Article(id=1319998834189644398, tenantId=1146029695717560320, journalId=1316781310003478668, issueId=1319998767835754596, articleNumber=null, orderNo=null, doi=10.11983/CBB25131, pmid=null, cstr=32102.14.CBB25131, oa=null, hot=null, price=null, onlineType=0, articleFormat=0, articleType=null, articleTypeStr=research-article, receivedDate=1753200000000, receivedDateStr=2025-07-23, revisedDate=null, revisedDateStr=null, acceptedDate=1756742400000, acceptedDateStr=2025-09-02, onlineDate=1792641846462, onlineDateStr=2026-10-22, pubDate=1757433600000, pubDateStr=2025-09-10, doiRegisterDate=null, doiRegisterDateStr=null, onlineIssueDate=1792641846462, onlineIssueDateStr=2026-10-22, onlineJustAcceptDate=null, onlineJustAcceptDateStr=null, onlineFirstDate=null, onlineFirstDateStr=null, sourceXml=null, magXml=null, createTime=1792641846462, creator=13701087609, updateTime=1792641846462, updator=13701087609, issue=Issue{id=1319998767835754596, tenantId=1146029695717560320, journalId=1316781310003478668, year='2025', volume='60', issue='5', pageStart='669', pageEnd='853', issueExtLink='null', onlineDate='null', pubDate='1757433600000', pubDateStr='2025-09-10', beforeIssueId=null, nextIssueId=null, price=null, status=1, issueComplete=1, articleOrder=1, issueType=-1, specialIssue=null, createTime=1792641830643, creator='13701087609', updateTime=1792641943258, updator='13701087609', preIssue=null, nextIssue=null, articleTotal=null, ext={EN=IssueExt(id=1319999240240222831, tenantId=1146029695717560320, journalId=1316781310003478668, issueId=1319998767835754596, language=EN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=), CN=IssueExt(id=1319999240240222832, tenantId=1146029695717560320, journalId=1316781310003478668, issueId=1319998767835754596, language=CN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=)}, issueFiles=null, downloadFileDto=null}, startPage=693, endPage=703, ext={EN=ArticleExt(id=1319998834571326063, articleId=1319998834189644398, tenantId=1146029695717560320, journalId=1316781310003478668, language=EN, title=Plant Immunity Study: Mechanism Breakthroughs and Application Innovations, columnId=1317485696684773674, journalTitle=Chinese Bulletin of Botany, columnName=SPOTLIGHTS, runingTitle=null, highlight=null, articleAbstract=

The plant innate immune system serves as the primary defense against pathogen invasion, with well-established frameworks for receptor recognition and signal transduction mechanisms. This review highlights recent key breakthroughs in plant immunity research from Chinese institutions: (1) The discovery of novel mechanisms driving virulence evolution through asymmetric chromosome distribution in fungi and chromosome fusion in oomycetes; (2) Elucidation of the kinase MtLICK1/2-mediated molecular switch that precisely regulates the symbiosis-immunity trade-off via phosphorylation of MtLYK3 in legumes; (3) Identification of a “sensor-executor” paradigm where tandem kinases and NLR immune receptors cooperatively activate immunity in cereal crops; (4) Innovative strategies including co-transfer of sensor-helper NLR pairs to overcome taxonomic restrictions; and (5) Develop technology of autoactive NLR chimeras activated by pathogen protease cleavage for broad-spectrum resistance. These advances collectively deepen our understanding of plant-pathogen-environment interactions across three dimensions—pathogen adaptive evolution, sophisticated host immune regulation, and receptor engineering applications. Crucially, fundamental mechanistic insights have been successfully translated into crop genetic improvement practices. The integrated findings provide a robust theoretical foundation and actionable technological framework for designing novel crop varieties with durable, broad-spectrum disea- se resistance to address mounting agricultural biosecurity threats.

, authors=Xiao Yinyan, Yu Hua, Wan Li*, authorsList=Xiao Yinyan, Yu Hua, Wan Li, authorCompany=null, correspAuthors=Wan Li, authorNote=
†These authors contributed equally to this paper
, correspAuthorsNote=
*E-mail:
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作为抵御病原入侵的核心防线, 植物天然免疫系统的受体识别与信号转导机制已建立较为完善的框架。该文对国内近期植物免疫领域取得的5项关键突破进行了阐述。(1) 发现了真菌染色体非均等分配及卵菌染色体融合驱动毒力进化的新规律; (2) 阐明了豆科植物中激酶MtLICK1/2通过磷酸化修饰MtLYK3精确调控“共生-免疫”转换的分子开关机制; (3) 明确了禾本科作物中串联激酶与NLR免疫受体以“sensor-executor”配对模式协同激活免疫的新范式; (4) 创新性提出“感受型与辅助型NLR共转移”策略以克服物种限制; (5) 开发出基于病原蛋白酶切割的自激活NLR嵌合体工程化改造技术, 赋予了植物广谱抗性。上述研究成果从病原适应性进化、宿主免疫精细调控及受体工程应用3个维度, 深化了对植物-病原-环境互作复杂网络的理解, 成功将基础机制认知转化为作物抗病遗传改良实践, 为设计培育具有持久、广谱抗性的作物新品种提供了坚实的理论基础与可操作的技术路径。

