Pathogen-associated molecular patterns(PAMPs)-triggered immunity(PTI)is an important component of plant innate immunity.In a previous study,we showed that the PAMP flg22 from Xanthomonas citri ssp.citri(Xflg22),the ca...Pathogen-associated molecular patterns(PAMPs)-triggered immunity(PTI)is an important component of plant innate immunity.In a previous study,we showed that the PAMP flg22 from Xanthomonas citri ssp.citri(Xflg22),the causal agent of citrus canker,induced PTI in citrus,which correlated with the observed levels of canker resistance.Here,we identified and sequenced two bacterial flagellin/flg22 receptors(FLS2-1 and FLS2-2)from‘Duncan’grapefruit(Citrus paradisi,CpFLS2-1 and CpFLS2-2)and‘Sun Chu Sha’mandarin(C.reticulata,CrFLS2-1 and CrFLS2-2).We were able to isolate only one FLS2 from‘Nagami’kumquat(Fortunella margarita,FmFLS2-1)and gene flanking sequences suggest a rearrangement event that resulted in the deletion of FLS2-2 from the genome.Phylogenetic analysis,gene structure and presence of critical amino acid domains all indicate we identified the true FLS2 genes in citrus.FLS2-2 was more transcriptionally responsive to Xflg22 than FLS2-1,with induced expression levels higher in canker-resistant citrus than in susceptible ones.Interestingly,‘Nagami’kumquat showed the highest FLS2-1 steady-state expression levels,although it was not induced by Xflg22.We selected FmFLS2-1,CrFLS2-2 and CpFLS2-2 to further evaluate their capacity to enhance bacterial resistance using Agrobacterium-mediated transient expression assays.Both FmFLS2-1 and CrFLS2-2,the two proteins from canker-resistant species,conferred stronger Xflg22 responses and reduced canker symptoms in leaves of the susceptible grapefruit genotype.These two citrus genes will be useful resources to enhance PTI and achieve resistance against canker and possibly other bacterial pathogens in susceptible citrus types.展开更多
The receptor-like kinase FLAGELLIN-SENSITIVE 2(FLS2)functions as a bacterialflagellin receptor local-ized on the cell membrane of plants.In Arabidopsis,the co-receptor BRI1-ASSOCIATED RECEPTOR KI-NASE 1(BAK1)cooperate...The receptor-like kinase FLAGELLIN-SENSITIVE 2(FLS2)functions as a bacterialflagellin receptor local-ized on the cell membrane of plants.In Arabidopsis,the co-receptor BRI1-ASSOCIATED RECEPTOR KI-NASE 1(BAK1)cooperates with FLS2 to detect theflagellin epitopeflg22,resulting in formation of a signaling complex that triggers plant defense responses.However,the co-receptor responsible for recog-nizing and signaling theflg22 epitope in rice remains to be determined,and the precise structural mecha-nism underlying FLS2-mediated signal activation and transduction has not been claried.This study pre-sents the structural characterization of a kinase-dead mutant of the intracellular kinase domain of OsFLS2(OsFLS2-KDD1013A)in complex with ATP or ADP,resolved at resolutions of 1.98 A˚and 2.09 A˚,respectively.Structural analysis revealed that OsFLS2 can adopt an active conformation in the absence of phosphorylation,although it exhibits only weak basal catalytic activity for autophosphorylation.Subse-quent investigations demonstrated that OsSERK2 effectively phosphorylates OsFLS2,which reciprocally phosphorylates OsSERK2,leading to complete activation of OsSERK2 and rapid phosphorylation of the downstream substrate receptor-like cytoplasmic kinases OsRLCK176 and OsRLCK185.Through mass spectrometry experiments,we successfully identied critical autophosphorylation sites on OsSERK2,as well as sites transphosphorylated by OsFLS2.Furthermore,we demonstrated the interaction between OsSERK2 and OsFLS2,which is enhanced in the presence offlg22.Genetic evidence suggests that OsRLCK176 and OsRLCK185 may function downstream of the OsFLS2-mediated signaling pathway.Our study reveals the molecular mechanism by which OsFLS2 mediates signal transduction pathways in rice and provides a valuable example for understanding RLK-mediated signaling pathways in plants.展开更多
