PpyERF060-PpyABF3-PpyMADS71调控乙烯信号通路介导的梨芽休眠进程

PpyERF060-PpyABF3-PpyMADS71 Regulates Ethylene Signaling Pathway-Mediated Pear Bud Dormancy Process

为明确DAM(Dormancy-Associated MADS-box)基因参与休眠过程调控的分子机制,以‘砀山酥梨’花芽为研究材料,对前期鉴定出的梨DAM基因PpyMADS71的上游调控因子开展挖掘与初步分析。结果表明:(1)芽休眠过程中乙烯响应基因PpyERF060(ethylene responsive factor 060)与PpyMADS71共表达,酵母点突变实验及双荧光素酶实验确定PpyERF060结合在PpyMADS71启动子的DRE1元件处并激活其表达;过表达PpyERF060的‘茄梨’愈伤组织中PpyMADS71表达上调,进一步证明转录激活作用。(2)对PpyERF060的上游基因展开探索,发现可对其转录产生激活作用的ABA响应因子PpyABF3(ABRE BINDING FACTOR 3),并初步确定PpyABF3在PpyERF060启动子上的结合位点。(3)外源乙烯可诱导‘砀山酥梨’芽中PpyERF060表达,同时抑制PpyABF3转录;且在‘茄梨’愈伤组织中过表达PpyERF060对PpyABF3的转录起到抑制作用。通过上述结果,初步发现由PpyERF060、PpyABF3和PpyMADS71构成的互作网络可整合乙烯与脱落酸的信号通路进而调控梨芽休眠进程。

In order to clarify the specific regulation mode of DAM gene in dormancy,’Dangshan Suli’pear floral buds were used as the material to explore the upstream transcriptional factors of a DAM gene PpyMADS71 identified in previous studies.The results are as follows:(1)PpyERF060(Ethylene responsive factor 060)and PpyMADS71 were co-expressed during bud dormancy cycle.The yeast binding site mutation assays and the dual-luciferase assays were used to confirm that PpyERF060 could bind to the DRE1 element of the PpyMADS71 promoter and activates PpyMADS71’s expression,which was further proved by the upregulation of PpyMADS71 expression in PpyERF060 transgenic pear calli.(2)To explore the upstream genes of PpyERF060,we found that the transcription factor PpyABF3(ABRE BINDING FACTOR 3)could activate the expression of PpyERF060 and the binding site of PpyABF3 on the promoter of PpyERF060 was primarily determined.(3)Furthermore,ethylene treatment on’Dangshan Suli’buds caused up-regulation of PpyERF060 and down-regulation of PpyABF3.In addition,overexpression of PpyERF060 in pear calli caused inhibition of the transcription of PpyABF3.Based on the above results,we found that there is a regulatory network among PpyERF060,PpyABF3,and PpyMADS71,which could combine ethylene and ABA signaling pathway,and ultimately regulates the dormancy process.