Di Zhang, Liang Li, Chunxue Wang, Nuo Zhou, Zhilei Yu, Tianyi Shen, Baifan Wang, Qiang Bian, Dejun Ma, Yucheng Gu, Han Xu, Zhen Xi
The development of dual‐target inhibitors represents a cost‐effective strategy for integrated pest management. Here, we report the first dual‐target inhibitors designed against the evolutionarily conserved domain of phytoene synthase (PSY), a key enzyme in carotenoid biosynthesis. Using comparative genomics, we identified structural conservation between PSY in plants and squalene synthase (erg9) in fungi. Through virtual screening and structure‐based optimization of compounds targeting PSY, we identified lead compound 1c , which exhibited potent herbicidal and fungicidal activity. In vitro binding assays confirmed that 1c binds to both PSY and erg9. In plants, 1c treatment reduced chlorophyll content, downregulated photosynthesis‐associated genes, and caused substrate accumulation in the carotenoid pathway. In fungi, 1c induced a mycelial morphology identical to erg9 knockout mutants. Molecular dynamics simulations revealed the differential binding conformations of 1c to PSY and erg9, elucidating its mode of action. This work establishes PSY and its homologues as a promising target for the development of novel, broad‐spectrum dual‐action agrochemicals based on targetome structural similarity.
{"title":"A Novel Dual‐Target Compound Designed With Potent Herbicidal and Fungicidal Activity Inspired by Conserved Phytoene Synthase Domains","authors":"Di Zhang, Liang Li, Chunxue Wang, Nuo Zhou, Zhilei Yu, Tianyi Shen, Baifan Wang, Qiang Bian, Dejun Ma, Yucheng Gu, Han Xu, Zhen Xi","doi":"10.1111/pbi.70531","DOIUrl":"https://doi.org/10.1111/pbi.70531","url":null,"abstract":"The development of dual‐target inhibitors represents a cost‐effective strategy for integrated pest management. Here, we report the first dual‐target inhibitors designed against the evolutionarily conserved domain of phytoene synthase (PSY), a key enzyme in carotenoid biosynthesis. Using comparative genomics, we identified structural conservation between PSY in plants and squalene synthase (erg9) in fungi. Through virtual screening and structure‐based optimization of compounds targeting PSY, we identified lead compound 1c , which exhibited potent herbicidal and fungicidal activity. In vitro binding assays confirmed that 1c binds to both PSY and erg9. In plants, 1c treatment reduced chlorophyll content, downregulated photosynthesis‐associated genes, and caused substrate accumulation in the carotenoid pathway. In fungi, 1c induced a mycelial morphology identical to erg9 knockout mutants. Molecular dynamics simulations revealed the differential binding conformations of 1c to PSY and erg9, elucidating its mode of action. This work establishes PSY and its homologues as a promising target for the development of novel, broad‐spectrum dual‐action agrochemicals based on targetome structural similarity.","PeriodicalId":221,"journal":{"name":"Plant Biotechnology Journal","volume":"2 1","pages":""},"PeriodicalIF":13.8,"publicationDate":"2026-01-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145920215","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
HaiSheng Zeng, MeiHui Shi, ZhiRong Chen, XueJing Sun, HuiJie Zhang, Yue Huang, YuCheng Chen, Jun Ren, HuiLing Huang, Almaz Borjigidai, Man Zhang, SuJuan Duan, Yi‐Jun Chen, Hong‐Lei Jin, Hong‐Bin Wang
The synthesis and accumulation of active ingredients in medicinal plants are distributed in specific organs, tissues, and cell types, which are important for the exploitation of medicinal plants. However, the fine distribution of active ingredients is difficult to know. Here, the system of mass spectrometry imaging (MSI) integrated with single‐cell RNA sequencing was established for the first time in Andrographis paniculata ( A. paniculata ), a medicinal plant widely utilised in China and Southeast Asia. MSI shows specific distribution of andrographolides in A. paniculata , with higher accumulation in non‐veinal leaf regions and outer stem cortex (leaf > stem; outer > inner cortex), as validated by LC‐QQQ‐MS/MS assays. Leaf scRNA‐seq demonstrates that ApCPS2 (the key terpene synthase for andrographolide biosynthesis) exhibits pronounced cell‐type‐specific expression in photosynthetic mesophyll subclusters, indicating mesophyll cells as the primary site for light‐modulated andrographolide production. Interestingly, light may enhance the accumulation of andrographolide biosynthesis, confirming the light sensitivity of metabolism in mesophyll cells. This study explores medicinal components' multidimensional spatial distributions and biosynthetic pathways in A. paniculata via MSI combined with single‐cell technology, providing a novel strategy for determining plant metabolites' fine synthesis and distribution.
