Study On The Roles Of Brassinosteroid On Growth And Developmemnt Of Apple Nursery Trees

In the world,trend of apple production is dwarf and high density cultivation.However,mechanism on apple tree growth and development has not been fully revealed.The researches on above theme focused on auxin(IAA),gibberellin(GA),cytokinin(CK)and abscisic acid(ABA).Brassinosteroid(BR),named as the sixth hormone,is an important plant growth regulator.However,information of BR on apple trees is unclear.Therefore,it is essential to uncover functions of BR in regulating apple trees,which will enrich its information on apple and will be of great significance to transformation of apple production mode in China.In this study,roles of BR were analyzed in apple.Using proteomics,BR-mediated key proteins and metabolic pathways which controlled growth of apple nursery tree were identified.Genome-wide identification of BR synthesis,metabolism and signal transduction genes were completed,and their potential functions were preliminarily analyzed.In addition,a novel apple micro RNA(Mdmi RLn492),which was pridected to target BR signal receptor Md BAK1 gene,was characterized.Functions of Mdmi RLn492 and Md BAK1 were revealed in transgenic Arabidopsis thaliana and apple.RNA-seq showed key predicted pathways in Md BAK1-mediated apple nursery trees' growth.The main results are described as follows:1.Roles of BR were disscussed through proteomics in apple.(1)Plant height,average internode length,leaf area,leaf and main stem fresh weight and stem diameter of apple nursery tree were significantly greater in BR-treated group than control one.(2)Proteins differentially expressed between BR treatment and control group were related to chlorophyll synthesis,photosynthesis,carbohydrate metabolism,energy metabolism(glycolysis,tricarboxylic acid cycle and respiratory electron transportion),hormone signaling,cell growth and lignin metabolism.(3)Cell length and xylem thickeness in main stem were increased in BR-treated apple;Contents of mineral elements(nitrogen,phosphorus and potassium),carbohydrates(starch,sucrose,fructose,etc.),photosynthetic indexes(Pn,Gs and Ci)and phytohormones(BR,IAA and GA)were obviously greater in BR-treated trees compared with control ones.According to the above results,a hypothesis about BR-mediated apple nursery trees' growth by regulating energy metabolism,hormone metabolism,cell growth and lignin synthesis was constructed.2.Twenty-two BR synthesis and metabolism genes were identified in apple,and their expression patterns in apple were discussed.(1)These apple genes were highly conserved in characteristic domains,chemical characteristics and gene as well as protein structures with their Arabidopsis homologous genes,respectively.Homology analysis showed above gene duplications in apple.(2)Expression level of a number of BR synthesis and metabolism genes,including Md CPDs,Md ROT3,Md CYP90D1,etc.,were regulated by BR,IAA,and temperature during root and stem enlongation of apple nursery tree.BR synthesis and metabolism genes(Md BR6OX2 and Md DWFs)showed different responses to exogenous phytohormones in Arabidopsis and apple,and some genes may have different functions in growth of main stem and root.(3)Several genes(Md DWF1 s,Md CPDs,Md DWF4-3,etc.)may also take part in shoot elongation in different apple tree varieties and grafting combinations.3.The genome-wide identification of apple BR signal transduction genes was performed,and their functions in apple nursery tree were analyzed,initiallly.(1)Gene duplication analysis showed that apple BR signaling genes occurred to amplification and evolution.Their homologous genes in Arabidopsis were identified.(2)Exogenous BR affected expression patterns of several apple BR signaling genes(Md BRI1-5,Md BSK3-8,Md BKI1-2,etc.)during stem elongation.(3)Some BR signaling genes(Md BRI1-5,Md BSK1,Md BSK4/7,etc.)were predicted to be involved in dwarf rootstock Malling 9(M9)-mediated scion dwarfing.4.Targeted relationship of Mdmi RLn492 to Md BAK1 was identified,and functions of Md BAK1 were analyzed through genetic transformation and RNA-seq technologies.(1)Mdmi RLn492 could characteristically targeted Md BAK1(Md BAK1-1).However,there were four Md BAK1 homologous genes(Md BAK1-1,Md BAK1-2,Md BAK1-3 and Md BAK1-4),which may lead to unexpected phenotypes of transgenic apple trees overexpressing Mdmi RLn492.(2)Md BAK1 improved transgenic plant height,average internode length,cell length in main stem,etc.(3)Md BAK1 may enhanced sensitivity of Arabidopsis thaliana and apple to BR.(4)Results from transcriptome indicated that Md BAK1 may promoted growth of apple tree by activating sugar as well as energy metabolism,hormone metabolism,BR as well as ethylene signal,lignin metabolism,and stress responses.In general,a putative mechanism of Md BAK1 on plant growth and development was clarified.In summary,BR played important roles in growth and development of apple nursery tree.BR may promote growth of apple nursery tree by affecting energy metabolism,cell growth and lignin metabolism.The similarities and differences of BR synthesis,metabolism and signal genes between apple and Arabidopsis thaliana were identified.The key BR synthesis,metabolism and signal genes were identified during main stem and root elongation which were mediated by BR,IAA and temperature.The target regulation of Mdmi RLn492 to Md BAK1 was clarified,and functional redundancy of Md BAK1 homologous genes was disscussed.Mechanism of Md BAK1 in promoting growth of apple nursery trees referring to sugar as well as energy,BR as well as ethylene signal,lignin metabolism and stress sensibility was preliminarily explained.