The Mechanism Of Auxin Alleviate Aluminum Toxicity In Alfalfa

Aluminium（Al）is a light metal that makes up 7%of the earth’s crust and is the third abundant element in the earth,only behind to oxygen and silicon.Al is normally not toxic to plants,however,Al is solubilized into the Al（H₂O）₆³⁺(Al³⁺)when soils become acidic（p H<5.5）.Al³⁺is very phytotoxic and is a major limiting factor for plant productivity in acidic soils.Auxin（IAA）plays an important role in plant growth and development,it regulates tissue growth and organ differentiation,controls cell elongation and division,mediates synthesis and modification of the cell wall polysaccharide,and is also involved in plant responses to environmental stresses including aluminum stress.However,limited information is available on the effect of auxin on plant response to Al³⁺stress.Alfalfa（Medicago sativa L.）,an important legume,is known as“king of grass”,but its growth was markly inhibited under Al³⁺stress.So far,the mechanisms of alfalfa response to Al³⁺toxicity has not been explored clearly.In present study,we used two alfalfa varieties（WL525,Al-resistant and WL440,Al-sensitive）with different aluminum tolerance as plant materials,and used different treatments including Al treatment(0 and 100μM L^-1 Al Cl₃),exogenous application of IAA(0 and 6 mg L^-1IAA)or application of glucose（0 and 0.05g/L Glu）,and dealing apical buds of seedlings（keep or cut apical buds）to investigate:1)The effects of Al³⁺on the changes of root tissue and cell structure;2)The effects of Al³⁺on IAA synthesis,transport and related gene expression;3)The effects of IAA on the changes of cell structure,cell arrangement,and cell wall composition of Al³⁺stressed alfalfa seedlings;4)Glucose as a carbon source of cell wall formation,the effects of glucose on the cell wall composition of Al-stressed alfalfa and Al³⁺adsorption capacity of cell wall.The main conclusions are as follows:1)Aluminum stress significantly inhibited root growth,decreased weights of root and shoot,root activities and total chlorophyll contents,but significantly increased the MDA content in leaves;Exogenous application of IAA significantly alleviated Al³⁺-induced inhibition of plant growth.2)When alfalfa was exposed to Al³⁺-stressed conditions,IAA concentrations in apical buds,root tips and shoot stems were significantly decreased;meanwhile the IAA concentration in root tips was significantly decreased in plants without apical buds in both the CK and Al treatment,decreased by 89.8%and 87.6%compared with the normol plants,respectively,indicating that the synthesis of IAA is mainly in apical buds,and the synthesis of IAA in the root tips is very limited.Al³⁺stress strongly decreased IAA synthesis in apical buds.3)The distribution of IAA fluorescence signals in root tips was disturbed in Al³⁺-stressed alfalfa,in which the highest intensity of fluorescence signals was detected in the apical meristematic zone,indicating that IAA transportation from shoot base to root tip was inhibited and the IAA distribution in root tips was disturbed.In addition,transcriptional analysis demonstrated that PIN2 and AUX1 were up-regulated in root tips exposed to Al³⁺,positively mediating polar auxin transport from the QC of root tips to the upper root,leading to a higher IAA accumulation in the epidermis and cortex of the meristematic zone.4)Aluminium stress changed the shape and structure of root tips,in which the cell shapes were deformed,cell arrangement was disordered,root tips were abused,cell and vacuole were thickened,lengths of the meristematic zone were shorted,and cell senescence was accelerated.The disordered cell arrangement was similar with IAA fluorescence signal disturbance,meantimes,the collose content in shoot and root tips increased,indicating that the destroy of cell structure inhibited IAA transport from root base to root tips.Exogenous application of IAA markedly alleviated the Al³⁺-induced inhibition of root growth by increasing IAA accumulation in root tips and recovering the damaged cell structure of root tips.5)The Al contents in shoots,root tips and cell wall（CW）were significantly lower in WL525 than WL440.The WL525 had higher Al accumulation in pectin,and WL440 had higher Al accumulation in hemicellulose 1（HC1）compared to each other.Meanwhile,Al³⁺stress also significantly increased the number of negatively charged functional groups（-COOH,C-C and C-OH）in pectin,hemicellulose and cellulose,especially in the treatment of Al stressed seedling without apcal buds,which caused the increases of Al³⁺adsorption ability and Al accumulations in the cell wall.Exogenous application of IAA significantly decreased pectin methylesterase activity（PME）,reduced the number of functional groups（-COOH,C-C and C-OH）and Al³⁺adsorption ability of cell wall in two alfalfa varieties,and thus reduced Al contents in root tips,CW and pectin fractions under Al³⁺condition;in addition,exogenous application of IAA increased plasma membrane H⁺-ATPase activity and acidification of rhizosphere.These results indicated that IAA induced reduction of Al accumulation in cell wall could be associated with the decrease of Al³⁺bonding ability in pectin through increase H⁺secretion to rhizosphere and balance H⁺level between internal and external cells.6)Al³⁺stress caused a significant increase in the contents of main components of pectin,including Arabinose（Ara）,Galactose acid（Gal A）,Rhamnose galacturonic acid chitosan I（RG-I）and Homogalacturonan（HGA）,and the content of Xylose（Xyl）which is main components of HC1.Exogenous application of IAA markedly reduced the contents of Ara,Xyl,Gal A and RG-I in Al-stressed alfalfa.Glucose（Glu）as a carbon source for the formation of monosaccharides of pectin and hemicellulos.Exogenous addition of 0.05g/L Glu significantly decreased the content of HC1,but increased the number of negatively charged functional groups（-OH,-COOH,C-C and C-OH）in cell wall,as result,the Al contents in the root tips and cell walls were decreased,in which the Al contents in HC1 were significantly decreased,but the Al contents in pectins were significantly increased.The decreased Al contents in root tips and cell walls greatly alleviated Al³⁺toxicity,and protected the structure and shape of root tips.