Plant bacterial diseases have infiicted substantial economic losses in global crop,fruit,and vegetable production.The conventional methods for managing these diseases typically rely on the application of antibiotics.H...Plant bacterial diseases have infiicted substantial economic losses in global crop,fruit,and vegetable production.The conventional methods for managing these diseases typically rely on the application of antibiotics.However,these antibiotics often target the growth factors of the pathogenic bacteria,leading to the accumulation and emergence of drug-resistant strains,which exacerbates antibiotic resistance.Innovative methods are urgently needed to treat and prevent the toxicity caused by these pathogenic bacteria.Targeting virulence mechanisms in pathogens is a globally recognized and effective strategy for mitigating bacterial resistance.TypeⅢsecretion system(T3SS)serves as a crucial virulence determinant in Gram-negative pathogens,and its non-essentials for pathogen growth renders it an ideal target.Targeting the T3SS holds significant potential to alleviate selective pressure for resistance mutations in pathogens.Therefore,targeting T3SS in pathogenic bacteria,while preserving their growth,has emerged as a novel avenue for the development of antimicrobial drugs.In recent years,a multitude of small molecular inhibitors targeting T3SS have been identified.This article offers a comprehensive review of T3SS inhibitors in plant pathogens,while also presenting the latest research advancements in this research direction.展开更多
In this study,Hy322 gene was cloned from Vibrio alginolyticus.The total length of its gene was 969 bp,and it could encode 322 amino acids.The physicochemical properties,protein structure,genetic evolutionary relations...In this study,Hy322 gene was cloned from Vibrio alginolyticus.The total length of its gene was 969 bp,and it could encode 322 amino acids.The physicochemical properties,protein structure,genetic evolutionary relationship and antigenic characteristics of the effector protein Hy322 of V.alginolyticus HY9901 type Ⅲ secretion system were studied and analyzed by bioinformatics methods and tools.The results showed that Hy322 is an unstable hydrophilic and acidic protein without a transmembrane region and a signal peptide,and secondary structure to α-helix.The evolutionary analysis showed that V.alginolyticus HY9901 and V.harveyi were clustered together,which indicated that the genetic relationship between the two species was closest.HY322 contains a FliN super family conserved domain associated with Flagellar motor switch.Bioinformatics analysis showed that the B-cell preponderant epitopes of Hy322 might be localized in the regions of 32-33,100-102,138-140,215-216,235-238 and 246-249.The 3D structure model of Hy322 subunit was simulated by SWISS-MODEL software and itwas found that the yscQ of Yersinia were similar and the similarity was 42.25%.In this study,the feasibility of Hy322 as a common antigen of Vibrio was verified from the perspective of bioinformatics,which laid the foundation for the next step in vaccine development.展开更多
The innate immune system is the first line of defense against invading pathogens. Innate immune cells recognize molecular patterns from the pathogen and mount a response to resolve the infection. The production of pro...The innate immune system is the first line of defense against invading pathogens. Innate immune cells recognize molecular patterns from the pathogen and mount a response to resolve the infection. The production of proinflammatory cytokines and reactive oxygen species, phagocytosis, and induced programmed cell death are processes initiated by innate immune cells in order to combat invading pathogens. However, pathogens have evolved various virulence mechanisms to subvert these responses. One strategy utilized by Gram-negative bacterial pathogens is the deployment of a complex machine termed the type Ⅲ secretion system(T3SS). The T3SS is composed of a syringe-like needle structure and the effector proteins that are injected directly into a target host cell to disrupt a cellular response. The three human pathogenic Yersinia spp.