Resistance Physiology And Proteomic Analysis Of Apple Rootstocks Under Mixed Salt-alkaline Stress

The salt-alkalization of soil,threatening the normal growth,metabolism of plants,is increasingly serious in the world,with seriously restricting the apple industry in the northwest loess plateau.In apple production,salt-resistance apple rootstock is commonly used to alleviate saline-alkali stress,also it is important to reveal the mechanism of rootstock responding to stress.The resistant mechanism of rootstocks is extremely importantly revealed for apple production.The apple rootstocks Malus halliana and Malus baccata were taken as materials.Pot experiments were conducted to irrigate different concentrations of salt/alkali mixed solution,such as 0mmol·L^-1,50mmol·L^-1,100mmol·L^-1,150mmol·L^-1 and 200mmol·L^-11 NaCl+NaHCO₃?amount of substance ratio 1:1 mixed?.The physiological indices of Malus halliana and Malus baccata were measured to select the suitable concentration of salt/alkali stress for omics research.Meanwhile,the photosynthetic and fluorescence parameters of Malus halliana were determined,with observing microscopic structure of the leaves to identify the anatomical features.iTRAQ technology was used to study the differential protein expression of Malus halliana leaves on mixed saline alkali stress,and analyze the metabolic pathway.Main results were as follows:1.After 40 days of 100mmol·L^-1mixed saline alkali stress,CS still maintained normal growth,and SDZ showed obvious salt damage phenotype.Under 150 and 200 mmol·L^-1saline-alkali stress,the leaves of CS were withered,and the SDZ'leaves all dry and even death.With the prolongation of stress time,the chlorophyll content of CS decreased slowly,and the SDZ'decreased rapidly after treated 20 days.The total organic acid content,soluble sugar content,proline content and relative electrical conductivity of CS and SDZ increased with the stress time prolonging,and the increased degree of CS under 100 mmol·L^-1saline-alkali stress was significantly lower than that of 150 and 200 mmol·L^-1 stress.SOD of CS and SDZ increased firstly and then decreased,while POD continuously increased,and the two enzyme activity was the highest under 100 mmol·L^-1 saline-alkali stress.Principal components analysis was carried out on 8 physiological indices under different concentrations of mixed salt-alkali stress,and the resistance of two rootstocks were comprehensively evaluated.The results showed that comprehensive score of different concentration 50,100,150 and 200 mmol·L^-1 in Malus baccata was all negative,however,the comprehensive score of high concentration 150 and 200 mmol·L^-1 in Malus halliana was negative,while 100mmol·L^-1 was positive.It indicated that the 100 mmol·L^-11 NaCl+NaHCO₃ was the critical concentration of Malus halliana.2.Under different mixed saline alkali stress,with the prolongation of time under saline-alkali stress,the Pn,Gs,Ci,Tr,F₀,Fv/Fm,ETR,?PSII and qP of CS leaves gradually decreased,while WUE,Y?NPQ?,Y?NO?and qN all increased.With the increase degree of saline-alkali stress,the length of palisade tissue became shorter and the arrangement became loose,and the sponge tissue became loose and the gap gradually became big.Under 100mmol·L^-1 saline-alkali stress,leaf thickness,upper epidermis thickness,lower epidermis thickness,palisade tissue thickness and sponge tissue thickness were the thickest,and significantly higher than those in control and other treatment groups.3.The leaves of CS were yellow,but fresh and tender,the ten-year old plants grew normally under treated 100mM NaCl+NaHCO₃ at fifth day.When treated seventh day,the leaves near the root withered and shrunk,at ninth day,the whole plants died.With the extension of time,the root activity increased first and then decreased,and the leaf water content gradually decreased,also Na⁺/K⁺ratio increased.The contents of K⁺in roots,stems and leaves all decreased gradually,and the K⁺content of roots was the highest at the same time,which was significantly higher than that in stems and leaves,and the content of K⁺was the lowest in the stems.The content of Na⁺in roots,stems and leaves increased sharply,and the content of Na⁺in roots was the highest,followed by leaves.4.100mM NaCl+NaHCO₃ stressed to fifth day,the iTRAQ technology was used to identify a total of 179 differential proteins responding to saline-alkali stress.There were 105up-regulated proteins and 74 down-regulated proteins.According to the literature,55proteases were involved in 30 pathways by GO enrichment and KEGG pathway analysis.Differential protein related enzymes participated in different metabolic pathway,such as photosynthesis,photosynthetic carbon fixation,citric acid cycle,glyoxylate cycle,glycolysis,amino sugars and nucleic acid metabolism,glucose metabolism,ascorbate and aldarate metabolism,plant hormone signal transduction,phenylpropanoid biosynthesis,protein processing in endoplasmic reticulum and ribosome protein,and so on.The percent of proteins involved in energy metabolism was the largest,was 50%,therein,the photosynthesis and photosynthetic carbon fixation related protein accounted for 11.82%,the 38.18%proteins related to carbohydrate metabolism;participate in nucleotide and amino acid metabolism was25.45%,protein synthesis and processing accounted for 6.36%and lipid metabolism was6.36%,plant hormone signaling proteins was 4.55%,and the proteins of phenylpropanoid biosynthesis accounted for 7.27%.