Infections with Salmonella enterica belong to the most prominent causes of food poisoning and infected fruits and vegetables represent important vectors for salmonellosis. Recent evidence indicates that plants recogni...Infections with Salmonella enterica belong to the most prominent causes of food poisoning and infected fruits and vegetables represent important vectors for salmonellosis. Recent evidence indicates that plants recognize S. enterica and raise defense responses. Nonetheless, the molecular mechanisms controlling the interaction of S. enterica with plants are still largely unclear. Here, we show that flagellin from S. enterica represents a prominent pathogenassociated molecular pattern （PAMP） in Arabidopsis thaliana, which induces PAMP-triggered immunity （PTI） via the recognition of the fig22 domain by the receptor kinase FLS2. The Arabidopsis fls2 mutant shows reduced though not abolished PTI activation, indicating that plants rely also on recognition of other S. enterica PAMPs. Interestingly, the S. enterica type III secretion system （T3SS） mutant prgH- induced stronger defense gene expression than wild-type bacteria in Arabidopsis, suggesting that T3SS effectors are involved in defense suppression. Furthermore, we observe that S. enterica strains show variation in the fig22 epitope, which results in proteins with reduced PTI-inducing activity. Altogether, these results show that S. enterica activates PTI in Arabidopsis and suggest that, in order to accomplish plant colonization, S. enterica evolved strategies to avoid or suppress PTI.展开更多
Genetic pest control strategies based on precise sex separation and only releasing sterile males can be accomplished by site-specific genome editing.In the current study,we showed that the mutation of single-allele Px...Genetic pest control strategies based on precise sex separation and only releasing sterile males can be accomplished by site-specific genome editing.In the current study,we showed that the mutation of single-allele Pxfl(2)d can significantly impair the normal mating behavior and testis development in male adults of the notorious cruciferous insect pest Plutella xylostella,in addition to its known functions in the ovarian development in female adults and egg hatching.Subsequent CRISPR/Cas9-based knock-in experiments revealed that site-specific integration of an exogenous green fluorescent protein(GFP)gene into autosomal Pxfl(2)d for labelling mutants could be achieved.However,this gene is not a suitable target for GFP insertion to establish a genetically stable knock-in strain because of the severe decline in reproductive capacity.We further screened for the W-chromosome-linked and Z-chromosome-linked regions to test the knock-in efficiency mediated by CRISPR/Cas9.The results verified that both types of chromosomes can be targeted for the site-specific insertion of exogenous sequences.We ultimately obtained a homozygous knock-in strain with the integration of both Cas9 and cyan fluorescent protein(CFP)expression cassettes on a Z-linked region in P.xylostella,which can also be used for early sex detection.By injecting the sgRNA targeting Pxfl(2)d alone into the eggs laid by female adults of the Z-Cas9-CFP strain,the gene editing efficiency reached 29.73%,confirming the success of expressing a functional Cas9 gene.Taken together,we demonstrated the feasibility of the knock-in of an exogenous gene to different genomic regions in P.xylostella,while the establishment of a heritable strain required the positioning of appropriate sites.This study provides an important working basis and technical support for further developing genetic strategies for insect pest control.展开更多
Breast cancer remains a significant global health challenge, necessitating effective early detection and prognosis to enhance patient outcomes. Current diagnostic methods, including mammography and MRI, suffer from li...Breast cancer remains a significant global health challenge, necessitating effective early detection and prognosis to enhance patient outcomes. Current diagnostic methods, including mammography and MRI, suffer from limitations such as uncertainty and imprecise data, leading to late-stage diagnoses. To address this, various expert systems have been developed, but many rely on type-1 fuzzy logic and lack mobile-based applications for data collection and feedback to healthcare practitioners. This research