The Mechanism Of ERF Transcription Factors In Regulating The Membrane Lipid Metabolism Of Cucumber Fruit During Cold Storage

Cucumber being a typical cold-sensitive fruit is prone to CI when the storage temperature is lower than 10°C.The main symptoms of cucumber CI include the disappearance of fresh fruit odor,pitting,serious water loss and wilting,which leads to the reduced commercial value of the fruit.Ethylene responsive factor(ERF)is a plant-specific transcription factor,which plays an important role in biotic and abiotic stresses.However,there is little information about the mechanism underlying ERFs in regulating CI of cucumber fruits.Previous studies have shown that after CI,the structure of membrane lipids will be destroyed and the membrane lipid content and composition will also change.Thus the aims of this work are to study the changes in the membrane components of cucumber and the ERF transcription factors on membrane lipid anabolism in ‘Jinpin Pioneer' cucumber fruit stored at 25°C at room temperature and 4°C at low temperatures.The molecular mechanism of Cs ERFs in regulating membrane lipid anabolism will provide a theoretical basis for improving chilling tolerance of cucumber fruit.The main results are as follows:1?CI symptoms of cucumber appeared during storage at 4°C for 5 d.The content of unsaturated fatty acids in membrane lipids increased firstly and then decreased after 5 days of storage.It is deduced that the unsaturated fatty acids in cucumber fruits are involved in the mechanism of cold tolerance.2?Sixteen membrane lipid metabolism-related genes and three ERF transcription factors were characterized.q RT-PCR analysis showed that Cs SAD1/2?Cs FAD2/3/5/7?Cs PLD?1/?2?Cs PLD?1?Cs PLD?1?Cs DGK1/5.1?Cs SPK1/2 which are related to membrane lipid metabolism were induced in cucumber fruit of cold storage,while Cs FAD6 and Cs DGK5.2 were inhibited.in addition,expression of Cs ERF17/24 was obviously up-regulated in the fruit of cold storage,indicating that low temperature is involved in the chilling injury by affecting membrane lipid metabolism and the expression of Cs ERF27 of cucumber fruits.3?The full-length c DNA sequence of Cs ERF17/24/27 gene was cloned.The amino acid sequence analysis showed that the Cs ERF17/24/27 proteins contained a typcial ERF domain.Subcellular localization indicated that Cs ERF17/24/27 was located exclusively in the nucleus.Transcriptional activity analysis illustrated that Cs ERF17/24 have transcriptional activation activities,whereas Cs ERF27 possesses transcriptional repression capacity.4?The EMSA and dual luciferase reporter assays demonstrated that Cs ERF17 recognizes and binds to the promoters of Cs SPK1/2,Cs DGK1,Cs PLD?1,Cs PLD?1,Cs FAD2 and Cs SAD1,and activates their activities.Taken together,our results suggest that Cs ERFs may regulate the membrane lipid metabolism of cucumber at least in part by activating a subset of membrane lipid meatabolism-related genes.