药用植物有效成分的合成和积累分布在特定的器官、组织和细胞类型中，这对药用植物的开发具有重要意义。然而，有效成分的精细分布很难知道。本文首次对中国和东南亚广泛使用的药用植物穿心莲（Andrographis paniculata, a. paniculata）建立了质谱成像（MSI）与单细胞RNA测序相结合的系统。MSI显示穿心莲内酯在穿心莲中的特定分布，在非脉叶区域和外茎皮层（叶&茎；外&茎；内皮层）积累较多，LC - QQQ - MS/MS分析证实了这一点。叶片scRNA - seq表明ApCPS2（穿心莲内酯生物合成的关键萜烯合成酶）在光合叶肉亚群中表现出明显的细胞类型特异性表达，表明叶肉细胞是光调节穿心莲内酯生产的主要位点。有趣的是，光可能促进穿心莲内酯生物合成的积累，证实了叶肉细胞代谢的光敏感性。本研究通过MSI结合单细胞技术，探索了金针叶药用成分的多维空间分布和生物合成途径，为确定植物代谢产物的精细合成和分布提供了一种新的策略。
{"title":"Mass Spectrometry Imaging Combined With Single‐Cell Transcriptional Profiling Reveals the Multidimensional Spatial Distributions and Biosynthetic Pathways of Medicinal Components in Andrographis paniculata ","authors":"HaiSheng Zeng, MeiHui Shi, ZhiRong Chen, XueJing Sun, HuiJie Zhang, Yue Huang, YuCheng Chen, Jun Ren, HuiLing Huang, Almaz Borjigidai, Man Zhang, SuJuan Duan, Yi‐Jun Chen, Hong‐Lei Jin, Hong‐Bin Wang","doi":"10.1111/pbi.70534","DOIUrl":"https://doi.org/10.1111/pbi.70534","url":null,"abstract":"The synthesis and accumulation of active ingredients in medicinal plants are distributed in specific organs, tissues, and cell types, which are important for the exploitation of medicinal plants. However, the fine distribution of active ingredients is difficult to know. Here, the system of mass spectrometry imaging (MSI) integrated with single‐cell RNA sequencing was established for the first time in <jats:styled-content style=\"fixed-case\"> <jats:italic>Andrographis paniculata</jats:italic> </jats:styled-content> ( <jats:styled-content style=\"fixed-case\"> <jats:italic>A. paniculata</jats:italic> </jats:styled-content> ), a medicinal plant widely utilised in China and Southeast Asia. MSI shows specific distribution of andrographolides in <jats:styled-content style=\"fixed-case\"> <jats:italic>A. paniculata</jats:italic> </jats:styled-content> , with higher accumulation in non‐veinal leaf regions and outer stem cortex (leaf > stem; outer > inner cortex), as validated by LC‐QQQ‐MS/MS assays. Leaf scRNA‐seq demonstrates that <jats:italic>ApCPS2</jats:italic> (the key terpene synthase for andrographolide biosynthesis) exhibits pronounced cell‐type‐specific expression in photosynthetic mesophyll subclusters, indicating mesophyll cells as the primary site for light‐modulated andrographolide production. Interestingly, light may enhance the accumulation of andrographolide biosynthesis, confirming the light sensitivity of metabolism in mesophyll cells. This study explores medicinal components' multidimensional spatial distributions and biosynthetic pathways in <jats:styled-content style=\"fixed-case\"> <jats:italic>A. paniculata</jats:italic> </jats:styled-content> via MSI combined with single‐cell technology, providing a novel strategy for determining plant metabolites' fine synthesis and distribution.","PeriodicalId":221,"journal":{"name":"Plant Biotechnology Journal","volume":"3 1","pages":""},"PeriodicalIF":13.8,"publicationDate":"2026-01-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145920246","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
The regulation of serotonin metabolism during fruit development and ripening remains poorly understood, despite its potential roles in plant defence and human nutrition. Here, we demonstrated that the MADS‐box transcription factor FUL2 acts as a key repressor of serotonin accumulation in tomato by forming a functional module with MADS1 . CRISPR‐Cas9‐generated ful2‐cr mutants exhibited delayed ripening, reduced fruit size and a striking 10‐fold increase in serotonin levels, suggesting a previously unrecognised link between FUL2 and secondary metabolism. Immunoprecipitation‐mass spectrometry (IP‐MS) revealed that FUL2 physically interacts with MADS1, and genetic analyses showed that mads1‐cr mutants phenocopied both the developmental and serotonin hyperaccumulation phenotypes of ful2‐cr mutants. Furthermore, ChIP‐seq and transcriptomic profiling demonstrated that the FUL2‐MADS1 complex directly binds CArG‐box motifs in the promoter of ASMT5 (a key enzyme in serotonin‐to‐melatonin conversion), activating its expression while repressing TDC1 (tryptophan decarboxylase). Electrophoretic mobility shift assays (EMSA) and dual‐luciferase reporter assays confirmed their cooperative DNA binding and synergistic transcriptional regulation. Our work establishes a MADS‐box transcriptional module that gates serotonin flux by coordinately regulating biosynthetic and metabolic genes. These findings provided a framework for engineering serotonin content in crops and deepen understanding of how developmental transcription factors govern specialised metabolism during ripening.