investigates the development of an Enhanced Mobile-based Fuzzy Expert system (EMFES) for breast cancer pre-growth prognosis. The study explores the use of type-2 fuzzy logic to enhance accuracy and model uncertainty effectively. Additionally, it evaluates the advantages of employing the python programming language over java for implementation and considers specific risk factors for data collection. The research aims to dynamically generate fuzzy rules, adapting to evolving breast cancer research and patient data. Key research questions focus on the comparative effectiveness of type-2 fuzzy logic, the handling of uncertainty and imprecise data, the integration of mobile-based features, the choice of programming language, and the creation of dynamic fuzzy rules. Furthermore, the study examines the differences between the Mamdani Inference System and the Sugeno Fuzzy Inference method and explores challenges and opportunities in deploying the EMFES on mobile devices. The research identifies a critical gap in existing breast cancer diagnostic systems, emphasizing the need for a comprehensive, mobile-enabled, and adaptable solution by developing an EMFES that leverages Type-2 fuzzy logic, the Sugeno Inference Algorithm, Python Programming, and dynamic fuzzy rule generation. This study seeks to enhance early breast cancer detection and ultimately reduce breast cancer-related mortality.展开更多
Numerical simulation has been widely applied to the assessment of debris flow hazards. In East Asia and especially Taiwan, the most widely used numerical programs are FLO-2D and Debris-aD. Although these two programs ...Numerical simulation has been widely applied to the assessment of debris flow hazards. In East Asia and especially Taiwan, the most widely used numerical programs are FLO-2D and Debris-aD. Although these two programs are applied to the same engineering tasks, they are different in many aspects. These two programs were compared according to their fundamental theories, input and output data, computational algorithms and results. Using both programs, the simulations of a real debris flow with abundant granular material induced by landslides at Xinfa village in southern Taiwan are performed for comparison. The simulation results show that Debris- 2D gives better assessment in hazard area delineating and flow depth predicting. Therefore, Debris-2D is better for simulation of granular debris flows.展开更多
基金This research was financially supported by the Citrus Research and Development Foundation(CRDF).
文摘Pathogen-associated molecular patterns(PAMPs)-triggered immunity(PTI)is an important component of plant innate immunity.In a previous study,we showed that the PAMP flg22 from Xanthomonas citri ssp.citri(Xflg22),the causal agent of citrus canker,induced PTI in citrus,which correlated with the observed levels of canker resistance.Here,we identified and sequenced two bacterial flagellin/flg22 receptors(FLS2-1 and FLS2-2)from‘Duncan’grapefruit(Citrus paradisi,CpFLS2-1 and CpFLS2-2)and‘Sun Chu Sha’mandarin(C.reticulata,CrFLS2-1 and CrFLS2-2).We were able to isolate only one FLS2 from‘Nagami’kumquat(Fortunella margarita,FmFLS2-1)and gene flanking sequences suggest a rearrangement event that resulted in the deletion of FLS2-2 from the genome.Phylogenetic analysis,gene structure and presence of critical amino acid domains all indicate we identified the true FLS2 genes in citrus.FLS2-2 was more transcriptionally responsive to Xflg22 than FLS2-1,with induced expression levels higher in canker-resistant citrus than in susceptible ones.Interestingly,‘Nagami’kumquat showed the highest FLS2-1 steady-state expression levels,although it was not induced by Xflg22.We selected FmFLS2-1,CrFLS2-2 and CpFLS2-2 to further evaluate their capacity to enhance bacterial resistance using Agrobacterium-mediated transient expression assays.Both FmFLS2-1 and CrFLS2-2,the two proteins from canker-resistant species,conferred stronger Xflg22 responses and reduced canker symptoms in leaves of the susceptible grapefruit genotype.These two citrus genes will be useful resources to enhance PTI and achieve resistance against canker and possibly other bacterial pathogens in susceptible citrus types.
