拟南芥转录调节因子AtBT4抗灰霉病的分子机制

Gray mould caused by Botrytis cinerea is one of the important world diseases, and always results in significant economic losses. Identifying of genes related to resistance, studying the molecular mechanism and disease-resistant breeding are very important and meaningful contribution to the gray mould control. AtBT4 was isolated and identified as a novel resistance-related gene of Arabidopsis against B. cinerea by functional complementation in our preliminary studies. AtBT4 gene encodes a transcription regulator with DNA-binding activity. In order to further understand the molecular mechanism of AtBT4, relationship between AtBT4 and the key genes in SA, JA/ET signal pathway will be performed by Real-Time PCR, Northern bolt and double mutant technique. Interactive transcription factors of AtBT4 were screened from Arabidopsis cDNA library and identified through yeast two-hybrid system and Co-immunoprecipitation technique. Chromatin immunoprecipitation assay was utilized to screen unknown target genes regulated by AtBT4 or its transcription factors. These works would yield a more complete understanding of AtBT4 regulatory signaling pathways and further elucidated the complex transcriptional networks and provide a new resource for molecular breeding and effective control of B. cinerea.

灰霉病是一种世界性的重要病害，常给农业生产造成严重经济损失。分离抗灰霉病相关基因，研究其抗病分子机制，进而培育抗病新品种是防治灰霉病的有效途径。项目组已通过功能互补实验首次明确了AtBT4为拟南芥抗灰霉病的相关基因，并确定它是通过SA和JA/ET信号途径调控拟南芥对灰葡萄孢的抗性。该基因编码产物属于转录调节因子，具有DNA结合活性。本项目拟通过Real-Time PCR、Northern bolt等方法对AtBT4与SA、JA/ET信号途径关键基因的双突变体进行分析，确定AtBT4在SA、JA/ET信号途径中的作用；利用酵母双杂交、Co-IP技术，从拟南芥cDNA文库中获得与AtBT4互作的转录因子；利用ChIP技术，获得AtBT4及其互作转录因子调控的下游功能基因，最终明确AtBT4调控拟南芥抗灰霉病的分子机制，为完善植物抗病反应信号通路奠定基础，对指导灰霉病的防治提供理论依据。

灰霉病是一种世界性的重要病害，常给农业生产造成严重经济损失。分离抗灰霉病相关基因，研究其抗病分子机制，进而培育抗病新品种是防治灰霉病的有效途径。项目组前期通过功能互补实验首次明确了AtBT4为拟南芥抗灰霉病的相关基因，该基因编码产物属于转录调节因子，具有DNA结合活性。为了明确AtBT4调控拟南芥抗灰霉病的分子机制，本项目利用RT-PCR、Real-Time PCR等技术，检测SA、JA/ET对AtBT4基因表达的影响及AtBT4基因对抗病防御相关基因表达的影响，确定了AtBT4基因的表达受JA和灰葡萄孢的诱导，AtBT4基因突变影响抗病相关基因PR1、PR4、PDF1.2和BIK1的表达，从而确定AtBT4基因是通过SA、JA信号途径影响拟南芥对灰葡萄孢的抗性。利用酵母双杂交等技术，从拟南芥酵母双杂交cDNA文库中筛选获得了10个与AtBT4互作的蛋白；利用酵母双杂交技术和BiFC技术，确定了7个候选互作蛋白与AtBT4互作。利用生物信息学等技术，获得了转录调节因子AtBT4调控的8个候选靶基因；获得了AtBT4-flag、AtBT4-Ha、AtBT4-GFP和AtBT4-YFP等拟南芥转基因植株，为进一步靶基因的验证奠定了良好的基础。该研究结果为完善植物抗病反应信号通路奠定基础，对指导灰霉病的防治提供理论依据。