{"title":"Tandem MADS ‐Box Genes FUL2 and MADS1 Form a Regulatory Module to Repress Serotonin Biosynthesis via Direct ASMT5 Activation in Tomato Fruit","authors":"Yaping Xu, Huimin Jia, Dengguo Tang, Lijun Zhang, Xinqin Liu, Junjie Rong, Yunai Lv, Zhaobo Lang, Qingfeng Niu","doi":"10.1111/pbi.70538","DOIUrl":"https://doi.org/10.1111/pbi.70538","url":null,"abstract":"The regulation of serotonin metabolism during fruit development and ripening remains poorly understood, despite its potential roles in plant defence and human nutrition. Here, we demonstrated that the MADS‐box transcription factor <jats:italic>FUL2</jats:italic> acts as a key repressor of serotonin accumulation in tomato by forming a functional module with <jats:italic>MADS1</jats:italic> . CRISPR‐Cas9‐generated <jats:italic>ful2‐cr</jats:italic> mutants exhibited delayed ripening, reduced fruit size and a striking 10‐fold increase in serotonin levels, suggesting a previously unrecognised link between FUL2 and secondary metabolism. Immunoprecipitation‐mass spectrometry (IP‐MS) revealed that FUL2 physically interacts with MADS1, and genetic analyses showed that <jats:italic>mads1‐cr</jats:italic> mutants phenocopied both the developmental and serotonin hyperaccumulation phenotypes of <jats:italic>ful2‐cr</jats:italic> mutants. Furthermore, ChIP‐seq and transcriptomic profiling demonstrated that the FUL2‐MADS1 complex directly binds CArG‐box motifs in the promoter of <jats:italic>ASMT5</jats:italic> (a key enzyme in serotonin‐to‐melatonin conversion), activating its expression while repressing <jats:italic>TDC1</jats:italic> (tryptophan decarboxylase). Electrophoretic mobility shift assays (EMSA) and dual‐luciferase reporter assays confirmed their cooperative DNA binding and synergistic transcriptional regulation. Our work establishes a MADS‐box transcriptional module that gates serotonin flux by coordinately regulating biosynthetic and metabolic genes. These findings provided a framework for engineering serotonin content in crops and deepen understanding of how developmental transcription factors govern specialised metabolism during ripening.","PeriodicalId":221,"journal":{"name":"Plant Biotechnology Journal","volume":"162 1","pages":""},"PeriodicalIF":13.8,"publicationDate":"2026-01-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145920346","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Efficient viral proliferation within the host is a critical step in pathogenicity and requires adenosine triphosphate (ATP). The replication, movement and immune evasion of many plant viruses within their hosts are associated with phase separation (PS)‐derived aggregates formed by viral components. However, the host factors that drive the formation of these condensates remain largely unknown. This study provides evidence that the nucleocapsid protein (N) of tomato spotted wilt virus (TSWV) recruits the host factor phosphoglycerate kinase (NbPGK) from Nicotiana benthamiana to form phase‐separated condensates. This remodels the host glycolytic pathway to generate ATP, supplying energy for viral replication via ribonucleoprotein complexes and acting as a promoter to regulate the PS network, thereby facilitating condensate formation. Notably, we have developed a small‐molecule PS modulator, F10 . By combining drug affinity‐responsive target stability, molecular docking, microscale thermophoresis and bio‐layer interferometry techniques allowed F10 , we confirmed binding to sites Arg94, Lys192 and Gly228 on TSWV N, residues critical for maintaining NbPGK recruitment. F10 interacts with N, liberating the hijacked host factor NbPGK, and exhibits potent antiviral activity, outperforming the commercial virucide Ningnanmycin. This study elucidates the molecular machinery underlying viral exploitation of host cellular metabolism and identifies a lead compound that is amenable to managing TSWV by targeting this process.
{"title":"Tomato Spotted Wilt Virus Reprogrammes Host Glycolysis to Facilitate Proliferation by a Phase‐Separated Co‐Aggregate of Nucleocapsid Protein and Phosphoglycerate Kinase","authors":"Guangcheng Zu, Zhifu Xing, Jiao Li, Tangbing Yang, Huan Wu, Qiangsheng Ge, Yanju Wang, Baoan Song, Runjiang Song","doi":"10.1111/pbi.70529","DOIUrl":"https://doi.org/10.1111/pbi.70529","url":null,"abstract":"Efficient viral proliferation within the host is a critical step in pathogenicity and requires adenosine triphosphate (ATP). The replication, movement and immune evasion of many plant viruses within their hosts are associated with phase separation (PS)‐derived aggregates formed by viral components. However, the host factors that drive the formation of these condensates remain largely unknown. This study provides evidence that the nucleocapsid protein (N) of tomato spotted wilt virus (TSWV) recruits the host factor phosphoglycerate kinase (NbPGK) from <jats:italic>Nicotiana benthamiana</jats:italic> to form phase‐separated condensates. This remodels the host glycolytic pathway to generate ATP, supplying energy for viral replication via ribonucleoprotein complexes and acting as a promoter to regulate the PS network, thereby facilitating condensate formation. Notably, we have developed a small‐molecule PS modulator, F10 . By combining drug affinity‐responsive target stability, molecular docking, microscale thermophoresis and bio‐layer interferometry techniques allowed F10 , we confirmed binding to sites Arg94, Lys192 and Gly228 on TSWV N, residues critical for maintaining NbPGK recruitment. F10 interacts with N, liberating the hijacked host factor NbPGK, and exhibits potent antiviral activity, outperforming the commercial virucide Ningnanmycin. This study elucidates the molecular machinery underlying viral exploitation of host cellular metabolism and identifies a lead compound that is amenable to managing TSWV by targeting this process.","PeriodicalId":221,"journal":{"name":"Plant Biotechnology Journal","volume":"29 1","pages":""},"PeriodicalIF":13.8,"publicationDate":"2026-01-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145903587","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Plant leaves originate from the shoot apical meristem (SAM) and undergo a developmental process of highly coordinated gene expression regulation. To date, only a few key regulators have been identified and characterised, so the gene expression cascades responsible for leaf cell specification and differentiation from SAM remain largely elusive. Here, we optimised a spatial transcriptomics protocol using the 10× Genomics Visium system and developed computational pipelines to reconstruct three-dimensional gene expression profiles of the SAM and sequentially developing leaves in maize seedlings. These enabled positional indexing of cells sampled from consecutive developmental stages, revealing dynamic transitions from undifferentiated stem cells in the SAM to functionally differentiated leaf structures. Through spatial–temporal transcriptome analysis, we identified distinct transcriptional programs and key regulatory genes involved in meristem maintenance, leaf primordia initiation, vascular tissue differentiation, and cellular heterogeneity. This approach outperforms the single-cell transcriptome profiling, which lacks temporal and spatial contexts. Our optimised experimental pipeline, which goes from section preparation to data processing, enables the spatial resolution and 3-dimensional mapping of gene expression profiles. The established pipeline is readily applicable to delineating molecular events underlying developmental transitions, cell type specifications, and differentiation in plants.