基金supported by grants from the National Natural Science Foundation of China (32160064 and 32360085)the Guangxi Natural Science Foundation (2020GXNSFFA297007)+2 种基金the Ba-Gui Scholar Program of Guangxi (to Z.G.H.)the State Key Laboratory for Conservation and Utilization of Subtropical Agro-bioresources (sklcusa-a02)the Innovation Project of Guangxi Graduate Education (YCBZ2023037).
文摘The receptor-like kinase FLAGELLIN-SENSITIVE 2(FLS2)functions as a bacterialflagellin receptor local-ized on the cell membrane of plants.In Arabidopsis,the co-receptor BRI1-ASSOCIATED RECEPTOR KI-NASE 1(BAK1)cooperates with FLS2 to detect theflagellin epitopeflg22,resulting in formation of a signaling complex that triggers plant defense responses.However,the co-receptor responsible for recog-nizing and signaling theflg22 epitope in rice remains to be determined,and the precise structural mecha-nism underlying FLS2-mediated signal activation and transduction has not been claried.This study pre-sents the structural characterization of a kinase-dead mutant of the intracellular kinase domain of OsFLS2(OsFLS2-KDD1013A)in complex with ATP or ADP,resolved at resolutions of 1.98 A˚and 2.09 A˚,respectively.Structural analysis revealed that OsFLS2 can adopt an active conformation in the absence of phosphorylation,although it exhibits only weak basal catalytic activity for autophosphorylation.Subse-quent investigations demonstrated that OsSERK2 effectively phosphorylates OsFLS2,which reciprocally phosphorylates OsSERK2,leading to complete activation of OsSERK2 and rapid phosphorylation of the downstream substrate receptor-like cytoplasmic kinases OsRLCK176 and OsRLCK185.Through mass spectrometry experiments,we successfully identied critical autophosphorylation sites on OsSERK2,as well as sites transphosphorylated by OsFLS2.Furthermore,we demonstrated the interaction between OsSERK2 and OsFLS2,which is enhanced in the presence offlg22.Genetic evidence suggests that OsRLCK176 and OsRLCK185 may function downstream of the OsFLS2-mediated signaling pathway.Our study reveals the molecular mechanism by which OsFLS2 mediates signal transduction pathways in rice and provides a valuable example for understanding RLK-mediated signaling pathways in plants.
文摘Infections with Salmonella enterica belong to the most prominent causes of food poisoning and infected fruits and vegetables represent important vectors for salmonellosis. Recent evidence indicates that plants recognize S. enterica and raise defense responses. Nonetheless, the molecular mechanisms controlling the interaction of S. enterica with plants are still largely unclear. Here, we show that flagellin from S. enterica represents a prominent pathogenassociated molecular pattern （PAMP） in Arabidopsis thaliana, which induces PAMP-triggered immunity （PTI） via the recognition of the fig22 domain by the receptor kinase FLS2. The Arabidopsis fls2 mutant shows reduced though not abolished PTI activation, indicating that plants rely also on recognition of other S. enterica PAMPs. Interestingly, the S. enterica type III secretion system （T3SS） mutant prgH- induced stronger defense gene expression than wild-type bacteria in Arabidopsis, suggesting that T3SS effectors are involved in defense suppression. Furthermore, we observe that S. enterica strains show variation in the fig22 epitope, which results in proteins with reduced PTI-inducing activity. Altogether, these results show that S. enterica activates PTI in Arabidopsis and suggest that, in order to accomplish plant colonization, S. enterica evolved strategies to avoid or suppress PTI.
基金supported by the National Natural Science Foundation of China(32172503 and 32260721)the Natural Science Foundation of Fujian Province,China(2023J01069)+2 种基金the State Key Laboratory of Ecological Pest Control for Fujian and Taiwan Crops,China(SKL2022001)the Innovation Fund of Fujan Agriculture and Forestry University,China(KFB23014A)the Undergraduate Training Program for Innovation and Entrepreneurship of Fujian Province,China(S202210389101).
