the differences in the expression levels in NRT1.2ox-1,NRT1.2ox-2,Col-0 and nrtl.2 plants,we found that the ABA-mediated Cd uptake by NRT1.2 was mainly related to IRT1 and Nramp1.In summary,NRT1.2 mediated-ABA might inhibit IRT1 and Nrampl and thereby reducing Cd levels in plants.