| Grape(Vitis vinifera L.)is one of the main economic fruit trees in China.During storage,grape is subject to berry decaying and dropping,rachis drying and browning,which seriously affect fruit quality,the commercial quality and shelf life.Studies have shown that ethylene plays an important role in fruit ripening and obscission.As we know,grape berry is a typical non-climacteric fruit,while ethylene production of rachis is more than 10 times of the berry,which has a similar ethylene release mode of climacteric fruit.ACS is the key gene in the process of ethylene synthesis,and the results have been shown that the VvACS1 gene is specifically expressed in the rachis of the grape.In the signal transduction pathway of ethylene,ethylene receptor gene(ETR)is the most upstream regulatory element,while the ERF transcription factor family has regulatory effect on the key genes of ethylene synthesis.In this study,we investigated the effect of ectopic expression of VvACS1 on tomato to analyze the function of VvACS1.The role of ethylene receptors in ethylene signaling pathway was investigated by analyzing the spatiotemporal expression of VvETR genes in different grape varieties.By studying the interaction between VvERF75 and VvACS1 promoter,this study clarified how ethylene response factor regulated the expression of VvACS1 gene and then regulated the synthesis of ethylene.The main results are as follows:1.After overexpression of VvACS1 in tomato,the content of auxin was reduced by 32%,and the content of zeatin was increased by 70%in the lateral buds,which weakened the apical dominance and promoted the formation of lateral buds.The zeatin:auxin ratio was increased from 1:5 to 1:2,which inhibited adventitious root production in cuttings compared to the control.2.The relative expression levels of zeatin O-glucosyltransferase(ZOG),auxin repressed/dormancy-associated protein(ARP),and ERF transcription factors were higher in transgenic plants than those in WT,which further suggests that ethylene may regulate auxin transport and distribution in shoots and that adventitious root formation is regulated by the synergistic mechanism between auxin and ethylene.3.Ethylene production in two tested grape varieties was very low in berry,and ethylene production was close to 0 at berry maturation,which showed a decreasing trend during berry developing.Ethylene release pattern of rachis is different as the berry.There was an ethylene climacteric release peak(up to 8.83μL·Kg-1·h-1)at post-harvest stage in the rachis of ’Hanxiangmi’,while the highest ethylene release peak(2.67μL·Kg-1·h-1)in the rachis of ’Shine-Muscat’ occurred at the 14 days after full blooming.The.expression pattern of VvETR4 in two cultivars was different,and the expression patterns of other three VvETRs were basically similar in the berry,of which the expression levels were higher in the early of berry developing stage.The expression of VvETRs in rachis was higher in the later stage of developing or post-harvest stage.The correlation analysis revealed that the expression of VvETR2 and VvETR4 were significantly positively correlated with ethylene production in berry,while the expression of VvETR2 was significantly nagetively correlated with ethylene production in rachis.The results suggest that VvETR2 and VvETR4 may play an important role in ethylene signaling pathway.4.Gene expression level of grape ERF transcription factors were analyzed,and the results showed that the expression trend of VvERF75 and VvACS1 increased with the developing of rachis,and there was a significant correlation between the expression levels of two genes,the correlation coefficient was 0.833.Yeast one-hybird(Y1H)assay showed that VvERF75 could bind the promoter of VvACS1 in vitro;Dual-Luciferase Reporter assay showed that VvERF75 could inhibite the transcription of VvACS1. |
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