Identification and functional analysis of the type III effector AopBF1 in Acidovorax citrulli
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    Abstract:

    [Background] Bacterial fruit blotch is a prevalent quarantine disease caused by Acidovorax citrulli in cucurbit crops. A. citrulli secretes the key pathogenic factor, known as type III effectors (T3Es), into plant cells via the type III secretion system (T3SS). However, the pathogenesis of T3Es remains elusive. In previous studies, we have identified several candidate T3Es in A. citrulli FC440. [Objective] To reveal the sequence features, transport characteristics, and the role in the pathogenic process of the candidate T3E AopBF1 from A. citrulli FC440 strain, so as to lay a theoretical foundation for dissecting the mechanism of pathogenesis. [Methods] Bioinformatic tools were used to predict and analyze the T3E sequence characteristics of AopBF1. Then, RT-qPCR and an avirulent protein reporter system analysis were conducted to reveal the regulation and transport characteristics of AopBF1. The pathogenicity of aopBF1 mutant (insertion mutation) and overexpressing strain was investigated to unveil the role of AopBF1 in pathogenicity. [Results] AopBF1 had the typical T3E characteristics, possessing a protein kinase domain and no other conserved domain. The expression of AopBF1 was significantly down-regulated in the mutants of the T3SS core regulatory genes hrpX and hrpG. The co-expression of aopBF1 and the AvrBs1 functional region (59-445 aa) in the avrBs1 mutant induced hypersensitive reactions (HRs) in the leaves of ECW-10R pepper containing the Bs1 protein. The pathogenicity of the aopBF1 mutant in cucumber was significantly reduced, while the accumulation of hydrogen peroxide, superoxide anion radical, and callose significantly increased. The overexpression of aopBF1 significantly enhanced the pathogenicity and delayed HRs in Nicotiana benthamiana. The AopBF1 transiently expressed in N. benthamiana was localized in the cytoplasm, nucleus, and cell membrane, and then induced HRs and up-regulated the expression of PTI and hormone pathway-related genes. [Conclusion] AopBF1 is demonstrated to be a T3E with a protein kinase domain in A. citrulli. It can inhibit PTI immune responses such as host reactive oxygen species and callose accumulation to enhance the pathogenicity of A. citrulli and can trigger PTI and hormone-related immune responses in N. benthamiana containing R proteins.

    Reference
    [1] 赵廷昌, 孙福在, 王兵万. 西瓜细菌性果斑病研究进展[J]. 植保技术与推广, 2001(3):37-38, 36. ZHAO TC, SUN FZ, WANG BW. Review on the research of Acidovorax avenae subsp. citrulli[J]. Plant Protection Technology and Extension, 2001(3):37-38, 36(in Chinese).
    [2] WILLEMS A, GOOR M, THIELEMANS S, GILLIS M, KERSTERS K, de LEY J. Transfer of several phytopathogenic Pseudomonas species to Acidovorax as Acidovorax avenae subsp. avenae subsp. nov., comb. nov., Acidovorax avenae subsp. citrulli, Acidovorax avenae subsp. cattleyae, and Acidovorax konjaci[J]. International Journal of Systematic Bacteriology, 1992, 42(1):107-119.
    [3] SALMOND GPC, REEVES PJ. Membrance traffic wardens and protein secretion in Gram-negative bacteria[J]. Trends in Biochemical Sciences, 1993, 18(1):7-12.
    [4] ZHANG XX, ZHAO M, YAN JP, YANG LL, YANG YW, GUAN W, WALCOTT R, ZHAO TC. Involvement of hrpX and hrpG in the virulence of Acidovorax citrulli strain Aac5, causal agent of bacterial fruit blotch in cucurbits[J]. Frontiers in Microbiology, 2018, 9:507.
    [5] 张晓晓. 西瓜噬酸菌效应蛋白Ace1功能研究及光照黑暗条件下致病性差异分析[D]. 北京:中国农业科学院博士学位论文, 2018. ZHANG XX. Functional study of effector Ace1 and analysis of pathogenicity differences under light and dark conditions in Acidovorax citrulli[D]. Beijing:Doctoral Dissertation of Chinese Academy of Agricultural Sciences, 2018(in Chinese).
    [6] 杨琳琳. 西瓜噬酸菌效应蛋白Ace0201和Ace1242的鉴定及生物学功能初步分析[D]. 沈阳:沈阳农业大学硕士学位论文, 2019. YANG LL. Identification and biological function of the effector Ace0201 and Ace1242 in Acidovorax citrulli[D]. Shenyang:Master's Thesis of Shenyang Agricultural University, 2019(in Chinese).
    [7] ZHANG XX, YANG YW, ZHAO M, YANG LL, JIANG J, WALCOTT R, YANG SS, ZHAO TC. Acidovorax citrulli type III effector AopP suppresses plant immunity by targeting the watermelon transcription factor WRKY6[J]. Frontiers in Plant Science, 2020, 11:579218.