{"title":"Serial Spatial Transcriptomes Reveal Regulatory Transitions in Maize Leaf Development","authors":"Chi-Chih Wu, Ludvig Larsson, Kun-Ting Hsieh, Chun-Ping Yu, Yi-Hua Chen, Kai-Hsuan Ding, Ho-Chun Yang, Joakim Lundeberg, Chin-Min Kimmy Ho, Shu-Hsing Wu, Mei-Yeh Jade Lu, Wen-Hsiung Li","doi":"10.1111/pbi.70515","DOIUrl":"https://doi.org/10.1111/pbi.70515","url":null,"abstract":"Plant leaves originate from the shoot apical meristem (SAM) and undergo a developmental process of highly coordinated gene expression regulation. To date, only a few key regulators have been identified and characterised, so the gene expression cascades responsible for leaf cell specification and differentiation from SAM remain largely elusive. Here, we optimised a spatial transcriptomics protocol using the 10× Genomics Visium system and developed computational pipelines to reconstruct three-dimensional gene expression profiles of the SAM and sequentially developing leaves in maize seedlings. These enabled positional indexing of cells sampled from consecutive developmental stages, revealing dynamic transitions from undifferentiated stem cells in the SAM to functionally differentiated leaf structures. Through spatial–temporal transcriptome analysis, we identified distinct transcriptional programs and key regulatory genes involved in meristem maintenance, leaf primordia initiation, vascular tissue differentiation, and cellular heterogeneity. This approach outperforms the single-cell transcriptome profiling, which lacks temporal and spatial contexts. Our optimised experimental pipeline, which goes from section preparation to data processing, enables the spatial resolution and 3-dimensional mapping of gene expression profiles. The established pipeline is readily applicable to delineating molecular events underlying developmental transitions, cell type specifications, and differentiation in plants.","PeriodicalId":221,"journal":{"name":"Plant Biotechnology Journal","volume":"38 1","pages":""},"PeriodicalIF":13.8,"publicationDate":"2026-01-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145903584","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Histone H3 lysine 27 trimethylation (H3K27me3) is essential for fungal pathogenicity, yet its contribution to pathogen–host interactions remains incompletely understood. Here, we profiled H3K27me3 dynamics in Fusarium graminearum during infection and identified 132 H3K27me3‐marked genes ( FgHMGs ). Among these, FgHMG1 encodes a secreted glycoside hydrolase family 11 (GH11) protein that functions as a pathogen‐associated molecular pattern (PAMP), triggering PAMP‐triggered immunity (PTI) in Nicotiana benthamiana through the receptor kinases BAK1 and SOBIR1, independently of its enzymatic activity. FgHMG1 also induces reactive oxygen species (ROS) accumulation and upregulation of defence‐related genes in wheat plants. Remarkably, FgHMG1 expression is repressed during host invasion by the histone methyltransferase FgKMT6, a homologue of Enhancer of zeste (E(z)) from Drosophila, via H3K27me3 deposition, enabling immune evasion. Loss of FgKMT6 abolishes H3K27me3 enrichment, derepresses FgHMG1, and enhances host immunity, effects largely rescued in ΔFgKMT6–FgHMG1 double mutants. Notably, foliar application of recombinant FgHMG1 protein reduced Fusarium head blight severity in wheat by 35%–50% in 2‐year field trials. These findings reveal that fungal pathogens exploit H3K27me3‐mediated silencing of immunogenic PAMP genes to evade host recognition and highlight FgHMG1 as a promising candidate for crop protection.
