The Mechanism Of MdNAC104 Mediated Auxin-Regulated Ethylene Synthesis In Apple Fruit

Apple is a well-known climacteric fruit.Ethylene plays a key regulatory role in ripening of apple fruit.The synthesis of ethylene is affected by other hormones,and the auxin is one of them.Our previous studies that exogenous auxin treatment induce ethylene synthesis and mature during the development(after picking,there is no natural post-cooked ability)of apple fruit,however,its regulatory mechanism needs further research.This study takes apples as material,using naphthalene acetic acid processing harvest the fruits of development stage.Transcription sequencing to identify a gene MdNAC104 that is being inhibited by auxin.Yeast one-hybrid assay,electrophoretic mobility shift assay,chromatin immunoprecipitation analysis,GUS activity assay and other experiments that the MdNAC104 is transcribed by negative regulation of MdACS1 and MdERF3,inhibiting expression of MdACS1 and inhibiting the ethylene synthesis of apple fruit.The main findings are as follows:1.Exogenous auxin treatment significantly promotes ethylene production during the development stage of apple fruit.The development of apple fruit was treated with a 4 m M NAA,and 20 days was stored at room temperature.The results showed that NAA treatment significantly promote ethylene production in apple fruit,and the peak of breathing occurred in the treatment of 15 days;there was no significant change in ethylene production in untreated apple fruits.2.Exogenous auxin treatment significantly induces expression of key genes in ethylene synthesis and signal transduction.Expression analysis of the key genes in ethylene synthesis and signal transduction that the expression of MdACS1 and Md ACO1 in the fruit is extremely low;After NAA treatment,the expression of MdACS1 and Md ACO1 is significantly higher than the control;Compared with the control,the expression of MdERF3 gene increased significantly after NAA treatment,and similar to the ethylene production.3.Exogenous auxin treatment significantly inhibits the expression of MdNAC104 during the development stage of apple fruit.Transcription sequencing of the control and NAA treatment of room temperature storage 5 d.A total of 22 differentially expressed NAC transcription factors,and the MdNAC104 was significantly inhibited by the NAA.4.MdNAC104 is a negative regulator of ethylene synthesis.In order to clarify the function of the MdNAC104 in fruit ripening,using the Agrobacterium-mediated instantaneous expression system to silence it in the apple fruit.The results show that the ethylene production is significantly increased in the silencing MdNAC104 fruit,and the NAA treatment further promotes the generation of fruit ethylene.The expression of the ethylene biosynthesis pathway and the signal transduction pathway genes are increased in the silencing MdNAC104 fruit,and NAA treatment further induces its expression.5.The MdNAC104 binding to MdACS1 and MdERF3 promoter,and negative regulation of its expression.Yeast one-hybrid assay and electrophoretic mobility shift assay showed that MdNAC104 binding to MdACS1 and MdERF3 promoter in vitro;chromatin immunoprecipitation analysis showed that MdNAC104 binding to MdACS1 and MdERF3 promoter in vivo.GUS activity assay showed that MdNAC104 negative regulation of MdACS1 and MdERF3 promoter activity;NAA treatment inhibits the negative regulation of MdNAC104 on MdACS1 and MdERF3.In summary,the exogenous auxin inhibits the transcription of MdNAC104 in developing apple fruit,inhibits the negative regulation of MdACS1 and MdERF3 promoters by MdNAC104,induces MdACS1 expression,and promotes fruit maturity.This study initially clarifies MdNAC104-mediated molecular mechanisms that mediate the growth of apple fruit ethylene,which provides theoretical basis for the maturity and storage of manual regulation in the molecular level.