小麦叶锈病菌效应蛋白Pt2567对小麦TcLr28表现无毒效应

doi:10.7668/hbnxb.20195743

摘要/Abstract

摘要：

旨在揭示小麦叶锈菌分泌的效应蛋白Pt2567在小麦叶锈菌致病中发挥的作用,为明确效应蛋白与寄主的互作机制奠定基础。采用烟草瞬时表达技术、细菌Ⅲ型分泌系统、qRT-PCR和寄主诱导的基因沉默(HIGS)技术对效应蛋白Pt2567开展亚细胞定位、抑制BAX(小鼠Bcl-2家族促死亡蛋白)试验、对胼胝质沉积和活性氧积累的影响研究及Pt2567功能的初步分析。结果表明,效应蛋白Pt2567在烟草中定位于细胞核,可以在烟草上抑制BAX诱导的细胞程序性死亡(PCD)。与对照组相比,在TcLr28中过表达效应蛋白Pt2567显著增强了胼胝质的沉积;在TcLr28上,效应蛋白Pt2567过表达24 h后提高了活性氧的积累。与对照相比,在接种生理小种THTT的TcLr28上沉默效应蛋白Pt2567降低了TcLr28抗病性,小麦叶锈菌夏孢子堆数量增多,激光共聚焦组织观察发现沉默效应蛋白Pt2567后病原菌发育进程加快。这些结果均表明,效应蛋白Pt2567在TcLr28中起正调控作用,推测效应蛋白Pt2567是Lr28的候选无毒基因。

关键词: 小麦叶锈菌, 效应蛋白, Pt2567, 无毒基因, 致病机制

Abstract:

The aim was to reveal the role of effector protein Pt2567 secreted by Puccinia triticina(Pt) in the pathogenesis and to lay a foundation for clarifying the interaction mechanism between effector protein and wheat.Tobacco transient expression technology,bacterial type Ⅲ secretion system,host induced gene silencing (HIGS) and other technologies were employed to carry out the subcellular localization,inhibition of BAX (mouse Bcl-2 family death promoting protein),effects on callose deposition and reactive oxygen species accumulation and preliminary function analysis of Pt2567.The results showed that the effector protein Pt2567 was subcellular located in the nucleus,and could inhibit BAX-induced programed cell death (PCD).Compared with control group,overexpression of effector protein Pt2567 in TcLr28 significantly enhanced the callose deposition and could increase the active oxygen accumulation at 24 h after the overexpression of the effector protein Pt2567 significantly.The silencing of effector protein Pt2567 on the TcLr28 inoculated with physiological race THTT significantly reduced the disease resistance of TcLr28,and increased the number of uredinium of Pt,and histological observation by confocal laser showed that silencing effector protein Pt2567 accelerated the development of wheat leaf rust.These results all suggest that effector protein Pt2567 plays a positive regulatory role in TcLr28,and is presumed to be a candidate avirulent gene for Lr28.

Key words: Puccinia triticina, Effector protein, Pt2567, Avirulent gene, Pathogenicity

中图分类号:

张赢丹, 张悦, 常佳迎, 张娜, 杨文香. 小麦叶锈病菌效应蛋白Pt2567对小麦TcLr28表现无毒效应[J]. 华北农学报, 2026, 41(1): 177-183. doi: 10.7668/hbnxb.20195743.

ZHANG Yingdan, ZHANG Yue, CHANG Jiaying, ZHANG Na, YANG Wenxiang. Effector Protein Pt2567 Secried by Puccinia triticina Shows Avirulence in Wheat TcLr28[J]. Acta Agriculturae Boreali-Sinica, 2026, 41(1): 177-183. doi: 10.7668/hbnxb.20195743.

http://www.hbnxb.net/CN/Y2026/V41/I1/177

图/表 5

图1 Pt2567在烟草中的亚细胞定位 N.细胞核;标尺为50 μm。

Fig.1 Subcellular localization of Pt2567 in N.benthamiana N.Nucleus;Bar=50 μm.

图2 Pt2567在烟草中抑制BAX诱导的PCD A.烟草注射示意图;B.红色圈内是Pt2567与BAX共注。

Fig.2 Pt2567 inhibits BAX-induced PCD in tobacco A.Schematic diagram of tobacco injection;B.Inside the red circle represents Pt2567 co-injected with BAX.

图3 Pt2567在TcLr28中过表达促进胼胝质沉积和活性氧积累 A.观察小麦叶片注射4种菌液苯胺蓝染色后胼胝质情况,标尺为100 μm;B.4组处理后每平方毫米胼胝质的数据统计,不同小写字母表示差异显著(P<0.05),图4同;C.接种叶锈菌24 h后的活性氧积累情况,标尺为50 μm;D.3组处理后每平方毫米活性氧的面积统计。

Fig.3 Overexpression of Pt2567 in TcLr28 promotes callose deposition and reactive oxygen species accumulation A.The callose of wheat leaves after injection of four bacterial solutions and aniline blue staining was observed(Bar=100 μm);B.Data statistics per square millimeter after four groups of treatments,different lowercase letters indicate significant differences(P<0.05),the same as Fig.4;C.Accumulation of reactive oxygen species after 24 h of inoculation with leaf rust fungus(Bar=50 μm);D.The area of reactive oxygen species per square millimeter after the three groups of treatments was counted.

图4 沉默 Pt2567 的表型观察及表达结果 A.接种病毒15 d表型观察;B.接种KHHT后10 d叶片表型观察;C.检测Pt2567的相对表达量。

Fig.4 Phenotypic observation and expression results of silencing Pt2567 A.Observation of viral inoculation phenotypes for 15 days;B.Observation of leaf phenotype at 10 days after inoculation with KHHT;C.Detecting the gene silencing efficiency of Pt2567.

图5 在TcLr28中沉默 Pt2567 的激光共聚焦显微观察结果 AP.附着胞;IH.侵染菌丝;SV.气孔下囊;HMC.吸器母细胞;H.吸器;标尺为30 μm。

Fig.5 Histological observation of silencing Pt2567 in TcLr28 under laser confocal microscopy AP.Attachment cell;IH.Infection hyphae;SV.Stomatal sac;HMC.Haustorial mother cells;H.Suction device;Bar=30 μm.

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