棉花抗黄萎病相关基因GhDIR1的初步功能分析

苏晓峰; 陈析丰; 黄雨欣; 王省芬; 郭惠明; 马伯军; 程红梅; 谢涛

doi:10.11983/CBB24135

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DOI: https://doi.org/10.11983/CBB24135

研究论文

棉花抗黄萎病相关基因GhDIR1的初步功能分析

苏晓峰 ,
陈析丰 ,
黄雨欣 ,
王省芬 ,
郭惠明 ,
马伯军 ,
程红梅 ,
谢涛

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¹浙江师范大学生命科学学院, 金华 321004; ²中国农业科学院生物技术研究所农业微生物资源发掘与利用全国重点实验室,北京 100081; ³河北农业大学农学院, 保定 071000

收稿日期: 2024-09-04

修回日期: 2024-10-12

网络出版日期: 2024-11-15

基金资助

农业生物育种重大项目(No.2023ZD04039-03-3)和浙江省自然科学基金(No.LZ23C130004)

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Preliminary Analysis of Function of the Gene GhDIR1 Related With Verticillium Wilt in Cotton

SU Xiao-Feng ,
CHEN Si-Feng ,
HUANG Yu-Xin ,
YU Sheng-Fen ,
GUO Hui-Meng ,
MA Ba-Jun ,
CHENG Hong-Mei ,
XIE Shou

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¹College of Life Sciences, Zhejiang Normal University, Jinhua 321004, China; ²National Key Laboratory of Agricultural Microbiology, Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, China; ³College of Agriculture, Hebei Agricultural University, Baoding 071000, China

Received date: 2024-09-04

Revised date: 2024-10-12

Online published: 2024-11-15

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摘要

由大丽轮枝菌引起的黄萎病是棉花生产中最主要的威胁之一, 其可导致棉花大幅减产和纤维品质严重下降。课题组前期对接种大丽轮枝菌的拟南芥进行转录组分析, 其中DIR1类蛋白基因AT3G53980.2受病原菌强烈诱导表达。本研究发现, 棉花脂质转移蛋白基因GhDIR1 (Gh_A09G180700.1)与其表现出高度的同源性。生物信息学分析表明GhDIR1开放阅读框(open reading frame, ORF)为351 bp, 编码116个氨基酸。亚细胞定位结果证明, GhDIR1定位于细胞膜上。分析GhDIR1在大丽轮枝菌V991侵染后的表达模式, 发现其能快速响应大丽轮枝菌侵染。利用病毒诱导的基因沉默(virus-induced gene silencing, VIGS)技术下调该基因表达后, 棉花对黄萎病菌抗性显著降低。分析野生型和GhDIR1沉默植株转录组测序结果, 发现差异表达基因主要在“类黄酮生物合成”“倍半萜和三萜生物合成”和“α-亚麻酸代谢”三个途径富集, 同时荧光定量结果表明三个途径中的六个关键基因GhCHS、GhDFR、GhCAD、GhSEQ、GhLOX和GhAOC在GhDIR1沉默植株中均下调表达, 与转录组数据一致; 推测GhDIR1可能通过介导类黄酮和萜类化合物合成途径, 并调节茉莉酸(jasmonic acid, JA)等植物激素的次级代谢, 来激活相关信号通路以影响植株抗病性。综上, GhDIR1作为棉花抗黄萎病的正向调控因子, 通过参与多种激素和抗病信号网络调控植物免疫防御反应。

关键词： 棉花黄萎病; 陆地棉; GhDIR1; 基因表达模式分析; 病毒诱导的基因沉默(VIGS)

本文引用格式

苏晓峰 , 陈析丰 , 黄雨欣 , 王省芬 , 郭惠明 , 马伯军 , 程红梅 , 谢涛 . 棉花抗黄萎病相关基因GhDIR1的初步功能分析[J]. 植物学报, 0 : 1 -0 . DOI: 10.11983/CBB24135

Abstract

Verticillium wilt, caused by Verticillium dahliae, is one of the most major threats in cotton production, which can lead to a significant reduction of cotton yield and fiber quality. Our research team previously conducted a transcriptomic analysis on Arabidopsis thaliana inoculated with V. dahliae and discovered that the gene AT3G53980.2, which encodes the DIR1-like protein, was strongly induced by the pathogen. In this study, we discovered that the cotton lipid transfer protein gene GhDIR1 (Gh_A09G180700.1) displays high homology with AT3G53980.2. Bioinformatic analysis indicated that the open reading frame (ORF) of GhDIR1 is 351 bp, encoding 116 amino acids. The subcellular localization analysis using transient expression displayed that GhDIR1 was shown to be localized to the cell membrane. Quantitative real-time PCR (qRT-PCR) analysis determined that GhDIR1 expression was rapidly induced by V. dahliae. Virus-induced gene silencing revealed that downregulation of GhDIR1 significantly increased cotton susceptibility to V. dahliae. Further analysis through transcriptomic sequencing of wild-type and GhDIR1-silenced plants revealed that differentially expressed genes were primarily enriched in pathways related to "flavonoid biosynthesis","sesquiterpene and triterpene biosynthesis" and "α-linolenic acid metabolism". The expression levels of marker genes (GhCHS, GhDFR, GhCAD, GhSEQ, GhLOX, and GhAOC) involved in three pathways were downregulated. This data suggests that GhDIR1 contributes to the synthesis of protective compounds. We propose that GhDIR1 may mediate the synthesis of flavonoids and terpenoids, regulate the secondary metabolism of plant hormones such as jasmonic acid (JA), and activate related signaling pathways. These results suggested that GhDIR1 might act as a positive regulatory factor and modulate immune defense responses through its regulation in multiple hormone and resistance signaling networks against Verticillium wilt in cotton.

Key words： Verticillium wilt; Gossypium hirsutum; GhDIR1; gene expression pattern analysis; virus-induced gene silencing (VIGS)

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