Huan Chen, Tiange Zhou, Xianxin Wu, Vikranth Kumar, Xingguo Lan, Yuan Hu Xuan
SummaryLight signalling regulates plant growth and stress resistance, whereas its mechanism in controlling saline‐alkaline tolerance (SAT) remains largely unknown. This study identified that light signalling, primarily mediated by Phytochrome B (PhyB), inhibited ammonium transporter 1 (AMT1) to negatively regulate SAT. Our previous findings have shown that PhyB can impede the transcription factors indeterminate domain 10 (IDD10) and brassinazole resistant 1 (BZR1) to reduce NH4+ uptake, thereby modulating SAT and sheath blight (ShB) resistance in rice. However, inhibition of IDD10 and BZR1 in the phyB background did not fully suppress NH4+ uptake, suggesting that other signalling pathways regulated AMT1 downstream of PhyB. Further analysis revealed that PhyB interacted with Calcineurin B‐like protein‐interacting protein kinase 31 (CIPK31), which positively regulated AMT1 expression. CIPK31 also interacted with Teosinte Branched1/Cycloidea/PCF19 (TCP19), a key regulator of nitrogen use efficiency (NUE). However, PhyB neither degraded CIPK31 nor directly interacted with TCP19. Instead, PhyB inhibited the CIPK31‐TCP19 interaction, releasing TCP19, which repressed AMT1;2 directly and AMT1;1 and AMT1;3 indirectly, thereby inhibiting NH4+ uptake and SAT while reducing ShB resistance. Additionally, Phytochrome Interacting Factor‐Like 15 (PIL15) interacted with TCP19. Different from TCP19, PIL15 directly activated AMT1;2 to promote SAT, suggesting a balancing mechanism for NH4+ uptake downstream of PhyB. Furthermore, PIL15 interacted with IDD10 and BZR1 to form a transcriptional complex that collaboratively activated AMT1;2 expression. Overall, this study provides novel insights into how PhyB signalling regulates NH4+ uptake and coordinates SAT and ShB resistance in rice.
{"title":"Phytochrome B‐mediated light signalling enhances rice resistance to saline‐alkaline and sheath blight by regulating multiple downstream transcription factors","authors":"Huan Chen, Tiange Zhou, Xianxin Wu, Vikranth Kumar, Xingguo Lan, Yuan Hu Xuan","doi":"10.1111/pbi.14599","DOIUrl":"https://doi.org/10.1111/pbi.14599","url":null,"abstract":"SummaryLight signalling regulates plant growth and stress resistance, whereas its mechanism in controlling saline‐alkaline tolerance (SAT) remains largely unknown. This study identified that light signalling, primarily mediated by Phytochrome B (PhyB), inhibited <jats:italic>ammonium transporter 1</jats:italic> (<jats:italic>AMT1</jats:italic>) to negatively regulate SAT. Our previous findings have shown that PhyB can impede the transcription factors indeterminate domain 10 (IDD10) and brassinazole resistant 1 (BZR1) to reduce NH<jats:sub>4</jats:sub><jats:sup>+</jats:sup> uptake, thereby modulating SAT and sheath blight (ShB) resistance in rice. However, inhibition of IDD10 and BZR1 in the <jats:italic>phyB</jats:italic> background did not fully suppress NH<jats:sub>4</jats:sub><jats:sup>+</jats:sup> uptake, suggesting that other signalling pathways regulated <jats:italic>AMT1</jats:italic> downstream of PhyB. Further analysis revealed that PhyB interacted with Calcineurin B‐like protein‐interacting protein kinase 31 (CIPK31), which positively regulated <jats:italic>AMT1</jats:italic> expression. CIPK31 also interacted with Teosinte Branched1/Cycloidea/PCF19 (TCP19), a key regulator of nitrogen use efficiency (NUE). However, PhyB neither degraded CIPK31 nor directly interacted with TCP19. Instead, PhyB inhibited the CIPK31‐TCP19 interaction, releasing TCP19, which repressed <jats:italic>AMT1;2</jats:italic> directly and <jats:italic>AMT1;1</jats:italic> and <jats:italic>AMT1;3</jats:italic> indirectly, thereby inhibiting NH<jats:sub>4</jats:sub><jats:sup>+</jats:sup> uptake and SAT while reducing ShB resistance. Additionally, Phytochrome Interacting Factor‐Like 15 (PIL15) interacted with TCP19. Different from TCP19, PIL15 directly activated <jats:italic>AMT1;2</jats:italic> to promote SAT, suggesting a balancing mechanism for NH<jats:sub>4</jats:sub><jats:sup>+</jats:sup> uptake downstream of PhyB. Furthermore, PIL15 interacted with IDD10 and BZR1 to form a transcriptional complex that collaboratively activated <jats:italic>AMT1;2</jats:italic> expression. Overall, this study provides novel insights into how PhyB signalling regulates NH<jats:sub>4</jats:sub><jats:sup>+</jats:sup> uptake and coordinates SAT and ShB resistance in rice.","PeriodicalId":221,"journal":{"name":"Plant Biotechnology Journal","volume":"128 1","pages":""},"PeriodicalIF":13.8,"publicationDate":"2025-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143072416","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}