, authors=肖银燕†, 于华†, 万里*, authorsList=肖银燕, 于华, 万里, authorCompany=null, correspAuthors=万里, authorNote=
†共同第一作者
, correspAuthorsNote=
*E-mail:

万里, 中国科学院分子植物科学卓越创新中心研究员, 博士生导师, 入选国家青年千人计划, 获得上海市植物病理学会杰出青年科学家奖。万里课题组专注于植物免疫机制研究和广谱抗性农作物创制, 在植物细胞内免疫受体NLR工作的分子机理方面取得重要进展。相关成果以第一作者(含共同)发表在Science (2014, 2019), 以通讯作者(含共同)发表在Nature、Science、Proc Natl Acad Sci USA、Nature Plants、Science Advances和J Integr Plant Biol等学术期刊。

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Co-expression of sensor and helper NLRs enables cross-species regulation of disease resistance in angiosperms (upper panel); An artificially designed autoactive NLR, when cleaved by pathogen proteases, releases its activity to confer broad- spectrum disease resistance in plants (lower panel).

, figureFileSmall=/FgUlNZmCSHj+EFTOXo8aA==, figureFileBig=Yr99PdAPRgYTshvxstKRng==, tableContent=null), ArticleFig(id=1319998837255680654, tenantId=1146029695717560320, journalId=1316781310003478668, articleId=1319998834189644398, language=CN, label=图1, caption=NLR免疫受体改造拓宽植物的抗病性

共转移感受型和辅助型NLR, 实现NLR在被子植物中跨物种调控抗病性(上半部分); 人工设计的自激活NLR被病原菌蛋白酶切割后释放活性, 赋予植物广谱抗病能力(下半部分)。

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肖银燕†, 于华†, 万里*
作者信息
  • 中国科学院分子植物科学卓越创新中心, 上海 200032
通讯作者:

万里, 中国科学院分子植物科学卓越创新中心研究员, 博士生导师, 入选国家青年千人计划, 获得上海市植物病理学会杰出青年科学家奖。万里课题组专注于植物免疫机制研究和广谱抗性农作物创制, 在植物细胞内免疫受体NLR工作的分子机理方面取得重要进展。相关成果以第一作者(含共同)发表在Science (2014, 2019), 以通讯作者(含共同)发表在Nature、Science、Proc Natl Acad Sci USA、Nature Plants、Science Advances和J Integr Plant Biol等学术期刊。

作者简介:
†共同第一作者
Plant Immunity Study: Mechanism Breakthroughs and Application Innovations
Xiao Yinyan, Yu Hua, Wan Li*
Affiliations
  • CAS Center for Excellence in Molecular Plant Sciences, Shanghai 200032, China
  • Corresponding Author:
出版时间: 2025-09-10 doi: 10.11983/CBB25131
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作为抵御病原入侵的核心防线, 植物天然免疫系统的受体识别与信号转导机制已建立较为完善的框架。该文对国内近期植物免疫领域取得的5项关键突破进行了阐述。(1) 发现了真菌染色体非均等分配及卵菌染色体融合驱动毒力进化的新规律; (2) 阐明了豆科植物中激酶MtLICK1/2通过磷酸化修饰MtLYK3精确调控“共生-免疫”转换的分子开关机制; (3) 明确了禾本科作物中串联激酶与NLR免疫受体以“sensor-executor”配对模式协同激活免疫的新范式; (4) 创新性提出“感受型与辅助型NLR共转移”策略以克服物种限制; (5) 开发出基于病原蛋白酶切割的自激活NLR嵌合体工程化改造技术, 赋予了植物广谱抗性。上述研究成果从病原适应性进化、宿主免疫精细调控及受体工程应用3个维度, 深化了对植物-病原-环境互作复杂网络的理解, 成功将基础机制认知转化为作物抗病遗传改良实践, 为设计培育具有持久、广谱抗性的作物新品种提供了坚实的理论基础与可操作的技术路径。

植物免疫  /  植物-微生物互作  /  抗病蛋白  /  抗病遗传改良

The plant innate immune system serves as the primary defense against pathogen invasion, with well-established frameworks for receptor recognition and signal transduction mechanisms. This review highlights recent key breakthroughs in plant immunity research from Chinese institutions: (1) The discovery of novel mechanisms driving virulence evolution through asymmetric chromosome distribution in fungi and chromosome fusion in oomycetes; (2) Elucidation of the kinase MtLICK1/2-mediated molecular switch that precisely regulates the symbiosis-immunity trade-off via phosphorylation of MtLYK3 in legumes; (3) Identification of a “sensor-executor” paradigm where tandem kinases and NLR immune receptors cooperatively activate immunity in cereal crops; (4) Innovative strategies including co-transfer of sensor-helper NLR pairs to overcome taxonomic restrictions; and (5) Develop technology of autoactive NLR chimeras activated by pathogen protease cleavage for broad-spectrum resistance. These advances collectively deepen our understanding of plant-pathogen-environment interactions across three dimensions—pathogen adaptive evolution, sophisticated host immune regulation, and receptor engineering applications. Crucially, fundamental mechanistic insights have been successfully translated into crop genetic improvement practices. The integrated findings provide a robust theoretical foundation and actionable technological framework for designing novel crop varieties with durable, broad-spectrum disea- se resistance to address mounting agricultural biosecurity threats.

