Research On The Detoxification Mechanism Of Iron In Response To Robinia Pseudoacacia-rhizobium Symbiosis Under Mercury Stress

With the further strengthening of people’s concept of ecological environment protection and environmental pollution control,phytoremediation of heavy metal contaminated soil has become an inevitable trend.Plants are also stressed by heavy metals while repairing heavy metals.As one of the important methods of plant response to heavy metal stress,omics can reveal the detoxification mechanism of mercury under reasonable application.Robinia pseudoacacia is one of the tree species to control environmental pollution in China,and its planting range is widely distributed.Recent studies have found that Robinia pseudoacacia has strong enrichment ability of heavy metals（Pb,Cd,Zn,Hg,etc.）,which can be used for the remediation of soil heavy metal pollution.The research shows that iron is of great significance for phytoremediation of heavy metal pollution,and can promote or inhibit the absorption of heavy metals by plants.At present,the action mechanism of iron element in Robinia pseudoacacia is not clear.In view of the improvement of scientific research technology and methods and the improvement of the strategic value of plant microorganism in soil heavy metal remediation,the research on how the Robinia pseudoacacia-Rhizobium symbiotic acts and responds to the migration and transformation of heavy metal mercury has been paid attention to.The concentration of Fe²⁺in this study is set as 0mg·kg^-1,10mg·kg^-1,30mg·kg^-1;Hg²⁺concentration:0ng·g^-1,320ng·g^-1,32000ng·g^-1,a total of 9 treatments.This study takes the response characteristics of rhizosphere microenvironment of iron element in Robinia pseudoacacia root and the physiological changes of Robinia pseudoacacia in response to mercury stress as the research object,studies the action mechanism of iron element in Robinia pseudoacacia root and its effect under mercury stress from the level of transcriptome and metabolites,and explores the physiological and ecological change mechanism of Robinia pseudoacacia-Rhizobium symbiosis system with different mercury concentrations under the action of root iron element,To explore the response mechanism of Robinia pseudoacacia-Rhizobium symbiosis system to mercury stress under the action of iron,in order to lay a theoretical foundation for the remediation of soil heavy metal mercury by Robinia pseudoacacia-Rhizobium symbiosis system.The results are as follows:（1）Under mercury stress,the growth of Robinia pseudoacacia was further dwarfed,and the ratio of root to crown length of Robinia pseudoacacia increased by 5%-13%;The root shoot ratio of Robinia pseudoacacia increased by 8%-16%.The net photosynthetic rate,transpiration rate and stomatal conductance of Robinia pseudoacacia decreased significantly with the increase of mercury concentration（P<0.05）,and there was no significant change in intercellular CO₂concentration.In Hg-0 treatment group,compared with Fe-0 treatment,there was no significant change in Fe-10treatment,and the E and gsw of Fe-30 treatment were significantly increased by 124%and 168%;In Hg-320 treatment group,compared with Fe-0 treatment,A,E and gsw of Fe-10 treatment increased by 167%,135%and 140%respectively,and there was no significant change in Fe-30treatment;In Hg-32000 treatment group,compared with Fe-0 treatment,A,E and gsw of Fe-10treatment increased by 165%,184%and 217%respectively,and there was no significant change in Fe-30 treatment.On the whole,the increase of Fe-10 group was the largest.Mercury stress changed the ratio of chlorophyll a to chlorophyll b.Chla/chlb was about 1:1in Hg-0 treatment and 7:3 in Hg-320 and Hg-32000 treatment.In terms of total chlorophyll,Hg-0treatment was the highest,which decreased by 235%in Hg-320 treatment and 184%in Hg-32000treatment.The addition of iron did not significantly affect the chlorophyll ratio and content of Robinia pseudoacacia.