Cytosine methylation (mCG) is an important heritable epigenetic modification, yet its functions remain to be fully defined in important crops. This study investigates mCG in soybean following the loss-of-function mutation of two GmMET1 genes. We generated knockout mutants of GmMET1s by CRISPR-Cas9 and conducted comprehensive methylome and transcriptome analyses. Our findings unravel the functional redundancy of the two GmMET1s, with GmMET1b being more critically involved in maintaining mCG levels, and complete knockout of both copies is lethal. We establish that genome-wide mCG levels scale with aggregated expression of GmMET1s. We identify a set of mCG-regulated genes whose expression levels were quantitatively modulated by upstream, body, or downstream mCG. Moreover, we find genes that were negatively regulated by upstream or body mCG are enriched in specific biological processes such as that of jasmonic acid metabolism. Notably, >80% of the differentially methylated genes (DMGs) in the mutants also exist as DMGs in natural soybean populations. Phenotypically, mutants that are heterozygous for GmMET1a and homozygous for GmMET1b knockouts (GmMET1a+/-GmMET1b-/-) exhibited early flowering, which was inherited by their selfed progeny (GmMET1a+/+GmMET1b-/-) with otherwise normal growth and development. Moreover, mutation of either GmMET1s, with slight reduction of mCG levels and similar phenotypes compared to the wild type under normal conditions, showed enhanced tolerance to cold and drought stresses. Together, our results underscore highly orchestrated regulatory effects of mCG on gene expression in soybean, which dictates growth, development and stress responses, implicating its utility in the improvement of soybean for better adaptability and higher yield.
{"title":"A modulatory role of CG methylation on gene expression in soybean implicates its potential utility in breeding.","authors":"Ying Wang, Hongwei Xun, Jiameng Lv, Wanting Ju, Yuhui Jiang, Meng Wang, Ruihong Guo, Mengru Zhang, Xiaoyang Ding, Bao Liu, Chunming Xu","doi":"10.1111/pbi.14606","DOIUrl":"https://doi.org/10.1111/pbi.14606","url":null,"abstract":"<p><p>Cytosine methylation (mCG) is an important heritable epigenetic modification, yet its functions remain to be fully defined in important crops. This study investigates mCG in soybean following the loss-of-function mutation of two GmMET1 genes. We generated knockout mutants of GmMET1s by CRISPR-Cas9 and conducted comprehensive methylome and transcriptome analyses. Our findings unravel the functional redundancy of the two GmMET1s, with GmMET1b being more critically involved in maintaining mCG levels, and complete knockout of both copies is lethal. We establish that genome-wide mCG levels scale with aggregated expression of GmMET1s. We identify a set of mCG-regulated genes whose expression levels were quantitatively modulated by upstream, body, or downstream mCG. Moreover, we find genes that were negatively regulated by upstream or body mCG are enriched in specific biological processes such as that of jasmonic acid metabolism. Notably, >80% of the differentially methylated genes (DMGs) in the mutants also exist as DMGs in natural soybean populations. Phenotypically, mutants that are heterozygous for GmMET1a and homozygous for GmMET1b knockouts (GmMET1a<sup>+/-</sup>GmMET1b<sup>-/-</sup>) exhibited early flowering, which was inherited by their selfed progeny (GmMET1a<sup>+/+</sup>GmMET1b<sup>-/-</sup>) with otherwise normal growth and development. Moreover, mutation of either GmMET1s, with slight reduction of mCG levels and similar phenotypes compared to the wild type under normal conditions, showed enhanced tolerance to cold and drought stresses. Together, our results underscore highly orchestrated regulatory effects of mCG on gene expression in soybean, which dictates growth, development and stress responses, implicating its utility in the improvement of soybean for better adaptability and higher yield.</p>","PeriodicalId":221,"journal":{"name":"Plant Biotechnology Journal","volume":" ","pages":""},"PeriodicalIF":10.1,"publicationDate":"2025-01-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143062170","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}
Hui Du, Zefeng Zhai, Jin Pu, Jun Liang, Rongyan Wang, Zhong Zhang, Pei Wang, Yanhui Zhu, Lian Huang, Dawei Li, Kaiyuan Chen, Guangtao Zhu, Chunzhi Zhang