plant immunity  /  plant-microbe interactions  /  disease resistance proteins  /  genetic improvement for disease resistance
肖银燕, 于华, 万里. 植物免疫研究: 机制突破和应用创新. 植物学报, 2025 , 60 (5) : 693 -703 . DOI: 10.11983/CBB25131
Xiao Yinyan, Yu Hua, Wan Li. Plant Immunity Study: Mechanism Breakthroughs and Application Innovations[J]. Chinese Bulletin of Botany, 2025 , 60 (5) : 693 -703 . DOI: 10.11983/CBB25131
在植物与病原微生物长期进化过程中形成了先天免疫系统, 该系统由2部分组成, 一是由细胞表面受体(pattern recognition receptors, PRRs)识别细胞外病原菌相关分子模式诱发的免疫反应(PAMP-trigge- red immunity, PTI), 病原菌能够分泌效应因子(effec-tor)到植物细胞内抑制植物的PTI; 相应地, 植物进化出第二道免疫防线, 由细胞内免疫受体(nucleotide-binding leucine-rich repeat receptors, NLR)识别效应因子后诱导的免疫反应(effector-triggered imm-unity, ETI), 二者协同调控植物的抗病性(Jones et al., 2016; Zhou and Zhang, 2020; Ngou et al., 2021; Yuan et al., 2021)。
近几十年, 对植物NLRs介导抗病的机制研究取得了众多突破。NLRs蛋白不仅能以直接或间接方式识别病原菌分泌到细胞内的效应因子, 激活植物的免疫反应; 在发挥免疫调控作用时, 还能单独、成对或以复杂网络的形式调控ETI的发生(Liu and Wan, 2022)。根据NLR在免疫反应中的功能不同, 将其分为2类, 一类是能够“感知”病原菌效应因子的感受型NLRs (sensor NLRs), 负责识别效应因子并将此信号传递至下游; 另一类是负责感知上游sensor NLRs传递的“危险”信号, 起“辅助”作用, 执行免疫功能的辅助型NLRs (helper NLRs)。在拟南芥(Arabidopsis thaliana)中, ADR1s (activated disease resistance 1)和NRG1s (N requirement gene 1)作为辅助型NLRs调控植物的抗性和细胞死亡。茄科植物中, NRCs (NLR required for cell death)作为辅助型NLRs在感受型NLRs下游发挥调控免疫反应的作用(Liu and Wan, 2022; Gong et al., 2023)。
然而, 对于病原微生物增殖、植物区分有益微生物进而抑制免疫反应, 及禾本科作物中NLRs感知病原菌效应因子的机制还有待进一步研究。此外, 如何将目前已有植物免疫认知, 创新应用到植物病害防治中还需不断探索。本文聚焦上述问题, 对近期我国科学家在这些领域取得的突破性进展进行解读, 概述植物免疫研究中的机制研究突破和应用创新。
植物病原体种类繁多, 包括真菌、卵菌、细菌、病毒、线虫、吸虫和寄生植物等。其中, 真菌是最主要的植物病原菌, 占植物病害的70%-80% (Dean et al., 2012)。真菌具有极强的环境适应能力, 这主要得益于其高效的繁殖方式(有性和无性繁殖)及灵活的基因组变异机制(从点突变到全基因组复制)。此外, 转座元件、结构变异和倍性变化等基因组可塑性特征, 也有助于真菌快速适应环境压力和宿主防御(Zaccaron and Stergiopoulos, 2025)。深入研究这些进化机制, 可为开发新型抗真菌策略奠定分子基础。
四川大学张跃林团队最新研究揭示了一种颠覆性的真菌染色体分配机制, 为理解真菌发育及其适应性进化和耐药性产生提供了全新视角。以核盘菌(Sclerotinia sclerotiorum)为例, 其单倍体基因组包含16条染色体(1N=16), 每个子囊孢子通常含有2个细胞核。按照经典遗传学理论, 有丝分裂或减数分裂后, 每个子细胞核都应获得完整的单倍体染色体组(1N), 以保证细胞正常执行功能(Ekins, 1999)。然而, 通过综合运用染色体计数、荧光原位杂交、流式细胞术及单核PCR等多种技术手段, 发现16条染色体并未完整存在于每个细胞核中, 而是以非对称方式分配至2个核内(平均每个核约含8条染色体)。这种现象并非个例。在灰霉菌(Botrytis cinerea)分生孢子中也观察到类似的染色体分配模式: 其18条单倍体染色体(1N=18)被不均等地分配至4-6个细胞核, 每个核仅包含3-8条不等的染色体。这些发现表明, 染色体非均等分配机制可能是多核真菌中的普遍现象。这一机制也完美解释了为何在真菌遗传操作中, 对单倍体孢子进行基因编辑或诱变后可直接获得纯合突变体, 无需像二倍体生物那样进行繁琐得多代纯化步骤(Xu et al., 2025)。