文摘Genetic pest control strategies based on precise sex separation and only releasing sterile males can be accomplished by site-specific genome editing.In the current study,we showed that the mutation of single-allele Pxfl(2)d can significantly impair the normal mating behavior and testis development in male adults of the notorious cruciferous insect pest Plutella xylostella,in addition to its known functions in the ovarian development in female adults and egg hatching.Subsequent CRISPR/Cas9-based knock-in experiments revealed that site-specific integration of an exogenous green fluorescent protein(GFP)gene into autosomal Pxfl(2)d for labelling mutants could be achieved.However,this gene is not a suitable target for GFP insertion to establish a genetically stable knock-in strain because of the severe decline in reproductive capacity.We further screened for the W-chromosome-linked and Z-chromosome-linked regions to test the knock-in efficiency mediated by CRISPR/Cas9.The results verified that both types of chromosomes can be targeted for the site-specific insertion of exogenous sequences.We ultimately obtained a homozygous knock-in strain with the integration of both Cas9 and cyan fluorescent protein(CFP)expression cassettes on a Z-linked region in P.xylostella,which can also be used for early sex detection.By injecting the sgRNA targeting Pxfl(2)d alone into the eggs laid by female adults of the Z-Cas9-CFP strain,the gene editing efficiency reached 29.73%,confirming the success of expressing a functional Cas9 gene.Taken together,we demonstrated the feasibility of the knock-in of an exogenous gene to different genomic regions in P.xylostella,while the establishment of a heritable strain required the positioning of appropriate sites.This study provides an important working basis and technical support for further developing genetic strategies for insect pest control.
文摘Breast cancer remains a significant global health challenge, necessitating effective early detection and prognosis to enhance patient outcomes. Current diagnostic methods, including mammography and MRI, suffer from limitations such as uncertainty and imprecise data, leading to late-stage diagnoses. To address this, various expert systems have been developed, but many rely on type-1 fuzzy logic and lack mobile-based applications for data collection and feedback to healthcare practitioners. This research investigates the development of an Enhanced Mobile-based Fuzzy Expert system (EMFES) for breast cancer pre-growth prognosis. The study explores the use of type-2 fuzzy logic to enhance accuracy and model uncertainty effectively. Additionally, it evaluates the advantages of employing the python programming language over java for implementation and considers specific risk factors for data collection. The research aims to dynamically generate fuzzy rules, adapting to evolving breast cancer research and patient data. Key research questions focus on the comparative effectiveness of type-2 fuzzy logic, the handling of uncertainty and imprecise data, the integration of mobile-based features, the choice of programming language, and the creation of dynamic fuzzy rules. Furthermore, the study examines the differences between the Mamdani Inference System and the Sugeno Fuzzy Inference method and explores challenges and opportunities in deploying the EMFES on mobile devices. The research identifies a critical gap in existing breast cancer diagnostic systems, emphasizing the need for a comprehensive, mobile-enabled, and adaptable solution by developing an EMFES that leverages Type-2 fuzzy logic, the Sugeno Inference Algorithm, Python Programming, and dynamic fuzzy rule generation. This study seeks to enhance early breast cancer detection and ultimately reduce breast cancer-related mortality.
基金support from National Science Council of Chinese Taipei(Grant No.NSC 96-2625-Z-002-006-MY3)
文摘Numerical simulation has been widely applied to the assessment of debris flow hazards. In East Asia and especially Taiwan, the most widely used numerical programs are FLO-2D and Debris-aD. Although these two programs are applied to the same engineering tasks, they are different in many aspects. These two programs were compared according to their fundamental theories, input and output data, computational algorithms and results. Using both programs, the simulations of a real debris flow with abundant granular material induced by landslides at Xinfa village in southern Taiwan are performed for comparison. The simulation results show that Debris- 2D gives better assessment in hazard area delineating and flow depth predicting. Therefore, Debris-2D is better for simulation of granular debris flows.
