Study On The Physiological And Transcriptomical Response Of Ectomycorrhizal Pinus Massoniana To Manganese Stress

Heavy-metal mining industry destructed ecolocial system severely and produces millions of tonnes of mine wastes.Consequently,a huge amount of heavy metals was released into the evironments during the processing progress and caused severe heavy-metal pollution.Mycorrhizal-plants remediation has the characteristics of environmental friendly and sustainability,and is the most potential strategy for the ecological restoration of heavy-metal polluted soils.In this study,we inoculated the Pinus massoniana seedlings with a Mn-tolerant ectomycorrhizal(ECM)funal islates-Pisolithus tinctorius(Pt-104).We performed a pot experiment,the mycorrhizal and nonmycorrhizal P.massoniana seedlings were exposed to different level of Mn SO4(0,1 m M m M,15 m M,30 m M,50 m M).We montitored the seedlings growth during the treatment period.After 45 d growth,we measured the phothsynthetic parameters of seedlings and determined several physiological parameters,and finally performed transcriptomic sequencing on seedling roots and needles,respectively.In this study,we try to characterize the response of P.massoniana ECM seedling exposed to Mn stress at physiological and molecular levels,and provide theorial and technical basis for remediation of soil Mn pollution with ECM seedlings.The main results were as follows:(1)After Mn stress,the plant height and biomass of mycorrhizal and non-mycorrhizal P massoniana decreased with the increase of Mn concentration,compared to the reference,mycorrhizal inoculation alleviated the inhibition effect of Mn on the seedling growth.Under 30 m M Mn stress,non-mycorrhizal seedling's height and biomass decreased by 24% and 29%,while mycorrhizal seedling's height and biomass fell by 13% and 18%,respectively.(2)Under 30 m M Mn2+ stress,mycorrhizal inoculation markedly increased net photosynthetic rate,stomatal conductance,decrease intercellular CO2 concentration and lower transpiration rate of the seedlings,and therefore alleviate the destruction effect on photosynthetic system by Mn stress.Meanwhile,mycorrhizal inoculation significantly improved the nutrients(N,P,K,Ca and Mg)absorption.However,Mn were also accumulated more in mycorrhizal seedlings and particullary in the root system,and therefore reduce the translocation of Mn to shoots.Mycorrhizal inoculation increased the activity of APX,SOD and CAT activities,which were increased 74.4%,41.6% and 116.3% in needles,and 55.5%,69.8% and 25.7% in roots,respectively.Under 30 m M Mn2+ stress,O2-content in the needles and roots of non-mycorrhizal seedlings increased by 48% and 51%,and MDA content increased by 33.4% and 36.8%,respectively.In contrast,O2-content in mycorrhizal seedlings increased 30% and 30%,MDA increased by14.5% and 19.6%.Similarly,soluble protein in the needles and roots of non-mycorrhizal seedlings increased by 13.4% and 9.3%,respectively,and free proline content increased by 37.3% and 32.2%,respectively.In contrast,in mycorrhizal needles and roots,soluble protein increased by 39.3% and 27.4% and free proline content increased by 156.0% and 144.7% respectively.Therefore,mycorrhizal symbiosis can comprehensively alleviate the toxic effect on seedling caused by Mn stress.(3)RNA-sequencing of the roots and needles of non-mycorrhizated and mycorrhizal seedlings under Mn treatment were performed on Illumina Hiseq2500 platform.In total,273,600 unigenes of P.massoniana seedlings were generated based on de novo assembly.By comparing gene expression profile in mycorrhizal seedlings between Mn stress and non-Mn treatment,a total of 1,605 differentially expressed genes were found in needles,among which 931 genes were up-regulated and 674 genes were down-regulated.In addition,a total of 1,411 genes were differentially expressed in roots,among which 1,111 genes were up-regulated and 300 genes were down-regulated.GO and Pathway analysis revealed that genes involved in cell swallowed,plant hormone signaling,plant-pathogen interaction pathways,sucrose metabolism and starch were specially regulated when exposure to Mn stress.In addition,based on the released P.tinctorius genome,differential expressed genes in mycorrhizal roots were also analysed.In total,330 genes belonging to P.tinctorius were differential expressed between the Mn treatment and non-treatment,among which 194 genes was up-regulated and 136 genes were down regulated.By Go enrichment and Pathway analysis,ECM fungal genes involved in metabolic pathway and antibiotic synthesis were regulated under Mn stress.Thus,gene express of EMF and host plant-P.massoniana transcriptomes were co-regulated when exposure to Mn stress.