    [8] ZHANG XX, ZHAO M, JIANG J, YANG LL, YANG YW, YANG SS, WALCOTT R, QIU DW, ZHAO TC. Identification and functional analysis of AopN, an Acidovorax citrulli effector that induces programmed cell death in plants[J]. International Journal of Molecular Sciences, 2020, 21(17):6050.
    [9] 段凯莉, 江聪, 王光辉. 禾谷镰刀菌蛋白激酶研究进展[J]. 生物技术进展, 2021, 11(5):618-627. DUAN KL, JIANG C, WANG GH. Research progress of protein kinases in wheat scab fungus Fusarium graminearum[J]. Current Biotechnology, 2021, 11(5):618-627(in Chinese).
    [10] WANG CF, ZHANG SJ, HOU R, ZHAO ZT, ZHENG Q, XU QJ, ZHENG DW, WANG GH, LIU HQ, GAO XL, MA JW, KISTLER HC, KANG ZS, XU JR. Functional analysis of the kinome of the wheat scab fungus Fusarium graminearum[J]. Public Library of Science Pathogens, 2011, 7(12):e1002460.
    [11] DÜRRENBERGER F, WONG K, KRONSTAD JW. Identification of a cAMP-dependent protein kinase catalytic subunit required for virulence and morphogenesis in Ustilago maydis[J]. Proceedings of the National Academy of Sciences of the United States of America, 1998, 95(10):5684-5689.
    [12] 优丽图孜·乃比. 西瓜食酸菌与黄瓜互作转录组分析及T3SEs基因的初步鉴定[D]. 乌鲁木齐:新疆农业大学硕士学位论文, 2021. YOULITUZI NB. Transcriptome analysis of Acidovorax citrulli-cucumber interaction and preliminary identification of T3SE genes in Acidovorax citrulli[D]. Urumqi:Master's Thesis of Xinjiang Agricultural University, 2021(in Chinese).
    [13] 张美祥, 安玉艳, 刘廷利, 茹艳艳, 李文号, 窦道龙. 在本氏烟中瞬时表达效应因子PsCRN127基因提高其对寄生疫霉的抗性[J]. 南京农业大学学报, 2015, 38(6):930-935. ZHANG MX, AN YY, LIU TL, RU YY, LI WH, DOU DL. Transient expression of the PsCRN127 effector gene enhances Nicotiana benthamiana resistance to Phytophthora parasitica[J]. Journal of Nanjing Agricultural University, 2015, 38(6):930-935(in Chinese).
    [14] 韦红玉. 野油菜黄单胞菌效应物转运鉴定系统的构建[D]. 南宁:广西大学硕士学位论文, 2006. WEI HY. Construction of the identification system of the effector translocation in Xanthomonas campestris pv. campestris[D]. Nanning:Master's Thesis of Guangxi University, 2006(in Chinese).
    [15] 陈宝强, 马博雅, 李莹莹, 优丽图孜·乃比, 宋金迪, 刘君. 西瓜食酸菌Ⅲ型分泌效应物基因aopW功能初步分析[J]. 微生物学通报, 2023, 50(5):1973-1987. CHEN BQ, MA BY, LI YY, YOULITUZI NB, SONG JD, LIU J. Preliminary functional analysis of the type Ⅲ secreted effector gene aopW in Acidovorax citrulli[J]. Microbiology China, 2023, 50(5):1973-1987(in Chinese).
    [16] 池俊玲, 赵一博, 郭江波, 张龙, 刘汉阳, 辛翠花. 不同浓度Cd2+胁迫下烟草实时荧光定量PCR内参基因的筛选[J]. 南方农业学报, 2019, 50(10):2133-2140. CHI JL, ZHAO YB, GUO JB, ZHANG L, LIU HY, XIN CH. Screening of internal reference genes for real-time fluorescence quantitative PCR under different concentrations of Cd2+ stress in tobacco[J]. Journal of Southern Agriculture, 2019, 50(10):2133-2140(in Chinese).
    [17] 优丽图孜·乃比, 王希东, 刘君, 陈宝强, 宋金迪. 西瓜食酸菌与黄瓜互作转录组分析[J]. 微生物学通报, 2021, 48(10):3667-3681. YOULITUZI·NB, WANG XD, LIU J, CHEN BQ, SONG JD. Transcriptome analysis of the interaction between Acidovorax citrulli and cucumber[J]. Microbiology China, 2021, 48(10):3667-3681(in Chinese).
    [18] LOPEZ VA, PARK BC, NOWAK D, SREELATHA A, ZEMBEK P, FERNANDEZ J, SERVAGE KA, GRADOWSKI M, HENNIG J, TOMCHICK DR, PAWŁOWSKI K, KRZYMOWSKA M, TAGLIABRACCI VS. A bacterial effector mimics a host HSP90 client to undermine immunity[J]. Cell, 2019, 179(1):205-218.e21.