{"title":"H3K27me3 ‐Mediated Epigenetic Silencing of FgHMG1 Enables Fungal Host Immune Evasion","authors":"Xiaozhen Zhao, Bingqin Yuan, Peixue Ma, Yajie Cai, Yan Huang, Yaxuan Wang, Zhen Cheng, Yuan Chen, Minghong Zheng, Ran Zhang, Jinmei Wu, Xieyu Li, Mohan Wang, Huijun Wu, Chengqi Zhang, Xuewen Gao, Li Chen, Qin Gu","doi":"10.1111/pbi.70530","DOIUrl":"https://doi.org/10.1111/pbi.70530","url":null,"abstract":"Histone H3 lysine 27 trimethylation (H3K27me3) is essential for fungal pathogenicity, yet its contribution to pathogen–host interactions remains incompletely understood. Here, we profiled H3K27me3 dynamics in <jats:italic>Fusarium graminearum</jats:italic> during infection and identified 132 H3K27me3‐marked genes ( <jats:italic>FgHMGs</jats:italic> ). Among these, <jats:italic>FgHMG1</jats:italic> encodes a secreted glycoside hydrolase family 11 (GH11) protein that functions as a pathogen‐associated molecular pattern (PAMP), triggering PAMP‐triggered immunity (PTI) in <jats:italic>Nicotiana benthamiana</jats:italic> through the receptor kinases BAK1 and SOBIR1, independently of its enzymatic activity. FgHMG1 also induces reactive oxygen species (ROS) accumulation and upregulation of defence‐related genes in wheat plants. Remarkably, FgHMG1 expression is repressed during host invasion by the histone methyltransferase FgKMT6, a homologue of Enhancer of zeste (E(z)) from Drosophila, via H3K27me3 deposition, enabling immune evasion. Loss of FgKMT6 abolishes H3K27me3 enrichment, derepresses FgHMG1, and enhances host immunity, effects largely rescued in <jats:italic>ΔFgKMT6–FgHMG1</jats:italic> double mutants. Notably, foliar application of recombinant FgHMG1 protein reduced Fusarium head blight severity in wheat by 35%–50% in 2‐year field trials. These findings reveal that fungal pathogens exploit H3K27me3‐mediated silencing of immunogenic PAMP genes to evade host recognition and highlight FgHMG1 as a promising candidate for crop protection.","PeriodicalId":221,"journal":{"name":"Plant Biotechnology Journal","volume":"11 1","pages":""},"PeriodicalIF":13.8,"publicationDate":"2026-01-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145903716","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
<p>Tanshinone and phenolic acid are key therapeutic compounds in the medicinal plant <i>Salvia miltiorrhiza</i>, while rutin is the major bioactive metabolite in the medicinal plant <i>Fagopyrum dibotrys</i>. However, their natural levels in cultivated varieties remain low, limiting their pharmacological potential. Enhancing metabolite accumulation through the modification of upstream transcription factors and key biosynthetic enzymes has shown promise (Deng et al. <span>2020</span>). Compounding the challenge, medicinal plants like <i>S. miltiorrhiza</i> are perennials with long life cycles, making conventional breeding inefficient. While CRISPR-Cas9 has facilitated gene knockout strategies in medicinal plants, precise base editing technologies remain underutilised (Das et al. <span>2024</span>). Prime editing (PE) has emerged as a powerful tool for introducing targeted nucleotide changes; thus, offering a promising route for molecular breeding (Anzalone et al. <span>2019</span>). While their application in medicinal plants remains largely unexplored.</p> <p>To address these limitations in genome engineering of medicinal plants, we develop the prime editor in medicinal plants. Our previous work demonstrated that N-terminus M-MLV RT fused to Cas9 nickase plus epegRNA prime editor (NEPE) is highly efficient in rice (Zhong et al. <span>2024</span>), indicating that the N-terminal fusion of M-MLV reverse transcriptase is particularly well-suited for plant applications. Building on this, and with optimization of the expression system, we further adapted the prime editor for medicinal plants, termed MediPlant-NEPE (Figure 1a; Figure S1).</p> <figure><picture> <source media="(min-width: 1650px)" srcset="/cms/asset/227ed4e7-a506-4d78-a262-b21f293b3a66/pbi70532-fig-0001-m.jpg"/><img alt="Details are in the caption following the image" data-lg-src="/cms/asset/227ed4e7-a506-4d78-a262-b21f293b3a66/pbi70532-fig-0001-m.jpg" loading="lazy" src="/cms/asset/b14de0f4-3fe0-4aa2-9b5e-5fcefdd071f8/pbi70532-fig-0001-m.png" title="Details are in the caption following the image"/></picture><figcaption> <div><strong>FIGURE 1<span style="font-weight:normal"></span></strong><div>Open in figure viewer<i aria-hidden="true"></i><span>PowerPoint</span></div> </div> <div>Construction and application of MediPlant-NEPE for enhancing metabolic engineering in medicinal plants. (a) Schematic of MediPlant-NEPE. (b) Genotyping of <i>SmbZIP1</i>-edited hairy roots, red arrows indicate edits. (c) Phenotypes of <i>SmMYB36</i> homozygous mutants. (d) Genotyping of <i>SmTCP15</i>-edited lines, red arrows indicate edit sites. (e) Phenotypes of <i>SmTCP15</i> homozygous mutants. (f) HPLC quantification of anthocyanin in mutant flowers. (g) Genotyping of <i>SmbZIP1</i>-edited lines, red arrows indicate mutation sites. (h) Dry root powder of <i>SmbZIP1</i> mutants. (i) HPLC quantification of major tanshinones in roots. (j) Genotyping of <i>SmRAS</i>-edited hairy root lines, amino acid changes