(Y. pestis, Y. enterocolitica, and Y. pseudotuberculosis) are Gramnegative bacteria that share in common a 70 kb virulence plasmid which encodes the T3 SS. Translocation of the Yersinia effector proteins(YopE, YopH, YopT, YopM, YpkA/YopO, and YopP/J) into the target host cell results in disruption of the actin cytoskeleton to inhibit phagocytosis, downregulation of proinflammatory cytokine/chemokine production, and induction of cellular apoptosis of the target cell. Over the past 25 years, studies on the Yersinia effector proteins have unveiled tremendous knowledge of how the effectors enhance Yersinia virulence. Recently, the long awaited crystal structure of YpkA has been solved providing further insights into the activation of the YpkA kinase domain. Multisite autophosphorylation by YpkA to activate its kinase domain was also shown and postulated to serve as a mechanism to bypass regulation by host phosphatases. In addition, novel Yersinia effector protein targets, such as caspase-1, and signaling pathways including activation of the inflammasome were identified. In this review, we summarize the recent discoveries made on Yersinia effector proteins and their contribution to Yersinia pathogenesis.展开更多
Ralstonia solanacearum is a widespread plant bacterial pathogen that can launch a range of type Ⅲ effectors(T3Es)to cause disease.In this study,we isolate a pathogenic R.solanacearum strain named P380 from tomato rhi...Ralstonia solanacearum is a widespread plant bacterial pathogen that can launch a range of type Ⅲ effectors(T3Es)to cause disease.In this study,we isolate a pathogenic R.solanacearum strain named P380 from tomato rhizosphere.Five out of 12 core T3Es of strain P380 are introduced into Pseudomonas syringae DC3000D36E separately to determine their functions in interacting with plants.DC3000D36E that harbors each effector suppresses FliC-triggered Pti5 and ACRE31 expression,ROS burst,and callose deposition.RipAE,RipU,and RipW elicit cell death as well as upregulate the MAPK cascades in Nicotiana benthamiana.The derivatives RipC1^(△DDXDX(T/V))and RipW^(△DDKXXQ)but not RipAE^(K310R) fail to suppress ROS burst.Moreover,RipAE^(K310R) and RipW^(△DDKXXQ) retain the cell death elicitation ability.RipAE and RipW are associated with salicylic acid and jasmonic acid pathways,respectively.RipAE and RipAQ significantly promote the propagation of DC3000D36E in plants.The five core T3Es localize in diverse subcellular organelles of nucleus,plasma membrane,endoplasmic reticulum,and Golgi network.The suppressor of G2 allele of Skp1 is required for RipAE but not RipU-triggered cell death in N.benthamiana.These results indicate that the core T3Es in R.solanacearum play diverse roles in plantpathogen interactions.展开更多
Objective Pseudomonas aeruginosa is a ubiquitous and opportunistic pathogen that uses the type Ⅲ secretion system （TTSS） to inject effector proteins directly into the cytosol of target cells to subvert the host cel...Objective Pseudomonas aeruginosa is a ubiquitous and opportunistic pathogen that uses the type Ⅲ secretion system （TTSS） to inject effector proteins directly into the cytosol of target cells to subvert the host cell＇s functions. Specialized bacterial chaperones are required for effective secretion of some effectors. To identify the chaperone of ExoS, the representative effector secreted by the TTSS of P aeruginosa, we analyzed the role of a postulated chaperone termed Orfl. Methods By allelic exchange, we constructed the mutant with the deletion of gene Orfl. Analysis of secreted and cell-associated fractions was performed by SDS-PAGE and Western blotting. Using strain expressing in trans Orfl, tagged by V5 polypeptide and histidine, protein-protein interaction was determined by affinity resin pull-down assay in combination with MALDI-TOF The role of Orfl in the expression of exoS was evaluated by gene reporter analysis. Results Pull-down assay showed that Orfl binds to ExoS and ExoT. Secretion profile analysis showed that Orfl was necessary for the optimal secretion of ExoS and ExoT. However, Orfl had no effect on the expression of exoS. Conclusion Orfl is important for the secretion of ExoS probably by maintaining ExoS in a secretion-competent conformation. We propose to name Orfl as SpcS for ＂specific Pseudomonas chaperone for ExoS＂.展开更多