染色体数目变异在卵菌致病性进化中同样发挥重要作用。卵菌(如疫霉属、腐霉属和霜霉属)虽形态类似真菌, 但在进化上属于不同的生物类群(Kamoun et al., 2015)。南京农业大学王源超团队通过端粒到端粒的全基因组测序分析, 发现染色体融合和分裂是导致卵菌染色体数目变异的主要原因。致病相关基因(如RxLR效应子)常富集在融合染色体的近端粒区域, 且表达水平较高, 表明这些区域在致病性进化中具有关键作用; 新发现的含Ankyrin重复序列的效应因子蛋白(ANK)在大豆疫霉(Phytophthora sojae)的融合染色体上特异性扩增并在感染过程中高表达, 可能直接参与病原菌的致病过程(Zhang et al., 2024)。
核盘菌染色体的非均等分配现象挑战了传统认知, 而染色体融合、断裂或非整倍性变异可能是毒力基因(如效应蛋白)复制或重排的重要驱动力。这些变异使病原菌通过染色体丢失或获得来调整基因剂量, 从而适应不同宿主。研究病原菌的这些独特进化规律, 不仅为理解植物免疫机制提供了新视角, 也为开发可持续的病害防控策略提供了理论依据。
与豆科植物形成共生关系的根瘤菌能将大气中的氮气(N2)转变为植物可吸收利用的铵(NH4+), 植物则为根瘤菌提供自身光合作用的产物作为碳源(Martin et al., 2017; Zipfel and Oldroyd, 2017; Poole et al., 2018; van Velzen et al., 2019)。这一共生关系的建立需要植物在根瘤菌侵染时抑制其本身的免疫反应, 从而启动共生信号通路。宿主免疫反应的抑制是植物根瘤共生形成的重要前提。
豆科植物通过含有LysM结构域的类受体激酶MtNFP和MtLYK3识别根瘤菌分泌的结瘤因子(Nod factor)。其中, MtLYK3具有激酶活性且对结瘤因子响应至关重要, 但在识别后未引起其与MtLYK3之间发生相互磷酸化, 表明存在其它组分参与共生信号过程(Limpens et al., 2003; Madsen et al., 2003; Arrighi et al., 2006; Radutoiu et al., 2007; Haney et al., 2011; Oldroyd, 2013; Moling et al., 2014)。此外, 当结瘤因子被识别, 共生信号被放大时, 植物免疫反应如何被抑制以协调“共生-免疫”两个近乎矛盾的过程也是重要的生物学问题(Liang et al., 2013)。
围绕结瘤因子被识别及共生信号被放大这一问题, 中国科学院分子植物科学卓越创新中心王二涛团队通过酵母双杂交筛选, 鉴定出与MtLYK3互作的胞质类受体激酶MtLICK1 (LYK3-interacting cytoplasmic kinase 1 ) (Wang et al., 2025a)。实验表明, MtLICK1/2能与MtLYK3在质膜上发生相互作用, 且Mtlick1/2与Mtlyk3突变体在根瘤菌侵染后的表型一致, 共生相关基因MtNIN (NODULE INCEPTION)和MtENOD11几乎不被结瘤因子诱导, 根瘤形成受到抑制。
MtLICK1/2具有激酶活性, 在结瘤因子响应过程中发挥重要作用。当MtLYK3磷酸化MtLICK1/2的位点突变为模拟非磷酸化状态(丙氨酸, A)时, MtLICK1/2的激酶活性丧失, 根瘤形成受阻; 突变为模拟磷酸化状态(天冬氨酸, D)时, 根瘤正常形成。进一步分析显示, 结瘤因子诱导MtLYK3依赖的MtLICK1/2磷酸化, 磷酸化的MtLICK1/2与MtLYK3互作减弱。此外, MtLICK1/2能反向磷酸化MtLYK3的第483和488位酪氨酸(Y), 这些位点突变为苯丙氨酸(F)时, MtLYK3的激酶活性丧失, 根瘤形成受到抑制, 表明在根瘤共生信号途径中MtLICK1/2介导的MtLYK3磷酸化在根瘤形成中具有关键作用。激活后的MtLYK3可磷酸化下游MtDMI2, 介导共生信号向下游传递。
MtLICK1/2不仅参与共生信号的传递, 还在根瘤菌侵染时的免疫反应抑制中发挥重要作用。基于前期研究报道的LjNFR1的YAQ motif在免疫-共生转换中发挥作用, 推测MtLICK1/2通过磷酸化MtLYK3的YAQ motif参与免疫-共生信号转换。Mtlick1/2在根瘤菌处理后表现出更高的ROS积累量和MAPK激活水平, 当对植物进行青枯菌(Ralstonia solanacearum)抗性测试时, Mtlick1/2突变体表现出比野生型更高的抗性。
用PAMPs (如肽聚糖(peptidoglycans, PNGs)或根瘤菌粗提物)处理植物后, 植物的结瘤能力显著下降, 表明植物的免疫反应抑制根瘤共生。用外源结瘤因子处理植物可抑制其免疫响应, 且依赖于MtLICK1/2。这些结果表明, MtLICK1/2被结瘤因子激活后发挥植物免疫调节的作用, 抑制植物的免疫反应, 促进植物根瘤共生形成。
该研究揭示了在根瘤菌侵染时, “共生-免疫”两条信号通路如何被精细调控以帮助植物区分环境中对自身有益和有害的微生物。
麦类作物中, 具有串联激酶的一类胞内蛋白(tandem kinase proteins, TKPs)在植物免疫反应中发挥重要作用(Chen et al., 2020), 如小麦(Triticum aestivum)抗茎锈病蛋白(Sr60/WTK2和Sr62/WTK5) (Chen et al., 2020; Yu et al., 2022)、抗条锈病蛋白(Yr15/ WTK1) (Klymiuk et al., 2018)、抗叶锈病蛋白(Lr9/ WTK6-vWA) (Wang et al., 2023)、抗白粉病蛋白(Pm24/WTK3、WTK4、Pm36/WTK7-TM和Pm57/ WTK6b-vWA)和抗麦瘟病蛋白(Rwt4) (Lu et al., 2020; Gaurav et al., 2022; Arora et al., 2023; Li et al., 2024; Zhao et al., 2024)。但这些串联激酶蛋白在植物免疫反应中的具体功能有待进一步研究。
