Study On The Resistance Function Of Melatonin Biosynthetic Gene HIOMT、MdASMT9 In The Regulation Of Salinealkali Stress In Apples

Apple（Malus domestica Borkh.）is one of the four major cultivated fruits in the world.The Northwest Loess Plateau has the advantages of sufficient sunlight,deep soil layer,and large temperature difference between day and night,and is one of the best eugenic areas for apples.However,the soil of the Loess Plateau is dominated by Heilong soil,Malan loess and loess soil,and its Na Cl and Na HCO₃ contents are relatively high,so that the overall soil p H is8.0-8.5.It is contrary to the conditions that the slightly acidic soil is suitable for the growth of apple plants,and is not conducive to the sustainable development of the apple industry.Melatonin,an indole tryptamine with various cellular and physiological activities,is widely present in plants.It can be used as a growth regulator,which can delay senescence,reduce the accumulation of reactive oxygen species（ROS）in organisms,and improve plant resistance to abiotic and biotic stresses.The previous research of our group showed that exogenous melatonin plays a vital role in apple’s response to saline-alkali stress.In this study,we focused on exploring the functions of exogenous and endogenous melatonin synthase genes in apple plants response to saline-alkali stress.We used apple plants which overexpressed two genes related to melatonin synthesis（human oxindole-O-methyltransferase gene HIOMT and apple N-acetyl-serotonin-methyltransferase gene MdASMT9）as objects,treated with salt stress,alkali stress and salt-alkali compound stress.The function of heterologously expressed HIOMT in apple response to salt stress and alkali stress and the function of overexpressed MdASMT9 in apple response to salt stress and salt-alkali compound stress and its possible regulatory mechanism were revealed.The main findings are as follows:1.Heterogeneous expression of HIOMT in apple plants were used as the test material,and the role of heterologous expression of HIOMT in regulating the response of apple plants to salt stress was studied through hydroponic experiments.The results of the study showed that the heterologous expression of HIOMT increased the concentration of endogenous melatonin,thereby increasing the activity of antioxidant enzymes and inhibiting the accumulation of ROS.In addition,the stomatal aperture could be adjusted by regulating ABA metabolism to inhibit water loss and enhance the absorption of carbon dioxide,thereby improving the photosynthetic capacity of plants.The heterologous expression of HIOMT could also effectively stabilize the ion balance by increasing the expression of salt stress-related genes,and alleviate osmotic stress by changing the metabolism of amino acids to improve the adaptability of apple leaves to salt stress.2.Heterogeneous expression of HIOMT in apple plants were used as the test material,and the role of heterologous expression of HIOMT in regulating the response of apple plants to alkali stress was studied through hydroponic experiments.The results of the study showed that the heterologous expression of HIOMT increased the endogenous melatonin content,relieved the inhibitory effect of alkali stress on plant growth,and promoted the growth and development of apple seedling roots.Under alkaline stress,the accumulation of ROS in apple plants increased,and the transgenic lines enhanced the activities of SOD and POD,improved the antioxidant system,and alleviated the damage of chloroplasts by alkali stress and improved the photosynthetic system.In addition,the heterologous expression of HIOMT increased the organic acid content in leaves and roots of apple plants by regulating the TCA cycle and up-regulating the gene expression of enzymes related to organic acid metabolism.The roots,stems and leaf organs of the transgenic lines all maintained a low Na⁺/K⁺ratio,which was beneficial to alleviate the alkali stress.3.Overexpressing MdASMT9 in apple plants were used as the test material,the role of overexpressing MdASMT9 in regulating the response of apple plants to salt stress was studied through hydroponic experiments.The results showed that overexpression of MdASMT9enhanced the tolerance of apple plants to salt stress.MdASMT9-overexpressing apple lines improved cell membrane permeability,enhanced photosynthetic capacity,enhanced antioxidant capacity,and maintained intracellular ion homeostasis by increasing melatonin content under salt stress.4.Overexpressing MdASMT9 in apple plants were used as the test material,the role of overexpressing MdASMT9 in regulating the response of apple plants to mixed salt-alkali stress was studied through hydroponic experiments.The results showed that after 15 d of treatment under Na Cl:Na HCO₃=1:1,25 m M Na Cl and Na HCO₃,p H 8.5±0.2,the overexpression of MdASMT9 could alleviate the effect of mixed salt-alkali stress on apple plant growth.The relative electrolyte leakage（REL）and MDA of the overexpression lines under the mixed salt-alkali stress were lower than WT,but the root activity was higher than WT.The chlorophyll content and Pn of the overexpression lines were higher than WT,and the overexpression lines reduced the stress inhibition of PSⅡ,and activated the heat dissipation mechanism（Y（NPQ）and q N）to resist damage.Roots,stems,and leaf organs all maintained a low Na⁺/K⁺ratio,as well as the amino acid metabolism was changed to alleviate osmotic stress.