Moderate Production Of Betacyanins Enhances The Aluminum Tolerance Of Arabidopsis And Rice

About 40%of the cultivated land in the world is acid soil,and aluminum（Al）toxicity is one of the major factors restricting plant growth in acid soil.Al toxicity is mainly limiting the absorption of water and nutrients by roots and inhibiting the growth and development of plant roots.In response to Al toxicity,plants developed two mechanisms:external exclusion and internal detoxification:the former mainly chelates plant rhizosphere Al by secreting organic acids,while the latter mainly chelates Al and redistributes Al in cells.Betacyanins are watersoluble pigments,and they have been found that it can improve plant resistance to drought,salinity and other abiotic stresses.However,there is no report on betacyanins contribute to the tolerance of plants to Al toxicity.We created transgenic Arabidopsis thaliana and rice that can synthesize betacyanins in vivo,in order to explore the effect of endogenous betacyanins on the Al resistance of Arabidopsis and rice.The main research contents and results are as follows:（1）Under Al stress,Arabidopsis plants with an appropriate amount of betacyanins（BtOE-At）had longer root length and heavier fresh weight in aboveground and underground parts than wild type（WT）,while there was no phenotypical difference when they were grown on 1/2 MS medium,indicating that betacyanins may improve the Al tolerance of plants.Chlorophyll content decreased after Al stress,but there was no significant difference on chlorophyll content in the leaves of BtOE-At and WT.Hematoxylin staining showed that Al content in the cell wall of BtOE-At was lower than that in WT.（2）Since aluminum stress often occurs under acidic conditions,we further subjected BtOE-At and WT to acid-aluminum stress.The results showed that root length and fresh weight of aboveground and underground parts of BtOE-At were significantly higher than those of WT.The chlorophyll content of each line was lower than that of the untreated group,but there was no significant difference among these lines.3,3’-diaminobenzidine（DAB）staining was used to investigate H₂O₂ content in plants.It was found that H₂O₂ content in BtOE-At was lower than that in WT.MDA content in BtOE-At was significantly lower than that in WT.The cell wall and cytoplasm Al content of BtOE-At were lower than that of WT,revealed by hematoxylin staining and morin staining.The results suggested that betacyanins might reduce the oxidative stress of plants by reducing the content of Al in plants,so as to maintain the growth and development of plants under acid-aluminum stress.（3）ALMT1 encode malic acid transporters located in the plasma membrane of cells,which are critical for plant response to Al toxicity.It was found that the expression level of ALMT1 in BtOE-At was significantly higher than that in WT,and the expression level of its positive regulator STOP1 also up-regulated.ALS3 encodes a bacterial-type ABC transporter with 7 TMD that redistributes Al from plant tissue to relieve Al toxicity.qRT-PCR showed that the ALS3 expression level of BtOE-At was significantly lower than that of WT.Combined with the results of phenotype and Al content analysis,we preliminarily suggested that Al tolerance improvement and low Al content in root tips of BtOE-At were related to the increase of malic acid excretion.（4）Rice,one of the most important cereals worldwide,is easily affected by Al toxicity.The root elongation of rice seedlings exposed to Al toxicity will be significantly inhibited.The root length of BtOE-Os was lightly inhibited than that of WT after acid-aluminum stress.The results of hematoxylin and morin staining showed that the content of Al in cytoplasm and cell wall of BtOE-Os was lower than that of WT after acid-aluminum stress,suggesting that betacyanins may alleviate the Al toxicity of rice by preventing the absorption of Al in root tips.In this study,through a series of physiological,biochemical and molecular biological experiments,we found that BtOE-At and BtOE-Os had stronger Al toxicity resistance than WT.This may be caused by the decrease of plant root tip Al content and the decrease of plant oxidative stress.The lower content of Al in root tip of BtOE-At may be related to the substantial increase of the excretion of malic acid.In this study,by exploring the mechanism of endogenous betacyanins that enhances the Al tolerance of Arabidopsis and rice,it will provide theoretical and application supporting for the improvement of plant Al tolerance.