前期研究表明, 小麦白粉病抗性品种葫芦头(Hulutou, HLT)中的WTK3 (wheat tandem kinase 3, Pm24)对白粉病抗性具有重要作用(Lu et al., 2020)。为阐明WTK3调控白粉病的抗性机制, 中国科学院遗传与发育生物学研究所刘志勇团队通过EMS诱变定位到1个与以往典型NLR不同的NLR蛋白WTN1 (wheat tandem NBD 1), 该蛋白具有N端的CC结构域、C端的LRR结构域以及位于二者之间的2个串联NBD结构域, WTK3与WTN1均位于小麦1D染色体, 距离114 kb (Lu et al., 2025)。遗传学分析表明, WTK3和WTN1的完整性为葫芦头小麦白粉病抗性所必需; 在野生型Fielder中包含了wtk3 (WTK3的等位基因, 对白粉病敏感)和位于1DS上有功能的WTN1, 表现为白粉病敏感; Fielder中过表达WTK3表现出对白粉病抗性, 在此基础上敲除WTN1后可恢复其敏感表型。进一步研究表明, WTK3和WTN1协同互作在小麦抗病过程中发挥作用。
Lu等(2025)利用WTK3的等位基因Rwt4 (Arora et al., 2023), 及Rwt4能识别的麦瘟菌(Magnapor- the oryzae)效应因子PWT4 (Inoue et al., 2017), 对WTN1介导的植物免疫反应激活机制进行研究, 发现小麦原生质体和烟草(Nicotiana benthamiana)中共表达PWT4-WTK3-WTN1或PWT4-Rwt4-WTN1能诱导细胞死亡, 表达其中1个或2个时不发生超敏反应。互作分析表明, 位于WTK3 N端的假激酶片段(pseudo-kinase fragment, PKF)负责识别效应因子PWT4, C端的激酶结构域II与WTN1互作, 通过WTK3/Rwt4对PWT4的识别介导WTN1激活。激活的WTN1形成1个高分子量多聚体(>880 kDa), 多聚体包含PWT4-WTK-3/Rwt4-WTN1三个组分。这种功能模式与此前已报道的ZAR1和Sr35类似——被效应因子激活的NLR形成抗病小体, 发挥Ca2+通道作用(Wang et al., 2019a, 2019b; Förderer et al., 2022)。电生理学实验表明, 在PWT4和WTK3/Rwt4同时存在的情况下WTN1被激活, 发挥离子通道作用, 其中WTN1的第1个α螺旋参与膜上通道的形成, 该区域被删除后离子通道活性下降, 细胞死亡受到抑制。
这些结果表明, 在麦类物种中, 串联激酶蛋白与NLR协同调控植物的抗病性, 串联激酶蛋白通过发挥感知(sensor)作用, 识别病原菌效应因子, 介导与其“成对”发挥免疫执行功能(executor) NLR的激活。系统发育分析表明, 多种禾本科植物中均存在这种“sensor-executor”, 二者协同行使功能调控植物的抗病性。
同期, Chen等(2025)的研究也表明, 小麦的串联激酶Sr62TK与距其20.4 kb的Sr62NLR以“sensor- executor”形式成对地在小麦中识别小麦秆锈菌(Puccinia graminis f. sp. tritici, Pgt)效应因子AvrSr62, 进而在调控秆锈病抗性中发挥作用。上述两项研究均表明, 串联激酶负责感知病原菌效应因子并介导与其协作的NLR激活和作用发挥(尽管二者在具体激活机制上存在差异)。
这种通过TKPs-NLRs成对调控植物的抗病性丰富了目前人们对NLR以单个蛋白、成对及成网状行使功能的认识, 为作物抗病育种及性状塑造提供了新的思路和策略。
植物通过效应因子触发的免疫反应(ETI)来抵御病原微生物的侵染, 这一过程由细胞内免疫受体NLRs蛋白介导, 引发局部细胞死亡和活性氧(ROS)爆发, 从而赋予植物强大的抗病能力(Jones et al., 2016; Jian et al., 2023)。在NLRs与病原菌长期进化过程中, NLRs表现出对病原菌识别的特异性; 此外, 特定效应因子在某类植物病原菌中又表现出保守性。因此, 将现有关于NLRs蛋白和效应因子激活ETI的认知应用于作物抗病育种, 已成为植物免疫研究和应用的重要方向(Tai et al., 1999; Liu et al., 2021)。
NLRs蛋白在跨物种行使功能时有2个特点, 即抗病基因只能在其近缘物种中或者同一科内(intrafamily)发挥作用, 跨科发挥作用时存在物种限制(restricted taxonomic functionality, RTF) (Tai et al., 1999)。这与细胞表面受体在植物跨科(interfamily)发挥作用有明显不同。例如, 将拟南芥中能识别青枯菌翻译延伸因子(translation elongation factor, EF-Tu)的细胞表面受体EFR, 转移至茄科植物或其它作物中能够增强植物的抗病能力(Lacombe et al., 2010; Schoonbeek et al., 2015; Schwessinger et al., 2015)。但NLRs跨物种行使功能受到明显限制。