丹参酮和酚酸是药用植物丹参中的关键治疗化合物，而芦丁是药用植物金荞麦的主要生物活性代谢物。然而，它们在栽培品种中的天然含量仍然很低，限制了它们的药理潜力。通过修饰上游转录因子和关键的生物合成酶来增强代谢物的积累已经显示出前景（Deng et al. 2020）。更复杂的是，像丹参这样的药用植物是多年生植物，生命周期长，使得传统的育种效率低下。虽然CRISPR-Cas9促进了药用植物的基因敲除策略，但精确的碱基编辑技术仍未得到充分利用（Das et al. 2024）。引体编辑（PE）已成为引入靶向核苷酸变化的强大工具；因此，为分子育种提供了一条有希望的途径（Anzalone et al. 2019）。而它们在药用植物中的应用在很大程度上仍未被探索。为了解决药用植物基因组工程的这些局限性，我们开发了药用植物基因组编辑器。我们之前的工作表明，n端M-MLV RT与Cas9 nickase + epegRNA引物编辑器（NEPE）融合在水稻中是非常高效的（Zhong et al. 2024），这表明M-MLV逆转录酶的n端融合特别适合植物应用。在此基础上，通过对表达系统的优化，我们进一步调整了药用植物的引物编辑器，称为MediPlant-NEPE（图1a；图S1）。图1打开图形查看器powerpointmediplant - nepe在药用植物代谢工程中的构建与应用(a) MediPlant-NEPE示意图。(b) smbzip1编辑的毛状根的基因分型，红色箭头表示编辑。(c) SmMYB36纯合突变体的表型。(d) smtcp15编辑细胞系的基因分型，红色箭头表示编辑位点。(e) SmTCP15纯合突变体的表型。(f)高效液相色谱法测定突变花中花青素含量。(g) smbzip1编辑细胞系的基因分型，红色箭头表示突变位点。(h) SmbZIP1突变体的干根粉。(1)丹参酮根中主要丹参酮的HPLC定量分析。(j) smras编辑的毛状根系基因分型，氨基酸变化如图所示。(k) SmRASG49A毛状根在液体培养基中培养56天后。(l)高效液相色谱法测定SmRAS突变体中酚酸含量。(m) fdufgt3编辑的F. dibotrys毛状根的基因分型，红色箭头表示编辑。(n)野生型和转基因dibotrys的毛状根培养物（上）和甲醇提取物（下）。(o)高效液相色谱法测定FdUFGT3突变体中槲皮素和芦丁含量。我们首先以丹参中的SmMYB36和SmTCP15为目标，测试MediPlant-NEPE的效率，并研究它们在花色素沉着中的作用。SmMYB36基因Q22位点的靶向C>；T转换和SmTCP15基因E24位点的靶向G>；T转换诱导过早终止密码子。经农杆菌介导转化后，获得了SmMYB36和SmTCP15的阳性转基因系，突变频率分别为21.21%和21.21%（双等位基因突变率分别为15.15%和12.12%；图1b、d；表S1）。SmMYB36双等位突变体的花颜色较浅，而SmTCP15双等位突变体的花颜色较深（图1c，e）。高效液相色谱分析显示花青素含量发生了显著变化：SmMYB36突变体中飞燕苷含量急剧下降，花青素和天龙花苷含量减少30%，而SmTCP15突变体中飞燕苷含量增加1.66倍，花青素含量增加1.43倍（图1f；图S2a，b）。与之前的报道不同（Hsu et al. 2024）， SmMYB36突变体没有表现出白化花，可能是由于SmANS表达减少（图S2c）。相比之下，SmTCP15突变体的SmANS表达增加了2.3至2.6倍（图S2d）。这项研究为SmTCP15作为花青素生物合成负调控因子的作用提供了新的见解，并证明了启动编辑对表型性状微调的潜力。接下来，我们应用MediPlant-NEPE通过靶向丹参酮和酚酸生物合成的关键调控因子SmbZIP1来提高丹参根次生代谢物的产生。在残基V43处引入GTG>；TAA突变，产生终止密码子，突变频率为26.47%，双等位基因突变率为17.65%（图1g）。纯合子突变体的根颜色更深（图1h；图S3a），丹参酮含量增加（图1i；图S3b）。其中，隐丹参酮和丹参酮IIA含量分别增加1.86倍和1.40倍，丹参酮I含量变化不显著。SmbZIP1突变体的酚酸含量降低（图S3c，d），与之前的报道一致（Deng et al. 2020）。我们还针对毛状根中的SmRAS来促进迷迭香酸和丹酚酸B的产生（图S4a）。 G49A突变由G>；C转化产生，编辑效率为39.47%，双等位基因突变率为15.79%（图1j）。选择2个杂合株系（#HM04-02、#HM04-03）和1个纯合株系（#HM04-06）进行进一步分析（图1k）。HPLC证实，纯合突变体#HM04-06迷迭香酸增加1.98倍，丹酚酸B增加1.80倍（图11）。所有纯合突变体迷迭香酸增加1.93倍，丹酚酸B增加2.07倍（图S4b-e）。分子对接显示，G49A突变使酶袋大小最小化，并与底物和产物产生新的相互作用（图S4f）。结果表明，SmRASG49A提高了迷迭香酸和丹酚酸B的含量。本研究进一步利用MediPlant-NEPE系统对野生植物黄芪中的芦丁含量进行检测，以期促进其作为药用植物的快速驯化。UFGT3基因是芦丁生物合成的关键酶（He et al. 2022）（图S5a）。预测FdUFGT3蛋白的第120位（Q120H）和第170位（D170G）的氨基酸替换可以增强酶活性并增加芦丁的产量（图S5b）。使用MediPlant-NEPE，我们引入了这些突变，Q120H的编辑效率为12.00%，D170G的编辑效率为8.00%，Q120H和D170G的双突变体的编辑效率为12.00%（图1m）。选择纯合子突变株进行进一步分析（图1n）。HPLC分析显示，三个品系的槲皮素含量均有所下降（图10；图S5c），其中双突变体的芦丁含量增加了2.75倍，而单突变体的芦丁含量改善有限。这些结果表明，MediPlant-NEPE可以通过酶工程有效地提高次级代谢物的产生。分子对接显示，突变改变了FdUFGT3酶袋，单个突变导致槲皮素和udp -葡萄糖在单独的口袋中结合（图S5d）。相比之下，双突变扩大了酶袋以容纳底物和产物芦丁（图S5e）。这些发现突出了MediPlant-NEPE在改善药用植物代谢物生产方面的潜力。总的来说，MediPlant-NEPE为药用植物丹参（S. miltiorrhiza）和黄芪（F. dibotrys）的先导编辑提供了概念验证。我们的研究结果显示了其在其他药用物种中的广泛应用潜力，并强调了其在分子育种、增强次生代谢物和种质创新方面的前景，从而加速了改良品种的开发，缩短了具有经济和治疗价值的植物的育种周期。
{"title":"Enhancing Metabolic Engineering in Medicinal Plants Through Prime Editing","authors":"Haomiao Yu, Xiao Feng, Xiaohang Zheng, Xiao Wang, Wenxin Zheng, Zhizhou Zhang, Yuanyuan Jiang, Ruiwu Yang, Li Zhang, Zhaohui Zhong","doi":"10.1111/pbi.70532","DOIUrl":"https://doi.org/10.1111/pbi.70532","url":null,"abstract":"<p>Tanshinone and phenolic acid are key therapeutic compounds in the medicinal plant <i>Salvia miltiorrhiza</i>, while rutin is the major bioactive metabolite in the medicinal plant <i>Fagopyrum dibotrys</i>. However, their natural levels in cultivated varieties remain low, limiting their pharmacological potential. Enhancing metabolite accumulation through the modification of upstream transcription factors and key biosynthetic enzymes has shown promise (Deng et al. <span>2020</span>). Compounding the challenge, medicinal plants like <i>S. miltiorrhiza</i> are perennials with long life cycles, making conventional breeding inefficient. While CRISPR-Cas9 has facilitated gene knockout strategies in medicinal plants, precise base editing technologies remain underutilised (Das et al. <span>2024</span>). Prime editing (PE) has emerged as a powerful tool for introducing targeted nucleotide changes; thus, offering a promising route for molecular breeding (Anzalone et al. <span>2019</span>). While their application in medicinal plants remains largely unexplored.