杀香鱼假单胞菌是大黄鱼内脏白点病的致病菌,感染该菌常会导致养殖大黄鱼很高的死亡率和严重的经济损失。病原株NB2011编码典型的Ⅲ型分泌系统,可能是该菌重要的毒力因子,ExsA是控制此分泌系统表达的重要调控蛋白。为确认ExsA在NB2011...杀香鱼假单胞菌是大黄鱼内脏白点病的致病菌,感染该菌常会导致养殖大黄鱼很高的死亡率和严重的经济损失。病原株NB2011编码典型的Ⅲ型分泌系统,可能是该菌重要的毒力因子,ExsA是控制此分泌系统表达的重要调控蛋白。为确认ExsA在NB2011致病过程中的作用,开发有效疫苗,实验采用双交换同源重组法构建了ExsA内部序列被卡那霉素基因替换的突变株,检测突变株与野生株对鼠巨噬细胞J774的黏附、内化和胞内增殖特性,并比较对大黄鱼的毒力变化,同时,通过透射电子显微镜观察人工感染后大黄鱼内脏组织的病理变化。研究表明,巨噬细胞对突变株的内化率降低,内化的细菌在12 h内被清除,野生株在内化后虽然一段时间内数量稍有下降,12~24 h期间数量急剧上升;突变株对大黄鱼的96 h LD_(50)为2.59×10~7/mL,比野生株高数百倍;电镜切片中未观察到组织内有菌体的存在,表明突变株的毒力明显减弱,可以作为弱毒疫苗的开发对象。展开更多
基金the financial support from the National Key Research and Development Program of China(No.2023YFD1701100)the National Natural Science Foundation of China(No.32072450)+2 种基金the National Science Fund for Distinguished Young Scholars of Guangdong Province(No.2021B1515020107)the Opening Foundation of Hubei Key Laboratory of Novel Reactor and Green Chemical Technology(No.NRG202306)the Opening Foundation of Guangdong Province Key Laboratory of Microbial Signals and Disease Control(No.MSDC2023-19)。
文摘Plant bacterial diseases have infiicted substantial economic losses in global crop,fruit,and vegetable production.The conventional methods for managing these diseases typically rely on the application of antibiotics.However,these antibiotics often target the growth factors of the pathogenic bacteria,leading to the accumulation and emergence of drug-resistant strains,which exacerbates antibiotic resistance.Innovative methods are urgently needed to treat and prevent the toxicity caused by these pathogenic bacteria.Targeting virulence mechanisms in pathogens is a globally recognized and effective strategy for mitigating bacterial resistance.TypeⅢsecretion system(T3SS)serves as a crucial virulence determinant in Gram-negative pathogens,and its non-essentials for pathogen growth renders it an ideal target.Targeting the T3SS holds significant potential to alleviate selective pressure for resistance mutations in pathogens.Therefore,targeting T3SS in pathogenic bacteria,while preserving their growth,has emerged as a novel avenue for the development of antimicrobial drugs.In recent years,a multitude of small molecular inhibitors targeting T3SS have been identified.This article offers a comprehensive review of T3SS inhibitors in plant pathogens,while also presenting the latest research advancements in this research direction.
基金Supported by Shenzhen Science and Technology Project(JCYJ20170818111629778,JCYJ20170306161613251)National Natural Science Foundation of Guangdong Province(2017A030313174)+1 种基金Natural Science Foundation of Guangdong Ocean University(C17379)Undergraduate Innovative and Entrepreneurial Team Project(CCTD201802)
文摘In this study,Hy322 gene was cloned from Vibrio alginolyticus.The total length of its gene was 969 bp,and it could encode 322 amino acids.The physicochemical properties,protein structure,genetic evolutionary relationship and antigenic characteristics of the effector protein Hy322 of V.alginolyticus HY9901 type Ⅲ secretion system were studied and analyzed by bioinformatics methods and tools.The results showed that Hy322 is an unstable hydrophilic and acidic protein without a transmembrane region and a signal peptide,and secondary structure to α-helix.The evolutionary analysis showed that V.alginolyticus HY9901 and V.harveyi were clustered together,which indicated that the genetic relationship between the two species was closest.HY322 contains a FliN super family conserved domain associated with Flagellar motor switch.Bioinformatics analysis showed that the B-cell preponderant epitopes of Hy322 might be localized in the regions of 32-33,100-102,138-140,215-216,235-238 and 246-249.The 3D structure model of Hy322 subunit was simulated by SWISS-MODEL software and itwas found that the yscQ of Yersinia were similar and the similarity was 42.25%.In this study,the feasibility of Hy322 as a common antigen of Vibrio was verified from the perspective of bioinformatics,which laid the foundation for the next step in vaccine development.