例如, 黄单胞菌种广泛存在一类效应因子AvrBs2, 来自辣椒(Capsicum annuum)中的NLR蛋白Bs2能识别来自其病原黄单胞菌辣椒斑点病变种(Xanthomonas campe- stris pv. vesicatoria, Xcv)的效应因子AvrBs2, 将AvrBs2和Bs2转移至其近缘物种番茄(Solanum lycopersicum)中能提高番茄对Xcv的抗性; 但将Bs2转移至亲缘关系较远的木薯(Maihot esculenta)和拟南芥中时, 尽管二者的致病菌油菜黄单胞菌(X. campestris pv. campestris, Xcc)和地毯草黄单胞菌(X. axonopodis pv. manihotis, Xam)也能够分泌该效应因子, 但是转基因材料并未表现出抗性(Tai et al., 1999; Zhao et al., 2011; Li et al., 2015; Medina et al., 2018)。此外, 来自致病疫霉菌(Phytophthora infestans)的效应蛋白AvrAmr3在致病疫霉菌中广泛存在, 这些病原菌侵染包括大豆及茄科和十字花科等在内的植物, 但AvrAmr3只能被光果龙葵(S. americanum)中依赖于NRCs的Rpi-amr3识别, 激活免疫反应(Lin et al., 2020; Oh et al., 2024)。围绕抗病蛋白跨物种行使功能, 克服物种限制, 辅助作物抗病育种, 中国农业大学郭海龙团队提出了将感受型与辅助型NLRs共同转移, 以提高被子植物的抗病性新策略(图1) (Du et al., 2025)。
为测试抗病蛋白跨物种行使功能的可行性, 研究者进行了一系列实验。首先克隆了来自水稻黄单胞菌稻生致病变种(X. oryzae pv. oryzicola, Xoc)的效应蛋白AvrBs2Xoc, 与AvrBs2Xcv相似, 在烟草中共表达AvrBs2Xoc和Bs2能诱导细胞死亡, 表明来自不同病原微生物的AvrBs2能被感受型NLRs——Bs2识别。将AvrBs2Xoc和Bs2在水稻(Oryza sativa)原生质体中共表达时, 并不能诱导原生质体死亡。原因是烟草中Bs2的功能发挥依赖于下游的辅助型NLRs——NbNRC2/ 3/4, 当共转入NRCs时, AvrBs2Xoc和Bs2的识别便能诱导水稻原生质体死亡。类似地, 在大豆原生质体中, 共表达AvrBs2Xcv、Bs2和NRC2/3/4也能诱导细胞死亡。此外, 将能识别AvrAmr1, 来自茄科植物光果龙葵的晚疫病抗性基因Rpi-amr1及其发挥功能依赖的SaNRC2/3, 在大豆原生质体中共表达也能诱导细胞死亡。AvrAmr3和Rpi-amr3也能以SaNRC2/3/4依赖的方式促进大豆原生质体细胞死亡。感受型NLRs (Rpi-amr3/Bs2)或感受型+辅助型NLRs (NbNRC2/SaNR-C2)的拟南芥转基因株系中, 利用无效应因子的丁香假单胞菌番茄致病变种DC3000 (Pseudomonas sy- ringae pv. tomato DC3000 D36E)分泌效应蛋白(AvrAmr3/AvrBs2)到植物细胞中时, 单独表达感受型NLRs的转基因株系并不能被激活行使功能, 只有当辅助型NLRs存在时, 效应因子与感受型NLRs的识别才能引起细胞死亡。这些结果表明, 茄科植物的感受型NLRs在识别效应因子后可通过与其匹配的辅助型NLRs协作, 在其远缘植物中实现跨物种行使功能。
由水稻黄单胞菌稻生致病变种(Xoc)引起的水稻细菌性条斑病(bacterial leaf streak, BLS)是威胁我国粮食安全的重要病害之一。早期研究表明, 玉米抗病基因Rxo1能识别Xoc携带的AvrRxo1效应蛋白(III型分泌系统效应子), 并触发抗性反应, 在水稻中能抑制病原菌的生长。该基因可用于水稻抗病育种, 防治细菌性条斑病(Zhao et al., 2005)。Xoc能分泌AvrBs2Xoc, 利用感受型与辅助型NLRs模块跨物种行使功能。研究者构建了表达Bs2与不同NbNRCs组合的转基因水稻, 结果显示, 在无病原菌侵染时, 转基因植株与野生型无显著差异; 而在Xoc侵染时, 转基因植株表现出更强的抗病性, 病斑减小且病原菌积累量降低。
这种感受型与辅助型NLRs在跨物种行使功能时依赖每个部分的完整性。当NbNRCs中CC结构域突变时, 转基因材料抗性丧失; 敲除Xoc中的AvrBs2时, 转基因株系恢复与野生型相似的病原菌敏感性; 单独表达Bs2或与NbNRCs共表达但不提供AvrBs2时, 水稻对病原菌的敏感性不变。此外, AvrBs2被Bs2识别后能诱导植物下游的NbNRCs寡聚化。表明感受型与辅助型NLRs在跨物种行使功能时机制保守。
值得注意的是, 这种感受型与辅助型NLRs模块转移至水稻中并不影响水稻本身的株高、分蘖数和千粒重等农艺性状, 也不改变水稻的基础免疫响应及对稻瘟病的敏感性, 这些结果进一步展示了这种感受型与辅助型NLRs在跨物种发挥抗病作用中的应用价值。
综上所述, 通过感受型与辅助型NLRs, 可实现跨物种利用抗病蛋白的功能, 为目前缺乏内源抗病基因的作物抗病育种提供了新思路。未来可进一步探索不同作物中保守的辅助型NLRs系统, 优化感受型与辅助型NLRs模块设计, 从而开发出更高效、广谱的抗病作物新品种。
植物免疫相关基因的克隆及其作用机制的解析为培育具有广谱、持续抗病性作物新品种奠定了重要理论基础。通过工程化手段改良或创造新型免疫识别特异性一直是植物病理学和作物育种领域的核心目标。早期的免疫受体设计主要依赖随机诱变、嵌合受体构建和结构域交换等传统技术(Wulff et al., 2001; Segretin et al., 2014; Steinbrenner et al., 2015)。随着对植物免疫机制的深入理解及结构生物学的快速发展, 理性改造植物免疫受体已成为创制新型抗病性状的关键策略(Tamborski and Krasileva, 2020)。