Anthocyanin biosynthesis and accumulation determines the colour of tuber flesh in potato (Solanum tuberosum) and influences nutritional quality. However, the regulatory mechanism behind anthocyanin biosynthesis in potato tuber flesh remains unclear. In this study, we identified the Pigmented tuber flesh (Pf) locus through a genome-wide association study using 135 diploid potato landraces. Genome editing of two tandem R2R3 MYB transcription factor genes, StMYB200 and StMYB210, within the Pf locus demonstrated that both genes are involved in anthocyanin biosynthesis in tuber flesh. Molecular and biochemical assays revealed that StMYB200 promotes StMYB210 transcription by directly binding to a 1.7-kb insertion present in the StMYB210 promoter, while StMYB210 also regulates its own expression. Furthermore, StMYB200 and StMYB210 both activated the expression of the basic helix–loop–helix transcription factor gene StbHLH1 and interacted with StbHLH1 to regulate anthocyanin biosynthesis. An analysis of the StMYB210 promoter in different diploid potato accessions showed that the 1.7-kb insertion is associated with flesh colour in potato. These findings reveal the genetic and molecular mechanism by which the Pf locus regulates anthocyanin accumulation in tuber flesh and provide an important reference for breeding new potato varieties with colourful flesh.
{"title":"Two tandem R2R3 MYB transcription factor genes cooperatively regulate anthocyanin accumulation in potato tuber flesh","authors":"Hui Du, Zefeng Zhai, Jin Pu, Jun Liang, Rongyan Wang, Zhong Zhang, Pei Wang, Yanhui Zhu, Lian Huang, Dawei Li, Kaiyuan Chen, Guangtao Zhu, Chunzhi Zhang","doi":"10.1111/pbi.14602","DOIUrl":"https://doi.org/10.1111/pbi.14602","url":null,"abstract":"Anthocyanin biosynthesis and accumulation determines the colour of tuber flesh in potato (<i>Solanum tuberosum</i>) and influences nutritional quality. However, the regulatory mechanism behind anthocyanin biosynthesis in potato tuber flesh remains unclear. In this study, we identified the <i>Pigmented tuber flesh</i> (<i>Pf</i>) locus through a genome-wide association study using 135 diploid potato landraces. Genome editing of two tandem R2R3 MYB transcription factor genes, <i>StMYB200</i> and <i>StMYB210</i>, within the <i>Pf</i> locus demonstrated that both genes are involved in anthocyanin biosynthesis in tuber flesh. Molecular and biochemical assays revealed that StMYB200 promotes <i>StMYB210</i> transcription by directly binding to a 1.7-kb insertion present in the <i>StMYB210</i> promoter, while StMYB210 also regulates its own expression. Furthermore, StMYB200 and StMYB210 both activated the expression of the basic helix–loop–helix transcription factor gene <i>StbHLH1</i> and interacted with StbHLH1 to regulate anthocyanin biosynthesis. An analysis of the <i>StMYB210</i> promoter in different diploid potato accessions showed that the 1.7-kb insertion is associated with flesh colour in potato. These findings reveal the genetic and molecular mechanism by which the <i>Pf</i> locus regulates anthocyanin accumulation in tuber flesh and provide an important reference for breeding new potato varieties with colourful flesh.","PeriodicalId":221,"journal":{"name":"Plant Biotechnology Journal","volume":"40 1","pages":""},"PeriodicalIF":13.8,"publicationDate":"2025-01-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143057028","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}
Zhongze Li, Minjae Kim, Jose Roberto da Silva Nascimento, Bertrand Legeret, Gabriel Lemes Jorge, Marie Bertrand, Fred Beisson, Jay J. Thelen, Yonghua Li-Beisson
The first step in chloroplast de novo fatty acid synthesis is catalysed by acetyl-CoA carboxylase (ACCase). As the rate-limiting step for this pathway, ACCase is subject to both positive and negative regulation. In this study, we identify a Chlamydomonas homologue of the plant carboxyltransferase interactor 1 (CrCTI1) and show that this protein interacts with the Chlamydomonas α-carboxyltransferase (Crα-CT) subunit of the ACCase by yeast two-hybrid protein–protein interaction assay. Three independent CRISPR-Cas9 mediated knockout mutants for CrCTI1 each produced an ‘enhanced oil’ phenotype, accumulating 25% more total fatty acids and storing up to fivefold more triacylglycerols (TAGs) in lipid droplets. The TAG phenotype of the crcti1 mutants was not influenced by light but was affected by trophic growth conditions. By growing cells under