中国农业科学2025，Vol.58Issue(13)：2578-2590,13.DOI:10.3864/j.issn.0578-1752.2025.13.007
葡萄抗灰霉病相关转录因子VviERF045下游靶基因的筛选
Screening of Target Genes Downstream of VviERF045,a Transcription Factor Associated with Gray Mold Resistance in Vitis vinifera
摘要
Abstract
[Background]Gray mold is an important fungal disease that seriously endangers the grapevine(Vitis vinifera)industry worldwide.As an important transcription factor family in plants,ethylene-responsive factor(AP2/ERF)family plays a key role in regulating plant growth and development and stress response.[Objective]The study aimed to elucidate the molecular mechanisms by which the grapevine ethylene-responsive factor VviERF045 mediates defense against Botrytis cinerea.Through prediction and functional analysis of its downstream target genes,this research provides new insights into the transcriptional regulatory network associated with grapevine resistance to B.cinerea,establishing a foundation for breeding disease-resistant cultivars.[Method]Grapevine leaves were inoculated with B.cinerea using both agar disc and spore suspension methods.The expression profiles of VviERF045 and its predicted targets were evaluated via quantitative real-time PCR(qRT-PCR).Promoter cis-acting elements were analyzed using PlantCARE,while phylogenetic relationships and sequence alignments of VviERF045 were assessed using MEGA 7 and DNAMAN.Functional validation was conducted through Agrobacterium-mediated transient overexpression of VviERF045 in'Cabernet Sauvignon'leaves.To further explore its regulatory landscape,DNA affinity purification sequencing(DAP-seq)was performed on'Pinot Noir'leaves at 12 hours post-inoculation(hpi)with B.cinerea.[Result]Transient overexpression of VviERF045 significantly enhanced resistance to gray mold in'Cabernet Sauvignon'leaves.DAP-seq analysis was performed on V.vinifera'Pinot Noir'leaves at 12 hpi.Comparative analysis of peaks between two experimental replicates identified 51 806 consensus peaks.Subsequent genomic annotation revealed these peaks were predominantly located within±2 kb regions flanking promoter-transcription start sites(TSS).The top 2 000 most statistically significant peaks were selected for functional characterization through Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway enrichment analyses.Through integrated analysis of potential target genes and peak calling,three high-confidence candidate target genes(TCP8,SAP5,and bHLH48)were identified,all showing significant binding peaks in their promoter regions.Subsequent qRT-PCR validation confirmed the transcriptional upregulation of these genes,suggesting their cooperative involvement with VviERF045 in the grapevine's response to B.cinerea infection.[Conclusion]VviERF045 functions as a positive regulator of grapevine defense against B.cinerea,likely by activating key stress-responsive genes such as TCP8,SAP5,and bHLH48.These results provide mechanistic insights into the pathogen-responsive transcriptional network in grapevine and identify potential molecular targets for breeding resistant V.vinifera cultivars.关键词
欧洲葡萄/VviERF045/灰葡萄孢/灰霉病/靶基因/瞬时转化/DAP亲和纯化测序Key words
Vitis vinifera/VviERF045/Botrytis cinerea/gray mold/target gene/transient transformation/DAP-seq引用本文复制引用
赵钰磊,辛佳璐,李承男,李珊,谢旭飞,尹晓..葡萄抗灰霉病相关转录因子VviERF045下游靶基因的筛选[J].中国农业科学,2025,58(13):2578-2590,13.基金项目
宁夏自然科学基金优青(2023AAC05016)、国家自然科学基金(32302496) （2023AAC05016）
