Regulation Of NAC Transcription Factors On Drought Stress Response And Dwarfing Character In Malus

Apple(Malus domestica Borkh.)is one of the most widely cultivated fruit trees in the world.Both the yield and the cultivating area of apple are at the forefront.The Loess Plateau of China is one of the dominant producing areas for apple production.Located in the arid or semi-arid areas,facing light precipitation and water scarcity problems,drought has been the major limiting factor affecting the yield and quality of apple fruit in this area.In production,breeding and applying new apple resources which have strong drought tolerance is an important way to alleviate water stress.Dwarfing and high-density planting pattern in apple production is a desirable method which can accelerate early bearing,increase yield and benefit for mechanized farming and management,so that it can cut cost and improve economic returns.However,the inappropriate using of rootstocks or backward cultivation and management techniques or other problems has limited the advantages of dwarfing and high-density planting pattern in China,hindering the development of this planting pattern in apple production.Breeding new dwarfing apple varieties offers more choices for dwarfing and high-density planting in apple production.And it is one of the major methods to stimulate the development of modern apple production.In this study,the functions of related apple NAC transcription factors genes were analyzed by genetic transformation or other methods.The results provided some basis for breeding drought tolerant apple varieties or dwarf apple rootstocks.The main results are shown as follows:1.In a genome-wide scale,175 NAC transcription factors were identified in apple.These NAC transcription factors consist of a large protein family.The members of these apple NAC proteins can be divided into 26 sub-families.And the members within a same sub-family tend to have similar protein sequences and gene structures.2.Fifteen NAC genes were identified to be abiotic stress related genes in apple.The expression patterns of 15 apple NAC gene were analyzed under drought,salt,cold and high temperature conditions by real-time quantitative PCR methods.These results showed that these genes were responsive to at least one kind of abiotic stress,indicating they are abiotic stresses related genes.Five NAC genes from the above 15 genes were cloned from ‘Golden delicious' apple.And their functional analysis showed that all those 5 NAC proteins localized in the cell nucleuses and 4 of them had transcriptional activation functions.3.A dwarfing and drought related NAC gene,Md NAC1,was cloned from ‘Golden Delicious' apple,whose function was analyzed in overexpressing lines of GL-3 apple.Md NAC1 conferred dwarfing phenotype in transgenic apple lines.Their reduction in plant size is manifested by shorter,thinner stems and roots,and smaller leaf areas.These transgenic lines also have shorter internodes and fewer cells in stems.Levels of endogenous abscisic acid(ABA)and brassinosteroid(BR)were lower in the transgenic lines,and the expression changed significantly for some genes involved in the biosynthesis of those two hormones.All of these findings demonstrated that Md NAC1 has a role in plant dwarfism,probably by regulating the content and signal pathways of ABA and BR.4.Overexpression of Md NAC1 enhanced drought tolerance in transgenic apple lines.The tolerance to natural drought and long-term drought stresses was analyzed in Md NAC1 overexpressing lines for self-rooted and the grafted seedings.Under drought conditions,the transgenic lines showed higher relative water content,lower relative electrolyte leakage,less accumulation of MDA,and higher photosynthetic pigments(chlorophyll a,chlorophyll b and carotenoids).And the photosynthetic rate was higher in the transgenic lines under drought conditions.Under long-term drought conditions,the relative growth was higher in Md NAC1 overexpressing lines than that in GL-3 wild-type plants.In addition,the levels of reactive oxygen species(H2O2 and O2-)were lower in transgenic lines comparing with in WT plants,however,the activities of relatived antioxidant enzymes(SOD,POD and CAT)were higher in transgenic lines under natural drought conditions.All these above results indicated that overexpression of Md NAC1 enhanced the plant ROS-scavenging ability and alleviated the damage suffered from drought stress.5.Apple Md SNAC1 is a abiotic stress responsive NAC transcription factor gene.It was strongly induced by drought,salt,cold and high temperature stresses.The expression of apple Md SNAC1 in tomato(Lycopersicon esculentum Mill.)plants improved the tolerance to drought stress.Under drought conditions,the Md SNAC1 transgenic lines showed higher relative relative water content and lower relative electrotype leakage in leaves;and the photosynthetic rate and photosynthetic pigments were all higher.On the contrary,the accumulation of MDA and reactive oxygen species(H2O2 and O2-)were lower,but the corresponding activities of antioxidant enzymes(SOD,POD,and CAT)were higher in transgenic lines.The tomato plants expressing Md SNAC1 showed enhanced tolerance to drought stress,and better physiological or biochemical parameters,indicating the positive regulatory role of apple Md SNAC1 in response to drought stress.