Identification Of AP1/FUL Subfamily Members And Functional Analysis Of PrupeFUL4 In Peach Fruit Ripening And Softening

AP1/FUL as a subfamily of MADS-box transcription factors and involves in regulating plant growth and development.Studies have confirmed that PPERFUL plays an important role in the process of peach fruit nucleus split,but its regulation mechanism during peach fruit ripening and softening needs further study.This study first identified five members of the AP1/FUL subfamily in the peach genome;then analyzed the expression characteristics of the five members in different tissues,fruit development different stages,and fruit ripening and softening stages to identify that may be involved in peach fruit ripening and softening of the AP1/FUL subfamily member.Secondly,'Bu Mu Zao Sheng' was used as the test material to study the function of PrupeFUL4 gene in the process of peach fruit ripening and softening by transiently silencing and overexpressing.In addition,FUL as a bait protein,explore its interaction with other transcription factors to regulate the mechanism of peach fruit ripening and softening.Mainly achieved the following research results:(1)Through bioinformatics analysis,five AP1/FUL subfamily members were identified in the peach genome.Phylogenetic tree analysis showed that Prupe AP1 was located in the eu AP1 clade,while the other four members were located in the eu FUL clade,so they were named Prupe AP1 and Prupe FUL1,2,3,4,respectively.Like other FUL-like proteins,members of the AP1/FUL subfamily have a highly conserved FUL motif(L/MPPMWL)at the C-terminal except for the MADS-box conserved domain,while the Prupe AP1 protein exhibits a different C-terminus motif.(2)The expression of members of the AP1/FUL subfamily has obvious tissue specificity.Prupe AP1 is mainly expressed in flowers,Prupe FUL1,2,3 is mainly expressed in leaves or roots,and PrupeFUL4 is mainly expressed in fruits and flowers.The expression level of PrupeFUL4 was higher in the second expansion and ripening stages of the fruits,but the expression level in the ripening and softening stage of the fruits was not affected by the treatment of ethylene and ABA.(3)Virus-mediated PrupeFUL4 gene silencing,the relative expression of PrupeFUL4 gene in flesh is significantly down-regulated,fruit softening is significantly inhibited,and ethylene biosynthesis related genes(Prupe ACO1 and Prupe ACS2),ABA biosynthesis related genes(Prupe NCED5)and cell wall degradation related genes(Prupe PME3,Prupe Lox1,Prupe EXP2,Prupe PG,and Prupe?-Man)were also down-regulated to varying degrees.The transient overexpression of PrupeFUL4 in fruits significantly promoted fruit softening,and the expression levels of genes related to ripening and softening were also up-regulated to varying degrees.Car G-box elements are present in the promoter regions of genes related to ripening and softening,indicating that PrupeFUL4 transcription factor may participate in the process of fruit ripening and softening by regulating the transcription of genes related to ripening and softening.(4)Fourteen proteins interacting with PrupeFUL4 were selected by yeast sieve library test.Yeast two-hybrid results show that PrupeFUL4 interacts with Prupe SEP1,Prupe SEP2,Prupe SEP3,Prupe OFP5,Prupe GL3,and Prupe AGL11 proteins,and these proteins are closely related to flower development and fruit ripening.Therefore,PrupeFUL4 may also regulates peach fruit ripening and softening by interacting with other transcription factors,but its molecular regulatory mechanism still needs further study.