受体-配体复合物(如FLS2-flg22和Sr50-AvrSr50)的高分辨率结构解析为LRR结构域的工程化改造奠定了重要基础。这些精确的结构信息显著缩小了靶点筛选范围, 对LRR结构域中参与配体结合或决定识别特异性的关键氨基酸进行精准修饰, 可实现受体识别的定向改造和扩展(Sun et al., 2013; Wei et al., 2020; Ortiz et al., 2022; Tamborski et al., 2023)。
许多植物NLR蛋白通过“guard”机制监测病原菌效应蛋白对植物靶蛋白的修饰来激活免疫反应(Cesari et al., 2014; Wang et al., 2015)。这一特性已被成功应用于受体设计。例如, 水稻Pik-1/Pik-2中的Pik-1蛋白含有重金属相关结构域(heavy metal-associated, HMA), 通过将其改造为NLR免疫受体——纳米体融合蛋白, 与Pik-2结合可赋予植物新的抗病能力(Kourelis et al., 2023)。另一典型模型是RPS5监测RLCK激酶PBS1的完整性, 当病原体分泌的蛋白酶切割PBS1时, RPS5即被激活。基于此原理, 通过改造PBS1等诱饵蛋白可有效扩展NLR介导的抗病识别范围(DeYoung et al., 2012; Narusaka et al., 2014; Mukhi et al., 2021)。
清华大学刘玉乐团队最新研究进展提出了一种突破性的植物免疫工程策略: 通过对自激活型NLR受体(autoactive NLR, aNLR)进行改造, 可实现植物广谱、持久和完全的病害抗性。在自激活NLR蛋白的N端融合一段含有病原菌蛋白酶切割位点(protease cleavage site, PCS)的柔性多肽, 形成嵌合蛋白。当病原体入侵时, 其特异性蛋白酶切割该嵌合蛋白, 释放游离的aNLR结构域, 从而触发植物的免疫反应(图1) (Wang et al., 2025b)。
他们构建的嵌合蛋白HA-PCSPVY-aTm-2²含有马铃薯Y病毒(potato virus Y, PVY)的PCS和自激活CNL蛋白aTm-2², 嵌合蛋白单独表达时不诱导细胞死亡, 但在与PVY的NIa蛋白酶共表达时能显著诱导细胞死亡。实验观察到蛋白积累量减少, 同时带有C端HA标签的嵌合蛋白被切割为更小的aTm-2²-HA片段, 证实切割后释放的aTm-2²激活了免疫反应。转基因烟草(HA-PCSPVY-aTm-2²)接种PVY-GFP 21天后未检测到病毒荧光和症状, RT-PCR也未检出病毒RNA, 而野生型植物严重感病, 表明该嵌合蛋白能赋予植物对PVY的完全抗性。进一步研究发现, 转基因烟草对芜菁花叶病毒(turnip mosaic virus, TuMV)、辣椒斑驳病毒(pepper mottle virus, PepMoV)、辣椒脉斑驳病毒(chilli veinal mottle virus, ChiVMV)以及李痘病毒(plum pox virus, PPV)均表现出完全抗性, 接种21天后无病毒积累, 抗性率达70%-96%, 证实该设计策略能实现广谱抗性。
同样, 他们构建了含有PCSPVY和自激活RNL蛋白aAtNRG1.1的嵌合蛋白HA-PCSPVY-aAtNRG1.1, 转基因本氏烟草对PVY及多种马铃薯(S. tuberosum) Y病毒属病毒表现出极端抗性或经典完全抗性, 并能够抵抗混合感染。将烟草蚀纹病毒(tobacco etch potyvirus, TEV)的PCS (PCSTEV)与PCSPVY串联, 构建HA-PCSTEV-PCSPVY-aAtNRG1.1后, 转基因本氏烟草对PVY和TEV均表现抗性, 部分植株对TEV的完全抗性率达55%。在大豆中, 表达含大豆花叶病毒(soybean mosaic virus, SMV) PCS的嵌合蛋白HA-PCSSMV-aAtNRG1.1的转基因大豆对SMV表现完全抗性, 并且农艺性状正常, 表明这种策略在农作物中同样适用。
与现有“诱饵工程”等方法相比, 该策略无需依赖特定NLR蛋白, 具有操作简单、适用范围广和抗性持久等优势, 未来有望扩展至更多病原体的防控。结合AlphaFold预测和进化分析等新技术, 受体改造工程的发展将进一步推动广谱抗病性研究; 结合传统遗传学和分子生物学手段, 有望在从模式植物到农作物的应用中实现更高效的抗病育种, 为农业可持续发展提供新的解决方案。
肖银燕: 撰写论文, 绘图, 修订文章内容; 于华: 撰写论文, 绘图; 万里: 修订文章内容及审校图, 提供资金和技术支持。
  • 国家自然科学基金面上项目(32270304)
参考文献 引证文献
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2025年第60卷第5期
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doi: 10.11983/CBB25131
  • 接收时间:2025-07-23
  • 首发时间:2026-10-22
  • 出版时间:2025-09-10
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  • 收稿日期:2025-07-23
  • 录用日期:2025-09-02
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国家自然科学基金面上项目(32270304)
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    中国科学院分子植物科学卓越创新中心, 上海 200032