heterotrophic conditions, we observed a crucial function of CrCTI1 in balancing lipid accumulation and cell growth. Mutating a previously mapped in vivo phosphorylation site (CrCTI1 Ser108 to either Ala or to Asp), did not affect the interaction with Crα-CT. However, mutating all six predicted phosphorylation sites within Crα-CT to create a phosphomimetic mutant reduced this pairwise interaction significantly. Comparative proteomic analyses of the crcti1 mutants and WT suggested a role for CrCTI1 in regulating carbon flux by coordinating carbon metabolism, antioxidant and fatty acid β-oxidation pathways, to enable cells to adapt to carbon availability. Taken together, this study identifies CrCTI1 as a negative regulator of fatty acid synthesis in algae and provides a new molecular brick for the genetic engineering of microalgae for biotechnology purposes.
{"title":"Knocking out the carboxyltransferase interactor 1 (CTI1) in Chlamydomonas boosted oil content by fivefold without affecting cell growth","authors":"Zhongze Li, Minjae Kim, Jose Roberto da Silva Nascimento, Bertrand Legeret, Gabriel Lemes Jorge, Marie Bertrand, Fred Beisson, Jay J. Thelen, Yonghua Li-Beisson","doi":"10.1111/pbi.14581","DOIUrl":"https://doi.org/10.1111/pbi.14581","url":null,"abstract":"The first step in chloroplast <i>de novo</i> fatty acid synthesis is catalysed by acetyl-CoA carboxylase (ACCase). As the rate-limiting step for this pathway, ACCase is subject to both positive and negative regulation. In this study, we identify a Chlamydomonas homologue of the plant carboxyltransferase interactor 1 (CrCTI1) and show that this protein interacts with the Chlamydomonas α-carboxyltransferase (Crα-CT) subunit of the ACCase by yeast two-hybrid protein–protein interaction assay. Three independent CRISPR-Cas9 mediated knockout mutants for CrCTI1 each produced an ‘enhanced oil’ phenotype, accumulating 25% more total fatty acids and storing up to fivefold more triacylglycerols (TAGs) in lipid droplets. The TAG phenotype of the <i>crcti1</i> mutants was not influenced by light but was affected by trophic growth conditions. By growing cells under heterotrophic conditions, we observed a crucial function of CrCTI1 in balancing lipid accumulation and cell growth. Mutating a previously mapped <i>in vivo</i> phosphorylation site (CrCTI1 Ser108 to either Ala or to Asp), did not affect the interaction with Crα-CT. However, mutating all six predicted phosphorylation sites within Crα-CT to create a phosphomimetic mutant reduced this pairwise interaction significantly. Comparative proteomic analyses of the <i>crcti1</i> mutants and WT suggested a role for CrCTI1 in regulating carbon flux by coordinating carbon metabolism, antioxidant and fatty acid β-oxidation pathways, to enable cells to adapt to carbon availability. Taken together, this study identifies CrCTI1 as a negative regulator of fatty acid synthesis in algae and provides a new molecular brick for the genetic engineering of microalgae for biotechnology purposes.","PeriodicalId":221,"journal":{"name":"Plant Biotechnology Journal","volume":"207 1","pages":""},"PeriodicalIF":13.8,"publicationDate":"2025-01-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143057033","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}
Jingfang Dong, Shaohong Zhang, Haifei Hu, Jian Wang, Risheng Li, Jing Wu, Jiansong Chen, Lian Zhou, Yamei Ma, Wenhui Li, Shuai Nie, Shaokui Wang, Guiquan Zhang, Bin Liu, Junliang Zhao, Tifeng Yang
Improving cold tolerance at the flowering stage (CTF) in rice is crucial for minimising yield loss, making the identification and application of cold-tolerant genes and QTLs imperative for effective molecular breeding. The long lead time, dependence on cold treatment conditions, and the inherent complexity of the trait make studying the genetic basis of CTF in rice challenging. To date, the fine-mapping or cloning of QTLs specific to CTF has not yet been achieved. In this study, single segment substitution lines (SSSLs) were constructed using HJX74 as the recipient and IR58025B, known for good CTF, as the donor. This approach led to the identification of two cold tolerance QTLs, qCTF3 and qCTF6, in rice. qCTF6 has promising breeding potential. Further, we identified the causal gene CTF1 underlying qCTF6 through map-based cloning. CTF1 which encodes a conserved putative protein, has two SNPs within its coding sequence that influence CTF in rice. Additionally, genetic variations in the promoter of CTF1 also contributes to CTF. Thirteen variant sites of CTF1 in the four cold tolerance SSSLs are consistent with the IR58025B. Moreover, we analysed 307 accessions to characterise haplotypes based on the 13 variation sites, identifying five distinct haplotypes. The selection and evolutionary analysis indicate that the cold-tolerant haplotype of CTF1 is a newly generated mutation that has undergone selection in japonica during domestication. This study not only provides a novel favourable gene for molecular breeding of CTF but also highlights the potential of CTF1 in advancing rice breeding.
