Mechanism Analysis Of Aluminum Inhibiting Root Elongation By Co-repressor LUH In Arabidopsis

Aluminum(A1),the most abundant metal elements in the crust,causes a severe damage on the plant root growth in acid soil.Cell wall plays a critical role in root growth inhibition in response to A1 stress.The main components of the cell wall are pectin and hemicellulose,and pectin methyl esterification(DM)affects the ability of chelating aluminum by pectin.LUH regulates the mucilage release in Arabidopsis seeds and the luh mutant seeds show the mucilage extrusion defective phenotype and the pectin esterification is also altered.We found that the increased DM in cell wall may contribute to higher tolerance to Al stress in luh.The main conclusions are followed:(1)The higher tolerance to Al stress is correlated with the reduced Al accumulation in the luh mutant rootUsing liquid medium analysis,we found that luh mutants show aluminumtoxicity resistance.Hematoxylin staining,morin staining and aluminum content analysis all showed that roots of luh mutants accmulated less A1 compared with Col.(2)Less accumulated A1 in cell wall of the luh mutant was detectedWe analyzed the A1 accumulation in cell wall and symplast,and the results showed that the content of aluminum in the luh mutants was reduced compared with the WT control.These data suggest that resistance of luh mutants to A1 stress is due to the less accumulated Al in cell wall.(3)The increased pectin methyl esterification is consistent with the higher tolerance to Al stress in the luh mutantUsing ruthenium red staining and the phenotype of mum2&mum4 mutants analysis indicates that mucilage may be not contributed to the aluminum toxicity resistance in luh mutants.Next,we analyzed the pectin content and DM in luh mutants,and found that the DM significantly increased in luh mutants compared with the WT control,which caused the decrease of the carboxyl with negative charge as well as the aluminum binding sites.(4)LUH regulates the expression of PME46.RNA seq analysis shows that PME46 expression increase after exposure to A1 in luh mutants.PME46,as a repressor of PME,inhibits the activity of PME,and thus inhibits pectin deesterification.(5)The pme46 mutant is sensitive to A1 stress and displayed increased Al accumulation in root tips.To investigate the role of PME46 in aluminum stress,we analyse the phenotype of pme46 mutants to Al,and the pme46 mutants showed more sensitive to Al compared with Col.Consistently,hematoxylin staining,Morin staining and aluminum content analysis also showed that pme46 mutants binded more aluminum compared with wild type.(6)SLK regulates PME46 expression and controls root growth inhibition under Al stress.Previous studied showed that LUH interacting with SLK to inhibit downstream gene expression.slk2-l mutants,similar to luh mutants,showed more resistant to Al stress.Interestingly,slk2-1 mutants also have less aluminum content in root cell wall.Furthermore,PME46 expression was up-reguated in the slk2-1 mutant.These results suggest that both SLK and LUH act as repressors to inhibit PME46 expression in response to Al.This study suggests thatLUH plays an important role in aluminum tolerance through inhibiting PME46 expression together with SLK,which affect the degree of pectin methyl esterification,and thus influences the chelating Al accumulation in cell wall.