| The continuous increase in atmospheric carbon dioxide(CO2)certainly impacts on the growth and physiological metabolism of plants.As a result,plant tissue nitrogen and protein concentrations decrease,which consequently affects the nutritional quality and economic value of forage.Nitrogen(N)is the second most important nutrient for plant growth and is also the basis for photosynthesis and the production of stored proteins in plants.Exogenous N addition can improve soil N availability and promote the uptake and accumulation of N nutrients in plants.However,the mechanism for the effect of N addition on forage growth and its nitrogen nutrients under elevated CO2 concentration is not clear.In this study,the Agropyron mongolicum,the dominant herb in the desert steppe of Ningxia,was selected.The experiment was conducted focusing on atmospheric CO2 concentration as the main plot with a split-plot experimental design,using the open-top chambers(OTC)to simulate both elevated atmospheric CO2 concentrations(eCO2:800 ± 20μmol·mol-1)and natural ambient concentrations(aCO2:400 ± 20μmol·mol-1),with potted plants supplemented by the NH4NO3(four treatments:0(N0,CK),1.2(N1.2,low N),3.6(N3.6,medium N)and 10.8(N10.8,high N)g N m-2 yr-1)as subplot,for a total of 8 treatments.This study investigated the growth,physiological and nitrogen uptake characteristics of A.mongolicum under elevated CO2 concentration and N addition,and combined with statistical analysis and omics techniques to ultimately reveal mechanisms underlying the effects of N addition on growth,nitrogen uptake,and metabolism of A.mongolicum under elevated CO2 concentration.The main findings are as follows:(1)Moderate N additions promoted leaves growth and roots development,and increased the nitrogen uptake and biomass accumulation of the A.mongolicum under elevated CO2 concentration.Compared with the control(N0),medium N addition(3.6 g N m-2 yr-1)in eCO2 increased the total biomass and aboveground nitrogen uptake in A.mongolicum by 93.0%and 214.0%.N addition increased plant height,tiller number,leaf width,leaf length,leaf thickness,leaf area and leaf dry weight,and also significantly enhanced total root length,root surface area,root dry density.and root nitrogen absorption capacity.In addition,it also promoted the activities of nitrate reductase,glutamine synthetase,glutamate synthetase and glutamate dehydrogenase,while decreased the content of nitrate nitrogen/total nitrogen in roots and leaves,and soluble sugar and starch content in leaves,and above which contributing to the improvement of the biomass and nitrogen uptake in A.mongolicum.(2)Moderate N addition increased the photosynthesis of A.mongolicum under elevated CO2 concentration.The net photosynthetic rate in medium N addition(3.6 g N m-2 yr-1)of the eCO2 was the highest,and increased by 32.1%compared with N0 treatment.Nitrogen addition mainly enhanced the capture of light energy by increasing leaf nitrogen content and chlorophyll content;increases the PSII activity,photochemical efficiency and light saturation point to ensure that the captured light energy is used more efficiently for photosynthesis and improved its carboxylation capacity by enhancing the initial carboxylation rate,maximum carboxylation rate and maximum electron transfer rate,and thus improved photosynthetic capacity and promoted its growth and dry matter accumulation.(3)Elevated CO2 concentration and N addition changed the physicochemical properties and microbial community composition of the rhizosphere soil of A.mongolicum.With the nitrogen addition increasing,soil total phosphorus,available phosphorus,and pH showed a decreasing trend,while soil total nitrogen,nitrate nitrogen,microbial biomass carbon,microbial biomass nitrogen and microbial biomass phosphorus showed a gradually increasing trend.Soil organic carbon and ammonium nitrogen varied slightly,with no significant differences among treatments.The fungal Chao1 and Shannon indices showed an increasing trend,however,the bacterial Shannon index and Chao1 index shown an upward and downward trend,respectively.Elevated CO2 and N addition had a greater effect on the bacterial community.The abundances of Acidobacteria,Proteobacteria,Planctomycetes,and Rokubacteria decreased significantly with increasing N addition,and the abundances of Nitrospirae and Patescibacteria increased.However,there was no significant difference on the abundance of soil fungi at the phylum level under different treatments.(4)N addition strengthened the mutual-feedback relationship between microorganisms and soil,which significantly affected the nutrient content of plants.The results of redundancy analysis and hierarchical partitioning showed that soil pH,total nitrogen,available phosphorus,nitrate nitrogen,microbial biomass carbon,microbial biomass nitrogen,and microbial biomass phosphorus were the key driving changes in bacterial communities,and soil nitrate nitrogen and pH explained 28.1%and 27.8%of the community variation,respectively.Soil nitrate nitrogen,microbial biomass nitrogen,and microbial biomass phosphorus were the key factors controlling the variation in fungal communities,with microbial biomass nitrogen explaining 74.7%of the variation.Structural equation modeling(SEM)indicated that the interaction of soil physicochemical properties and microbial communities explained 78.0%of the variation in plant nutrients,as soil physicochemical properties had the greatest impact on total plant nutrients,followed by fungal diversity,bacterial community composition,and bacterial diversity.(5)N addition activated the expression of more genes involved in photosynthesis,carbon metabolism and amino acid biosynthesis.Transcriptome analysis showed that 223(149 up-regulated,74 downregulated)and 454(268 up-regulated,186 down-regulated)differentially expressed genes(DGEs)were identified in leaves and roots of A.mongolicum in middle nitrogen treatment under aCO2(ACN3.6),respectively;1116(518 up-regulated,598 down-regulated)and 455(268 up-regulated,186 downregulated)DGEs were identified in control treatment under eCO2(ECN0),respectively;1583(767 upregulated,816 down-regulated)and 2102(892 up-regulated,1210 down-regulated)DGEs were identified in middle nitrogen treatment under eCO2(ECN3.6),respectively.Compared with ACN3.6 and ECN0 treatment,ECN3.6 treatment obtained the most DGEs,which mainly enriched in photosynthesis,carbon metabolism,and amino acid biosynthesis.(6)N addition changed the accumulation of related metabolites in carbon metabolism,starch and sucrose metabolism and amino acid biosynthesis pathway under elevated CO2 concentration.A total of 733 metabolites were identified by ultra-high performance liquid chromatography-mass spectrometry(UPLC-ESI-MS/MS).123(18 up-regulated,105 down-regulated)and 107(24 up-regulated and 83 downregulated)differential metabolites(DEMs)were identified in roots and leaves under ACN3.6 treatment;72 DEMs were identified in ECN0 treatment(33 up-regulated,39 down-regulated)and 82(32 upregulated,50 down-regulated)differential metabolites;193(29 up-regulated,164 down-regulated)and 156(65 up-regulated,91 down-regulated)DEMs were identified under ECN3.6 treatment,respectively.ECN3.6 treatment obviously obtained more DEMs than ACN3.6 and ECN0 treatment.KEGG enrichment analysis showed that DEMs were mainly involved in biosynthesis pathways between carbon metabolism,starch,and sucrose metabolism and amino acid under ECHN treatment.(7)Transcription proteomics and metabolomics joint analysis revealed that N addition inhibited the accumulation of carbohydrates(glucose,fructose,trehalose)to synthesize amino acids and their derivatives using more carbon skeletons,thereby transferring to the N metabolic pathway,promoting more amino acids(glutamine,phenylalanine,methionine,cysteine,serine,valine,Lysine,Nacetylglutamic acid,and arginine)synthesis,and consequently increasing the nitrogen nutrient content under elevated CO2. |
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