</p>\u0000<p>To address these limitations in genome engineering of medicinal plants, we develop the prime editor in medicinal plants. Our previous work demonstrated that N-terminus M-MLV RT fused to Cas9 nickase plus epegRNA prime editor (NEPE) is highly efficient in rice (Zhong et al. <span>2024</span>), indicating that the N-terminal fusion of M-MLV reverse transcriptase is particularly well-suited for plant applications. Building on this, and with optimization of the expression system, we further adapted the prime editor for medicinal plants, termed MediPlant-NEPE (Figure 1a; Figure S1).</p>\u0000<figure><picture>\u0000<source media=\"(min-width: 1650px)\" srcset=\"/cms/asset/227ed4e7-a506-4d78-a262-b21f293b3a66/pbi70532-fig-0001-m.jpg\"/><img alt=\"Details are in the caption following the image\" data-lg-src=\"/cms/asset/227ed4e7-a506-4d78-a262-b21f293b3a66/pbi70532-fig-0001-m.jpg\" loading=\"lazy\" src=\"/cms/asset/b14de0f4-3fe0-4aa2-9b5e-5fcefdd071f8/pbi70532-fig-0001-m.png\" title=\"Details are in the caption following the image\"/></picture><figcaption>\u0000<div><strong>FIGURE 1<span style=\"font-weight:normal\"></span></strong><div>Open in figure viewer<i aria-hidden=\"true\"></i><span>PowerPoint</span></div>\u0000</div>\u0000<div>Construction and application of MediPlant-NEPE for enhancing metabolic engineering in medicinal plants. (a) Schematic of MediPlant-NEPE. (b) Genotyping of <i>SmbZIP1</i>-edited hairy roots, red arrows indicate edits. (c) Phenotypes of <i>SmMYB36</i> homozygous mutants. (d) Genotyping of <i>SmTCP15</i>-edited lines, red arrows indicate edit sites. (e) Phenotypes of <i>SmTCP15</i> homozygous mutants. (f) HPLC quantification of anthocyanin in mutant flowers. (g) Genotyping of <i>SmbZIP1</i>-edited lines, red arrows indicate mutation sites. (h) Dry root powder of <i>SmbZIP1</i> mutants. (i) HPLC quantification of major tanshinones in roots. (j) Genotyping of <i>SmRAS</i>-edited hairy root lines, amino acid changes","PeriodicalId":221,"journal":{"name":"Plant Biotechnology Journal","volume":"1 1","pages":""},"PeriodicalIF":13.8,"publicationDate":"2026-01-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145903583","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Sihao Zhang, Bao Li, Xiaoting Li, Song Yu, Yebitao Yang, Peipei Liu, Tanglingdian He, Wei Wang, Mei Li, Liang Guo, Jinxing Tu
Brassica napus is the second most important oil crop worldwide. Number of primary branches (Branch number, BN) and silique number on the main inflorescence (SMI) are key yield‐related quantitative traits. Here, we cloned a major QTL, qDB.A09 , which simultaneously influences BN and SMI. The causal gene, SHOOT APICAL MERISTEM ENLARGER1 ( SAME1 ), encodes a mutator‐like transposase‐derived transcription factor and is functionally confirmed to positively regulate BN and SMI in B. napus . A rare C‐to‐A single nucleotide polymorphism (SNP1055), located 1.4 kb downstream of SAME1 , is associated with its elevated expression in the boundary region between the organising centre and central zone of the shoot apical meristem (SAM). Functional analysis indicates that high SAME1 expression represses the expression of BnaC06.ARR15 and expands the BnaA09.WUS expression domain, resulting in enlarged SAM size and increased BN and SMI, with BN increased by 3.40 ± 0.45 and SMI increased by 32.67 ± 4.03. In addition, the seed yield per plant is increased by 22.05%. We further demonstrate that qDB.A09 significantly increases BN and SMI in the elite cultivar ZS11, with BN increased by 3.10 ± 0.67 and SMI increased by 27.35 ± 9.12. This study provides a new genetic locus that can be utilised for the genetic improvement of yield‐related traits in B. napus .