基金Supported by the ASM Robert D Watkins Graduate FellowshipUC Davis Hellman Fellowship
文摘The innate immune system is the first line of defense against invading pathogens. Innate immune cells recognize molecular patterns from the pathogen and mount a response to resolve the infection. The production of proinflammatory cytokines and reactive oxygen species, phagocytosis, and induced programmed cell death are processes initiated by innate immune cells in order to combat invading pathogens. However, pathogens have evolved various virulence mechanisms to subvert these responses. One strategy utilized by Gram-negative bacterial pathogens is the deployment of a complex machine termed the type Ⅲ secretion system(T3SS). The T3SS is composed of a syringe-like needle structure and the effector proteins that are injected directly into a target host cell to disrupt a cellular response. The three human pathogenic Yersinia spp.(Y. pestis, Y. enterocolitica, and Y. pseudotuberculosis) are Gramnegative bacteria that share in common a 70 kb virulence plasmid which encodes the T3 SS. Translocation of the Yersinia effector proteins(YopE, YopH, YopT, YopM, YpkA/YopO, and YopP/J) into the target host cell results in disruption of the actin cytoskeleton to inhibit phagocytosis, downregulation of proinflammatory cytokine/chemokine production, and induction of cellular apoptosis of the target cell. Over the past 25 years, studies on the Yersinia effector proteins have unveiled tremendous knowledge of how the effectors enhance Yersinia virulence. Recently, the long awaited crystal structure of YpkA has been solved providing further insights into the activation of the YpkA kinase domain. Multisite autophosphorylation by YpkA to activate its kinase domain was also shown and postulated to serve as a mechanism to bypass regulation by host phosphatases. In addition, novel Yersinia effector protein targets, such as caspase-1, and signaling pathways including activation of the inflammasome were identified. In this review, we summarize the recent discoveries made on Yersinia effector proteins and their contribution to Yersinia pathogenesis.
基金supported by the National Key R&D Program of China(2019YFD1002000)the Science and Technology Programs of the Shandong Tobacco(KN273)Zunyi Tobacco(2021XM03).
文摘Ralstonia solanacearum is a widespread plant bacterial pathogen that can launch a range of type Ⅲ effectors(T3Es)to cause disease.In this study,we isolate a pathogenic R.solanacearum strain named P380 from tomato rhizosphere.Five out of 12 core T3Es of strain P380 are introduced into Pseudomonas syringae DC3000D36E separately to determine their functions in interacting with plants.DC3000D36E that harbors each effector suppresses FliC-triggered Pti5 and ACRE31 expression,ROS burst,and callose deposition.RipAE,RipU,and RipW elicit cell death as well as upregulate the MAPK cascades in Nicotiana benthamiana.The derivatives RipC1^(△DDXDX(T/V))and RipW^(△DDKXXQ)but not RipAE^(K310R) fail to suppress ROS burst.Moreover,RipAE^(K310R) and RipW^(△DDKXXQ) retain the cell death elicitation ability.RipAE and RipW are associated with salicylic acid and jasmonic acid pathways,respectively.RipAE and RipAQ significantly promote the propagation of DC3000D36E in plants.The five core T3Es localize in diverse subcellular organelles of nucleus,plasma membrane,endoplasmic reticulum,and Golgi network.The suppressor of G2 allele of Skp1 is required for RipAE but not RipU-triggered cell death in N.benthamiana.These results indicate that the core T3Es in R.solanacearum play diverse roles in plantpathogen interactions.