{"title":"Natural variation in CTF1 conferring cold tolerance at the flowering stage in rice","authors":"Jingfang Dong, Shaohong Zhang, Haifei Hu, Jian Wang, Risheng Li, Jing Wu, Jiansong Chen, Lian Zhou, Yamei Ma, Wenhui Li, Shuai Nie, Shaokui Wang, Guiquan Zhang, Bin Liu, Junliang Zhao, Tifeng Yang","doi":"10.1111/pbi.14600","DOIUrl":"https://doi.org/10.1111/pbi.14600","url":null,"abstract":"Improving cold tolerance at the flowering stage (CTF) in rice is crucial for minimising yield loss, making the identification and application of cold-tolerant genes and QTLs imperative for effective molecular breeding. The long lead time, dependence on cold treatment conditions, and the inherent complexity of the trait make studying the genetic basis of CTF in rice challenging. To date, the fine-mapping or cloning of QTLs specific to CTF has not yet been achieved. In this study, single segment substitution lines (SSSLs) were constructed using HJX74 as the recipient and IR58025B, known for good CTF, as the donor. This approach led to the identification of two cold tolerance QTLs, <i>qCTF3</i> and <i>qCTF6</i>, in rice. <i>qCTF6</i> has promising breeding potential. Further, we identified the causal gene <i>CTF1</i> underlying <i>qCTF6</i> through map-based cloning<i>. CTF1</i> which encodes a conserved putative protein, has two SNPs within its coding sequence that influence CTF in rice. Additionally, genetic variations in the promoter of <i>CTF1</i> also contributes to CTF. Thirteen variant sites of <i>CTF1</i> in the four cold tolerance SSSLs are consistent with the IR58025B. Moreover, we analysed 307 accessions to characterise haplotypes based on the 13 variation sites, identifying five distinct haplotypes. The selection and evolutionary analysis indicate that the cold-tolerant haplotype of <i>CTF1</i> is a newly generated mutation that has undergone selection in <i>japonica</i> during domestication. This study not only provides a novel favourable gene for molecular breeding of CTF but also highlights the potential of <i>CTF1</i> in advancing rice breeding.","PeriodicalId":221,"journal":{"name":"Plant Biotechnology Journal","volume":"45 1","pages":""},"PeriodicalIF":13.8,"publicationDate":"2025-01-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143056341","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}
Jarst van Belle, Jan G. Schaart, Annemarie C. Dechesne, Danli Fei, Abraham Ontiveros Cisneros, Michele Serafini, Richard G.F. Visser, Eibertus N. van Loo
Mutants with simultaneous germline mutations were obtained in all three F5H genes and all three FAD2 genes (one to eleven mutated alleles) in order to improve the feed value of the seed meal and the fatty acid composition of the seed oil. In mutants with multiple mutated F5H alleles, sinapine in seed meal was reduced by up to 100%, accompanied by a sharp reduction in the S-monolignol content of lignin without causing lodging or stem break. A lower S-lignin monomer content in stems can contribute to improved stem degradability allowing new uses of stems. Mutants in all six FAD2 alleles showed an expected increase in MUFA from 8.7% to 74% and a reduction in PUFA from 53% to 13% in the fatty acids in seed oil. Remarkably, some full FAD2 mutants showed normal growth and seed production and not the dwarfing phenotype reported in previous studies. The relation between germline mutation allele dosage and phenotype was influenced by the still ongoing activity of the CRISPR/Cas9 system, leading to new somatic mutations in the leaves of flowering plants. The correlations between the total mutation frequency (germline plus new somatic mutations) for F5H with sinapine content, and FAD2 with fatty acid composition were higher than the correlations between germline mutation count and phenotypes. This shows the importance of quantifying both the germline mutations and somatic mutations when studying CRISPR/Cas9 effects in situations where the CRISPR/Cas9 system is not yet segregated out.