{"title":"Rare Natural SNP Activates SHOOT APICAL MERISTEM ENLARGER1 to Increase Branch Number and Silique Number on the Main Inflorescence in Brassica napus","authors":"Sihao Zhang, Bao Li, Xiaoting Li, Song Yu, Yebitao Yang, Peipei Liu, Tanglingdian He, Wei Wang, Mei Li, Liang Guo, Jinxing Tu","doi":"10.1111/pbi.70500","DOIUrl":"https://doi.org/10.1111/pbi.70500","url":null,"abstract":"<jats:styled-content style=\"fixed-case\"> <jats:italic>Brassica napus</jats:italic> </jats:styled-content> is the second most important oil crop worldwide. Number of primary branches (Branch number, BN) and silique number on the main inflorescence (SMI) are key yield‐related quantitative traits. Here, we cloned a major QTL, <jats:italic>qDB.A09</jats:italic> , which simultaneously influences BN and SMI. The causal gene, <jats:italic>SHOOT APICAL MERISTEM ENLARGER1</jats:italic> ( <jats:italic>SAME1</jats:italic> ), encodes a mutator‐like transposase‐derived transcription factor and is functionally confirmed to positively regulate BN and SMI in <jats:styled-content style=\"fixed-case\"> <jats:italic>B. napus</jats:italic> </jats:styled-content> . A rare C‐to‐A single nucleotide polymorphism (SNP1055), located 1.4 kb downstream of <jats:italic>SAME1</jats:italic> , is associated with its elevated expression in the boundary region between the organising centre and central zone of the shoot apical meristem (SAM). Functional analysis indicates that high <jats:italic>SAME1</jats:italic> expression represses the expression of <jats:italic>BnaC06.ARR15</jats:italic> and expands the <jats:italic>BnaA09.WUS</jats:italic> expression domain, resulting in enlarged SAM size and increased BN and SMI, with BN increased by 3.40 ± 0.45 and SMI increased by 32.67 ± 4.03. In addition, the seed yield per plant is increased by 22.05%. We further demonstrate that <jats:italic>qDB.A09</jats:italic> significantly increases BN and SMI in the elite cultivar ZS11, with BN increased by 3.10 ± 0.67 and SMI increased by 27.35 ± 9.12. This study provides a new genetic locus that can be utilised for the genetic improvement of yield‐related traits in <jats:styled-content style=\"fixed-case\"> <jats:italic>B. napus</jats:italic> </jats:styled-content> .","PeriodicalId":221,"journal":{"name":"Plant Biotechnology Journal","volume":"11 1","pages":""},"PeriodicalIF":13.8,"publicationDate":"2026-01-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145903599","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
The Common Symbiosis Signalling Pathway (CSSP) underpins interactions between plants and microbes, yet its potential for crop improvement remains underexplored. Here, we investigated the gain‐of‐function mutant SPD1 ( MtDMI1S760N ), which constitutively activates the symbiotic signalling pathway in Medicago truncatula , by expressing it in tomato ( Solanum lycopersicum cv. Micro‐Tom). Heterologous expression of SPD1 constitutively activated ethylene biosynthesis, leading to broad‐spectrum resistance against fungal, bacterial, and vascular pathogens. Beyond immunity, SPD1 reprogrammed tomato development, accelerating seed germination, flowering, and fruit ripening, while reducing arbuscular mycorrhizal colonisation and primary root growth. Transcriptome analysis revealed constitutive activation of ethylene biosynthesis and immune marker genes, consistent with increased ethylene emission and amplified ROS and MAPK response to both pathogenic and symbiotic elicitors. Ethylene inhibitor AVG reversed both immune activation and root defects, confirming a central role of ethylene signalling in SPD1‐mediated reprogramming. Our findings show that an autoactivate legume symbiotic component can reprogramme defence and development traits in a non‐legume via ethylene signalling, highlighting SPD1 as a promising tool for breeding early‐maturing and disease‐resistance crops.
{"title":"Autoactive MtDMI1 Reprogrammes Immunity and Development in Tomato via Ethylene Signalling","authors":"Haiyue Liu, Ji Xu, Fang Xie","doi":"10.1111/pbi.70533","DOIUrl":"https://doi.org/10.1111/pbi.70533","url":null,"abstract":"The Common Symbiosis Signalling Pathway (CSSP) underpins interactions between plants and microbes, yet its potential for crop improvement remains underexplored. Here, we investigated the gain‐of‐function mutant <jats:italic>SPD1</jats:italic> ( <jats:italic>MtDMI1</jats:italic> <jats:sup> <jats:italic>S760N</jats:italic> </jats:sup> ), which constitutively activates the symbiotic signalling pathway in <jats:styled-content style=\"fixed-case\"> <jats:italic>Medicago truncatula</jats:italic> </jats:styled-content> , by expressing it in tomato ( <jats:styled-content style=\"fixed-case\"> <jats:italic>Solanum lycopersicum</jats:italic> </jats:styled-content> cv. Micro‐Tom). Heterologous expression of <jats:italic>SPD1</jats:italic> constitutively activated ethylene biosynthesis, leading to broad‐spectrum resistance against fungal, bacterial, and vascular pathogens. Beyond immunity, SPD1 reprogrammed tomato development, accelerating seed germination, flowering, and fruit ripening, while reducing arbuscular mycorrhizal colonisation and primary root growth. Transcriptome analysis revealed <jats:italic>constitutive</jats:italic> activation of ethylene biosynthesis and immune marker genes, consistent with increased ethylene emission and amplified ROS and MAPK response to both pathogenic and symbiotic elicitors. Ethylene inhibitor AVG reversed both immune activation and root defects, confirming a central role of ethylene signalling in SPD1‐mediated reprogramming. Our findings show that an autoactivate legume symbiotic component can reprogramme defence and development traits in a non‐legume via ethylene signalling, highlighting SPD1 as a promising tool for breeding early‐maturing and disease‐resistance crops.","PeriodicalId":221,"journal":{"name":"Plant Biotechnology Journal","volume":"14 1","pages":""},"PeriodicalIF":13.8,"publicationDate":"2026-01-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145903588","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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