AP2/ERF在青花菜苗期响应黑腐病的功能研究  

Functional Study of AP2/ERF in Response to Black Rot at Broccoli Seedling Stage

作　　者：唐征[1] 陈思雀 徐谦[1] 钟伟杰 刘庆[1] 朱世杨[1] TANG Zheng;CHEN Sique;XU Qian;ZHONG Weijie;LIU Qing;ZHU Shiyang(Key Laboratory of Crop Breeding in South Zhejiang,Key Laboratory of Wenzhou Endemic Crop Biotechnology Innovation,Wenzhou Vocational College of Science&Technology/Wenzhou Academy of Agricultural Sciences,Wenzhou,Zhejiang 325006,China)

机构地区：[1]温州科技职业学院/温州市农业科学研究院,浙南作物育种重点实验室,温州特色作物生物技术创新重点实验室,浙江温州325006

出　　处：《园艺学报》2024年第11期2523-2539,共17页Acta Horticulturae Sinica

基　　金：浙江省农业(蔬菜)新品种选育重大科技专项(2021C02065-4-2);温州市科技项目(2019ZX007-2);浙江省自然科学基金项目(LY18C150006)。

摘　　要：通过对黑腐病病原菌侵染后的青花菜(Brassica oleracea L.var.italica)叶片进行转录组测序分析,探究青花菜对黑腐病的响应机制,挖掘并鉴定其中的抗病相关基因。Gene Ontology(GO)分析显示,与未接种相比,接种病原菌3、5和7 d后上调的差异表达基因均显著富集在胁迫应答和植物激素信号途径;3个比较组共有的差异表达基因中,上调表达的基因编码32个AP2/ERF转录因子;AP2/ERF编码的蛋白质序列长度在132~344 aa,且启动子中含有大量应答脱落酸、生长素、茉莉酸、赤霉素和水杨酸的顺式作用元件。水杨酸(SA)处理能够显著增强青花菜对黑腐病的抗性,强烈诱导AP2/ERF以及防御相关基因的表达。此外,BolC1t04894H编码的EAR型转录抑制子ERF4定位于细胞核中。瞬时表达青花菜ERF4导致烟草叶片对黑腐病病原菌处理敏感。EMSA结果显示,青花菜ERF4可以结合WRKY33启动子中的GCC-box序列,双荧光素酶报告系统试验表明ERF4抑制WRKY33的表达。综上,黑腐病病原菌侵染后,激活了青花菜激素信号转导途径,SA强烈诱导AP2/ERF的表达,进而调控一系列抗病相关基因的转录,启动防御反应。To explore the response mechanism of black rot and identify disease-resistance related genes through RNA-Seq analysis of broccoli(Brassica oleracea L.var.italica)leaves infected by black rot pathogen.Gene Ontology(GO)analysis showed that up-regulated differentially expressed genes(DEGs)were significantly enriched in stress response and phytohormone signaling pathways after 3,5,and 7 days infected by pathogens compared to uninoculated.At these three time points,the up-regulated genes in overlapped DEGs encoded up to 32 AP2/ERF transcription factors.Their encoded protein sequence ranged from 132 to 344 amino acids in length,and its promoter contained a large number of cis-elements in response to abscisic acid,auxin,jasmonic acid,gibberellin and salicylic acid(SA).SA treatment significantly enhanced the resistance of broccoli to black rot,and strongly induced the expression of AP2/ERF and defense-related genes.BolC1t04894H encodes an EAR type transcriptional suppressor ERF4,which is localized in the nucleus.Transient expression of broccoli ERF4 resulted in sensitivity of tobacco leaves to Xcc treatment.Electrophoretic Mobility Shift Assay(EMSA)result indicated that ERF4 could bind the GCC-box sequence in the promoter of WRKY33.Subsequently,double luciferase reporter system experiments showed that ERF4 inhibited WRKY33 expression.Taken together,the phytohormone signalling pathway are activated in broccoli after black rot pathogen infection,and SA strongly induces the AP2/ERF expression,thereby regulating a series of defense-related genes transcription to response black rot.

关 键 词：青花菜 AP2/ERF转录因子 黑腐病 水杨酸 抗病性 

分 类 号：S635.9[农业科学—蔬菜学]