{"title":"Direct and indirect effects of multiplex genome editing of F5H and FAD2 in oil crop camelina","authors":"Jarst van Belle, Jan G. Schaart, Annemarie C. Dechesne, Danli Fei, Abraham Ontiveros Cisneros, Michele Serafini, Richard G.F. Visser, Eibertus N. van Loo","doi":"10.1111/pbi.14593","DOIUrl":"https://doi.org/10.1111/pbi.14593","url":null,"abstract":"Mutants with simultaneous germline mutations were obtained in all three <i>F5H</i> genes and all three <i>FAD2</i> genes (one to eleven mutated alleles) in order to improve the feed value of the seed meal and the fatty acid composition of the seed oil. In mutants with multiple mutated <i>F5H</i> alleles, sinapine in seed meal was reduced by up to 100%, accompanied by a sharp reduction in the S-monolignol content of lignin without causing lodging or stem break. A lower S-lignin monomer content in stems can contribute to improved stem degradability allowing new uses of stems. Mutants in all six <i>FAD2</i> alleles showed an expected increase in MUFA from 8.7% to 74% and a reduction in PUFA from 53% to 13% in the fatty acids in seed oil. Remarkably, some full <i>FAD2</i> mutants showed normal growth and seed production and not the dwarfing phenotype reported in previous studies. The relation between germline mutation allele dosage and phenotype was influenced by the still ongoing activity of the CRISPR/Cas9 system, leading to new somatic mutations in the leaves of flowering plants. The correlations between the total mutation frequency (germline plus new somatic mutations) for <i>F5H</i> with sinapine content, and <i>FAD2</i> with fatty acid composition were higher than the correlations between germline mutation count and phenotypes. This shows the importance of quantifying both the germline mutations and somatic mutations when studying CRISPR/Cas9 effects in situations where the CRISPR/Cas9 system is not yet segregated out.","PeriodicalId":221,"journal":{"name":"Plant Biotechnology Journal","volume":"38 1","pages":""},"PeriodicalIF":13.8,"publicationDate":"2025-01-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143044199","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>Wood, one of the most abundant renewable natural resources globally, plays a crucial role in the timber, papermaking and bioenergy industries (Chutturi <i>et al</i>., <span>2023</span>). Wood (i.e. secondary xylem) is derived from vascular cambium, which is pivotal in determining the wood biomass in woody plants (Tang <i>et al</i>., <span>2022</span>). Reactive oxygen species (ROS) act as signalling molecules that regulate plant development, growth and responses to abiotic and biotic stresses (Wang <i>et al</i>., <span>2024</span>). Numerous studies underscore the significance of ROS in maintaining the root and shoot stem cell niches (Wang <i>et al</i>., <span>2024</span>). A recent study has indicated that LATERAL ORGAN BOUNDARIES DOMAIN 11 (LBD11) governs several ROS metabolic genes to manage the specific distribution of ROS within the cambium, thus affecting cambial cell proliferation in <i>Arabidopsis</i> root and shoot (Dang <i>et al</i>., <span>2023</span>). However, there remains a lack of clarity on the biological functions of ROS accumulation in tree vascular cambium activity. Additionally, the localized accumulation of ROS is required for lignin biosynthesis (Wang <i>et al</i>., <span>2024</span>). Therefore, ROS homeostasis enables woody plants to fine-tune the activity of cambium, increase wood yield and improve their quality.</p>