The Mechanism Of MabHLHs And Membrane Lipid Metabolism In Ethylene-induced Cold Tolerance In Banana Fruit

Banana is one of the most important tropical and subtropical fruits in the world,and it is a typical cold-sensitive fruit.Therefore,it is easy for banana fruit to suffer from cold stress during storage and transportation,which will seriously affect and reduce its commodity value bHLH transcription factors are involved in the regulation of banana biological and abiotic stress response mechanisms and play an important role in the post-harvest storage and transportation of banana fruits.The membrane lipid structure was destroyed and the content and components of membrane lipid were also changed in the process of chilling injury.In other study,it was found that ethephon treatment could induce the cold tolerance of banana fruits,but the mechanism was not completely clear,especially whether and how bHLH was involved in the regulation of membrane lipid anabolism genes under cold stress.In this study,we isolated two bHLH genes(MabHLH112/115)and DGK gene(Ma DGK2)and PLD genes(Ma PLDδ1/5),we also investigate the effect of the cold stress and ethephon treatment on the expession of these genes,we examined the relationship of MabHLH112/115 beteween the promoter of Ma DGK2 and Ma PLDδ1/5genes,which are helpful to clarify the role of bHLH genes and DGK gene and PLD genes in banana fruits treated with ethephon under low temperature stress.The main results are as follows.1.Two MabHLHs genes named MabHLH112 and MabHLH115 respectively were isolated from the banana fruit.Amino acid sequence and phylpgenetic tree analysis showed that the two genes both contained typical bHLH conserved domains,and belonged to the bHLH transcription factor;sucellular localization showed that MabHLH112/115 was located in the nucleus and was nuclear protein;transcription activity analysis illustrated that the MabHLH112/115 have transcriptional activation activities;Three membrane lipid anabolism genes named Ma DGK2 、 Ma PLDδ1 、Ma PLDδ5 respectively were characterized from the banana fruit.2.Ethephon treatment could alleviate the chilling injury of banana fruit.Real-time quantitative PCR analysis showed that the expressions of MabHLH112/115,Ma DGK2,Ma PLDδ1 and Ma PLDδ5 were significantly increased with the process of fruit chilling injury,while the expressions of these genes were gradually decreased under ethephon treatment.3.Electrophoretic mobility shift assay(EMSA)illustrated that MabHLH112/115 bind specifically to the E-box elements in the promoters of membrane lipid anabolism genes Ma DGK2,Ma PLDδ1 and Ma PLDδ5.Dual-luciferase reporter assay(DLR)indicated that MabHLH112/115 activate the promoter activities of these genes.4.Ma MYB4 could bind the promoters of MabHLH112/115 and inhibited the promoters activities of these genes.