Role Of Cell Wall For Aluminum Resistance And Toxicity Mechanisms In Plants

Aluminum(Al) toxicity is one of the most significant factors limiting plant growth and crop production in acid soils.Although Al-induced organic acid anions secretion has been proven as one of the most important mechanisms of Al resistance,in recent years, evidence supporting the hypothesis that the root cell wall plays an important role in Al detoxification is accumulating.Al inhibits the root elongation within few hours,but the mechanism of Al-induced inhibition of root growth is still not well understood.In the present study,we investigated the role of cell wall pectic polysaccharides in excluding Al from buckwheat root apex.We compared the organic acid anions secretion in buckwheat,wheat and Cassia tora L.,and a new Al resistant mechanism was proposed that organic acid anions protected cell wall from aluminum binding.To investigate the mechanism of Al inhibition root elongation,we use pectin monoclonal antibodies to study the effect of Al treatment on pectin distribution and cell wall property of maize root.Finally we examined the expression patterns of different reporter genes in the arabidopsis root tip under Al stress.The results were as follows:1.Role of cell wall for Al resistance in buckwheatFour buckwheat cultivars different in Al resistance were selected in this study.The Al contents in root apices of Al-resistant cultivars were lower than that in Al-sensitive cultivars,so the exclusion mechanism was involved here.PG treatment,which is an organic acid inhibitor,increased the Al contents in root apices of all cultivars,but the sensitive cultivar increased more,suggested that organic acids secretion can not fully explain the different Al tolerance.Although Al treatment increased pectin content in root apices of all cultivars,it was significantly lower in Al-resistant cv.Jiangxi than in other three cultivars irrespective of with or without Al treatments. Al contents in root sections were positively correlated to the pectin content. Localization of pectin by immunofluoresence revealed that Al-sensitive cv.Shanxi had a higher proportion of low-methylated pectin and a lower proportion of high-methylated pectin than Al-resistant cv.Jiangxi.The PME activity was lower in Jiangxi than Shanxi.We found the same phenomenon in rice,indicated that whether organic acid was secreted the degree of pectin methylation is related to Al resistance. The kinetics study with root cell wall extracted from 0-5 mm section showed that the total amount of Al adsorbed was higher and the desorption rate was lower in Shanxi than in Jiangxi.All the above results satisfactorily explain the difference in Al resistance between these two cultivars,which could not be explained by oxalate secretion.Thus we concluded that cell wall pectin content,PME activity,pectin de-methylation,cell wall binding capacity to Al contribute to Al resistance in buckwheat.2.Organic acid protect cell wall from binding AlAluminum-induced secretion of organic acids from the root apex has been demonstrated to be one major Al resistance mechanism in plants.However,whether the organic acid concentration is high enough to detoxify Al in the growth medium is frequently questioned.The genotypes of Al-resistant wheat,Cassia tora L.and buckwheat secrete malate,citrate and oxalate,respectively.In the present study we found that at a 35%inhibition of root elongation,the Al activities in the solution were 10,20,and 50μM with the corresponding at the rates of 15,20 and 21 nmol/cm2 per 12 h,respectively,for the above three plant species.When exogenous organic acids were added to ameliorate Al toxicity,twofold and eightfold higher oxalate and malate concentrations were required to produce the equal effect by citrate.Based on this calculation,the mounts of malate,citrate,and oxalate exudation should be in this proportion 4:1:5,it is different between the real exudation and theoretic secretion.So we propose that there are other roles of organic acids in Al resistance.After the root apical cell walls were isolated and preincubated in 1 mM malate,oxalate or citrate solution overnight,the total amount of Al adsorbed to the cell walls all decreased significantly to a similar level,implying that these organic acids own an equal ability to protect the cell walls from binding Al.These findings suggest that protection of cell walls from binding Al by organic acids may contribute significantly to Al resistance. 3.Perturbed distribution of HG epitopes is involved in the Al-induced root growth inhibition in maizeAl inhibits root elongaion within few hours,but the mechanism is still not well known. Here we use pectin monoclonal antibodies(JIM5,JIM7) to investigate the effect of Al treatment on pectin polysaccharide in maize root.In the absence of Al,JIM5 epitope was present around the cell wall with higher fluorescence intensity at cell corners lining the intercellular spaces,and JIM7 epitope was present throughout the cell wall. However,a 3 h treatment with 50μM Al produced 10%root growth inhibition in both cultivars and caused the disappearance of fluorescence in middle lamella of both epitopes.Prolonged Al treatment(24 h) with 50%root growth inhibition in B73,an Al-sensitive cultivar,resulted in faint and irregular distribution of both epitopes.At a 50%Al-induced root growth inhibition in Nongda3138(100μM Al,9 h),an Al-resistant cultivar,the distribution of HG epitopes was also restricted to the lining of intercellular spaces.Altered distribution of both epitopes was also observed when exposure of roots to 50μM LaCl3 for 24 h,which resulted in 40%of root growth inhibition.Changes in HG distribution and root growth inhibition were highly correlated.Washing by citrate to remove off the most Al in the root tip had no effect on the HG distribution.These results indicating that Al-induced perturbed distribution of HG epitopes is possibly involved in the Al-induced root growth inhibition in maize.4.Al effect the expression of reporter genes in arabidopsis root tipsAl stress inhibits root elongation,but what parts of cells in the root apex are influenced and whether auxin distribution is affected are still unclear.Here we tested the effect of Al stress on the expression of different cell and auxin reporter genes.The enhancer trap line J3411,in which GFP is expressed in the columella and lateral root cap,exposure to Al decreased the GFP expression in the columella cells,but had no effect in the lateral root cap,implying that the function of columella cells had been damaged,this can satisfactory explain that under Al stress generally root growth of primary roots is inhibited while the growth of lateral roots is enhanced.To investigate the impact of Al treatment on the quiet center,we examined the expression of a QC-specific promoter QC25 in the root tip,we found that the GUS activity had no difference with or without Al treatment,indicating that Al had no influence on quiet center cells,which is in accordance with the suggestion that Al mainly affects cell elongation but not cell division.Al treatment can change the expression pattern of DR5-GUS which is an auxin-response gene in the Arabidopsis root tips,indicated that Al treatment changed the distribution of auxin.