基金This research was supported by the association "Vaincre la Mucoviscidose" of France
文摘Objective Pseudomonas aeruginosa is a ubiquitous and opportunistic pathogen that uses the type Ⅲ secretion system （TTSS） to inject effector proteins directly into the cytosol of target cells to subvert the host cell＇s functions. Specialized bacterial chaperones are required for effective secretion of some effectors. To identify the chaperone of ExoS, the representative effector secreted by the TTSS of P aeruginosa, we analyzed the role of a postulated chaperone termed Orfl. Methods By allelic exchange, we constructed the mutant with the deletion of gene Orfl. Analysis of secreted and cell-associated fractions was performed by SDS-PAGE and Western blotting. Using strain expressing in trans Orfl, tagged by V5 polypeptide and histidine, protein-protein interaction was determined by affinity resin pull-down assay in combination with MALDI-TOF The role of Orfl in the expression of exoS was evaluated by gene reporter analysis. Results Pull-down assay showed that Orfl binds to ExoS and ExoT. Secretion profile analysis showed that Orfl was necessary for the optimal secretion of ExoS and ExoT. However, Orfl had no effect on the expression of exoS. Conclusion Orfl is important for the secretion of ExoS probably by maintaining ExoS in a secretion-competent conformation. We propose to name Orfl as SpcS for ＂specific Pseudomonas chaperone for ExoS＂.
文摘许多革兰氏阴性菌借助Ⅲ型分泌系统黏附在宿主细胞表面,然后跨越胞膜将特异性蛋白注入宿主细胞内,破坏宿主细胞内的多种信号通路,从而有利于细菌的感染及定殖。在肠致病性大肠杆菌(Enteropathogenic Escherichia coli,EPEC)中,除了肠细胞脱落位点(Locus of entericyte effacement,LEE)毒力岛编码的Ⅲ型分泌系统(Type Ⅲ secretion system,T3SS)外,在分析肠出血性大肠杆菌O157:H7的基因组序列时发现一个新的Ⅲ型分泌系统,大肠杆菌Ⅲ型分泌系统2(Escherichia coli type Ⅲ secretion system 2,ETT2)毒力岛。研究显示,ETT2可能在大多数菌株中不具有完整的分泌系统功能,但是其对于细菌毒力的发挥具有重要作用。因此,本文简要综述了大肠杆菌ETT2的基因特征、ETT2的分布与流行、ETT2的功能与机制等方面的主要研究进展。
文摘杀香鱼假单胞菌是大黄鱼内脏白点病的致病菌,感染该菌常会导致养殖大黄鱼很高的死亡率和严重的经济损失。病原株NB2011编码典型的Ⅲ型分泌系统,可能是该菌重要的毒力因子,ExsA是控制此分泌系统表达的重要调控蛋白。为确认ExsA在NB2011致病过程中的作用,开发有效疫苗,实验采用双交换同源重组法构建了ExsA内部序列被卡那霉素基因替换的突变株,检测突变株与野生株对鼠巨噬细胞J774的黏附、内化和胞内增殖特性,并比较对大黄鱼的毒力变化,同时,通过透射电子显微镜观察人工感染后大黄鱼内脏组织的病理变化。研究表明,巨噬细胞对突变株的内化率降低,内化的细菌在12 h内被清除,野生株在内化后虽然一段时间内数量稍有下降,12~24 h期间数量急剧上升;突变株对大黄鱼的96 h LD_(50)为2.59×10~7/mL,比野生株高数百倍;电镜切片中未观察到组织内有菌体的存在,表明突变株的毒力明显减弱,可以作为弱毒疫苗的开发对象。