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万里, 中国科学院分子植物科学卓越创新中心研究员, 博士生导师, 入选国家青年千人计划, 获得上海市植物病理学会杰出青年科学家奖。万里课题组专注于植物免疫机制研究和广谱抗性农作物创制, 在植物细胞内免疫受体NLR工作的分子机理方面取得重要进展。相关成果以第一作者(含共同)发表在Science (2014, 2019), 以通讯作者(含共同)发表在Nature、Science、Proc Natl Acad Sci USA、Nature Plants、Science Advances和J Integr Plant Biol等学术期刊。

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2种不同金属材料的力学参数
科
Family
属数
Number of
genus
种数
Number of
species
占总种数比例
Percentage of
total species (%)
属
Genus
种数
Number of
species
占总种数比例
Percentage of total
species (%)
鹅膏菌科Amanitaceae 2 11 5.26 鹅膏菌属 Amanita 10 4.78
小菇科 Mycenaceae 2 12 5.74 丝盖伞属 Inocybe 5 2.39
多孔菌科 Polyporaceae 8 14 6.70 蜡蘑属 Laccaria 5 2.39
红菇科 Russulaceae 3 23 11.00 小皮伞属 Marasmius 6 2.87
小菇属 Mycena 11 5.26
光柄菇属 Pluteus 5 2.39
红菇属 Russula 17 8.13
栓菌属 Trametes 5 2.39
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