bHLH转录因子MdbHLH4调控苹果抗旱性的功能鉴定及分子机理研究

研究证明bHLH转录因子能够结合到抗逆相关基因的启动子上，通过调控靶基因的表达调节植物的抗逆性。在该过程中，DREB家族基因是其调控的一类主要靶基因。虽然DREB基因对植物的抗旱性具有显著的调控作用，但是bHLH转录因子能否通过调节DREB基因的表达调节苹果的抗旱性目前尚不清楚。项目组前期从苹果中克隆到一个抗旱调节关键基因MdbHLH4，并已获得其过表达的转基因苹果植株。在此基础上，拟通过转基因材料鉴定MdbHLH4的抗旱调控功能；通过ChIP-Seq筛选受MdbHLH4调控的DREB基因，并利用ChIP-PCR、EMSA进行验证；另外，拟通过GUS染色、LUC/REN双荧光瞬时表达鉴定MdbHLH4对DREB基因启动子转录活性的调控功能。该研究可初步阐明MdbHLH4通过调控DREB基因的转录调节苹果抗旱性的分子机理，为苹果的抗旱育种提供理论依据和基因资源。

Studies showed that bHLH transcription factors (TFs) can bind to the promoters of stress resistance-related genes to mediate various stress resistance, by regulating the expression of these target genes in plants. In these processes, the DREB gene family is one of the main targets for bHLH TFs. It is well konwn that DREB genes play crucial roles in regulating plant drought resistance, however, whether bHLH TFs could modulate apple drought resistance by regulating the expression of DREB genes is unclear. In our preliminary study, a key drought responsive gene MdbHLH4 has been isolated from apple, and the transgenic apple plantlets with MdbHLH4 overexpressed have also been obtained. Based on this, we will test the function of MdbHLH4 in regulating drought resistance using the transgenic plants. Furthermore, we will identify candidate DREB genes regulated by MdbHLH4 by searching the whole-genome of MdbHLH4-GFP transgenic apple plantlets with ChIP-Seq. Then, ChIP-PCR and EMSA analyses will be performed to verify the binding capacity of MdbHLH4 with the promoter regions of target DREB genes. Moreover, the transcriptional activity of MdDREBs mediated by MdbHLH4 will be verified by GUS staining and LUC/REN double fluorescence transient expression analyses. From this study, the molecular mechanism of MdbHLH4 in regulating drought resistance of apple will be characterized, by mediating the transcription of target DREB genes. These results will provide theoretical basis and genetic resources for apple drought resistance breeding.