（2）After Mercury enters the plant,the content of roots is the most,followed by leaves and stems.The content of mercury ion in the root of Robinia pseudoacacia increased significantly,and the content of mercury ion in the root of Robinia pseudoacacia increased to a certain extent.The effect of Fe-10 treatment is the most significant.The mercury concentration in Robinia pseudoacacia symbiosis increased with the increase of mercury concentration in the environment,which increased ROS 147%-320%in Robinia pseudoacacia leaves,enzyme activities of various antioxidant stresses by 267%-625%,and GSH cycle ratio by 259%-1010%.The addition of iron alleviated the mercury stress of Robinia pseudoacacia in varying degrees.ROS decreased by 54%-75%,the enzyme activity of antioxidant stress increased by 180%-281%again,and the GSH cycle ratio decreased by 32%-65%.（3）Under mercury stress,the growth environment of Robinia pseudoacacia seedlings deteriorated,the water content increased,the electrical conductivity increased and the p H value decreased,which increased the organic acid content of root exudates of Robinia pseudoacacia Rhizobium symbiosis,oxalic acid,citric acid and malic acid by 104,7 and 8 times respectively,and tartaric acid content increased by 201ug.In terms of the final secretion,the order is oxalic acid,tartaric acid,citric acid and malic acid.After applying a certain amount of iron,the growth environment of Robinia pseudoacacia was improved.The contents of oxalic acid,citric acid,malic acid and tartaric acid in Hg-32000 treatment decreased first and then increased,among which Fe-10treatment had the best effect,reducing about 65%,48%,26%and 57%respectively.The content of organic acid in the sand and the surrounding environment increases,resulting in an increase in the conductivity of the sand.Fe-10 treatment forms an iron film on the root surface of Robinia pseudoacacia,adsorbs more mercury ions,and the addition of iron promotes the transport of mercury to the aboveground,resulting in the decrease of mercury ion content in the root of Robinia pseudoacacia,the degree of mercury stress in the root of Robinia pseudoacacia and the secretion of low molecular organic acids.（4）Mercury stress caused 8099 up-regulated and 9710 down-regulated genes in Robinia pseudoacacia,43 up-regulated and 29 down-regulated genes in Rhizobium;Lux R family transcription regulators between Robinia pseudoacacia-rhizobium symbionts are up-regulated,and regulate plant pathogen interaction through TGA/PR family in plant hormone signal,which affects the expression of ROS,HR,defense and other related genes in pattern recognition receptor（PRRs）to varying degrees.Iron alleviated mercury stress,resulting in up-regulation of 7514 and 5751genes in Robinia pseudoacacia,up-regulation of 3 and down regulation of 49 genes in Rhizobium,changes in the regulation of Lux R family regulators between symbionts,down regulation of Lux R family transcription regulators,up regulation of activity of two-component transcription regulators,and up regulation of TGA/PR,resulting in plant pathogen interaction regulating ROS,hypersensitivity.The expression of defense related genes was up-regulated.Under mercury stress,the expression of chitin elicitor receptor kinase 1 involved in pattern recognition receptor pathway K13429（CERK1）was down-regulated,the expression of cyclic nucleotide gated channel of K05391（CNGC）was up-regulated,and the expression of K13420（FLS2）LRR like serine/threonine protein kinase FLS2 was up-regulated;When iron alleviated mercury stress,CERK1 expression was up-regulated,CNGC expression was down-regulated,and FLS2 expression remained unchanged.Therefore,we screened its regulatory genes ATH:AT3G21630,DMK:116918470,ATH:AT5G46330 as candidate genes to resist mercury stress.In addition,transcriptome analysis showed that Mer T and Mer C in Robinia pseudoacacia were involved in the transport of mercury.The combined analysis of metabolomics and transcriptomics showed that the increase of cholinergic synaptic metabolism and the expression level of photosynthetic carbon sequestration could partly explain the phenomenon of iron promoting net photosynthesis under mercury stress;The addition of iron enhanced lipoic acid metabolism,protected Robinia pseudoacacia from the toxicity of heavy metal ions,and improved the expression level of ABC transporter.The transport process of mercury ions may be mediated by ABC transporter.