Screening Of Auxin Response Factor PsARF Related To Postharvest Softening Of Green Plum And Its Mechanism Of Actio

The Qingcui plum(Prunus salicina Lindl.Cv.Qingcui)is a characteristic fruit of Chongqing,loved by consumers for its sweet and crisp taste and unique flavor.The Qingcui plum is a climacteric fruit,with vigorous postharvest physiological activity and a short shelf life.Without treatment,it will soften in 2-3 days,which greatly reduces its economic value.Clarifying the mechanism of Qingcui plum fruit softening during postharvest storage and identifying key regulatory factors is of great significance for extending the storage time.Auxin plays an important role in many physiological processes of plants,such as cell elongation,root formation,and fruit ripening and softening.Auxin response factors(ARFs)are a class of transcription factors that regulate the expression of auxin response genes and play an important role in auxin signal transduction.In this study,the ARF transcription factor family was screened from the plum genome,and PsARF1 related to fruit softening was identified from the postharvest transcriptome database of Qingcui plum.Then,its expression and function were analyzed by q RT-PCR,subcellular localization,bimolecular fluorescence complementation,yeast one-hybrid,and tomato genetic transformation.The main results are as follows:(1)Seventeen PsARF gene family members with 2-14 exons contain were identified from the plum genome,located on 8 chromosomes could be divided into 5groups.Ten motif and Auxin＿resp domain were predicted by MEME and Blast CD respectively.Transcriptome data showed that only PsARF1 was up-regulated during postharvest storage of Qingcui plum,indicating that PsARF1 maybe be involved in fruit softening regulation.(2)The ORF(Open reading frame)of PsARF1 is 2103 bp,encoding 701 amino acids.Sequence alignment analysis showed that PsARF1 contained classic conserved domains: B3 domain,Auxin＿Resp domain,and AUX＿IAA domain.Further q RT-PCR analysis showed that the expression of PsARF1 gradually increased during postharvest storage of Qingcui plum,reaching a peak on day 4,about 13.2times of day 0,and then maintained at a high level.Its expression was induced by ethylene and inhibited by 1-MCP treatment.In addition,exogenous NAA treatment after harvesting of immature Qingcui plum has a similar effect to ethylene treatment,both of which can promote softening and the expression of the PsARF1 gene in Qingcui plum.(3)Subcellular localization analysis showed that PsARF1 protein was located in the nucleus.Bimolecular fluorescence complementation results showed that PsARF1 could interact with the mature softening-related transcription factors PsERF76 and PsIAA2 proteins in plants,and also interact with itself.Yeast one-hybrid experiments showed that PsARF1 could bind to the promoter of PsERF76.By using Agrobacterium-mediated transformation of micro-tom,18 transgenic plants overexpressing PsARF1 were obtained.In conclusion,this study identified PsARF1 as a regulator of postharvest maturation and softening of Qingcui plum,which may play a role by interacting with PsERF76 and PsIAA2 and regulating the expression of PsERF76.This study laid the foundation for further analysis of the molecular mechanism of ARF transcription factors in regulating postharvest softening of Qingcui plum.