    [19] 颉兵兵, 刘君, 优丽图孜·乃比, 张春博. 西瓜食酸菌抗铜基因cueR的生物信息学分析及功能验证[J]. 微生物学通报, 2020, 47(5):1534-1543. XIE BB, LIU J, YOULITUZI NB, ZHANG CB. Bioinformatics analysis and functional verification of copper resistance gene cueR in Acidovorax citrulli[J]. Microbiology China, 2020, 47(5):1534-1543(in Chinese).
    [20] 陆巍, 许晓明, 张荣铣, 戴新宾. 冰醋酸对于测定植物材料中超氧阴离子含量的灵敏度的影响[J]. 南京师大学报(自然科学版), 2004, 27(1):82-84. LU W, XU XM, ZHANG RX, DAI XB. Effece of adding acetic acid on improvement of determination of superoxide anion content in plants[J]. Journal of Nanjing Normal University (Natural Science Edition), 2004, 27(1):82-84(in Chinese).
    [21] 张美祥, 刘廷利, 茹艳艳, 张琪梦, 窦道龙. 效应因子PsCRN77基因在本氏烟中的表达降低其对寄生疫霉的抗性[J]. 植物病理学报, 2015, 45(6):619-625. ZHANG MX, LIU TL, RU YY, ZHANG QM, DOU DL. Expression of an effector gene PsCRN77 decreases Nicotiana benthamiana resistance to oomycete pathogen Phytophthora parasitica[J]. Acta Phytopathologica Sinica, 2015, 45(6):619-625(in Chinese).
    [22] CHEN CL, LIU SS, LIU Q, NIU JH, LIU P, ZHAO JL, JIAN H. An ANNEXIN-like protein from the cereal cyst nematode Heterodera avenae suppresses plant defense[J]. the Public Library of Science One, 2015, 10(4):e0122256.
    [23] 刘琳硕, 贺祥, 李红梅, 元青, 王暄. TRV介导的E3泛素连接酶基因NbE3R14沉默对烟草基础免疫及南方根结线虫寄生的影响[J]. 南京农业大学学报, 2020, 43(1):65-71. LIU LS, HE X, LI HM, YUAN Q, WANG X. Effects of TRV-mediated silencing of the E3 ubiquitin ligase gene NbE3R14 in Nicotiana benthamiana on the plant basal immunity and the parasitism of Meloidogyne incognita[J]. Journal of Nanjing Agricultural University, 2020, 43(1):65-71(in Chinese).
    [24] 季苇芹, 叶云峰, 张爱萍, 杨玉文, 关巍, 赵廷昌. 我国瓜类细菌性果斑病研究新进展[J]. 中国瓜菜, 2022, 35(9):1-8. JI WQ, YE YF, ZHANG AP, YANG YW, GUAN W, ZHAO TC. Advances of bacterial fruit blotch in China[J]. China Cucurbits and Vegetables, 2022, 35(9):1-8(in Chinese).
    [25] TAMPAKAKI AP, SKANDALIS N, GAZI AD, BASTAKI MN, PANAGIOTIS FS, CHAROVA SN, KOKKINIDIS M, PANOPOULOS NJ. Playing the "harp":evolution of our understanding of hrp/hrc genes[J]. Annual Review of Phytopathology, 2010, 48(1):347-370.
    [26] WENGELNIK K, BONAS U. HrpXv, an AraC-type regulator, activates expression of five of the six loci in the hrp cluster of Xanthomonas campestris pv. vesicatoria[J]. Journal of Bacteriology, 1996, 178(12):3462-3469.
    [27] FENSELAU S. Sequence and expression analysis of the hrpB pathogenicity operon of Xanthomonas campestris pv. vesicatoria which encodes eight proteins with similarity to components of the Hrp, Vsc, Spa, and Fli secretion systems[J]. Molecular Plant-Microbe Interactions, 1995, 8(6):845.
    [28] 成春燕. 一个用于鉴定十字花科黑腐病菌III型效应物转运与分泌的报告质粒的构建[D]. 南宁:广西大学硕士学位论文, 2013. CHENG CY. Construction of one reports palsmid for the identification of transport and secretion of type Ⅲ effector in Xanthomonas campestris pv. campestris[D]. Nanning:Master's Thesis of Guangxi University, 2013(in Chinese).
    [29] 徐荣旗. 野油菜黄单胞菌野油菜致病变种新的依赖于Ⅲ型分泌系统的效应物的鉴定[D]. 南宁:广西大学博士学位论文, 2006. XU RQ. Identification of novel effectors depended on type Ⅲ secretion system in Xanthomonas campestris pv. campestris[D]. Nanning:Doctoral Dissertation of Guangxi University, 2006(in Chinese).
    [30] XU RQ, BLANVILLAIN S, FENG JX, JIANG BL, LI XZ, WEI HY, KROJ T, LAUBER E, ROBY D, CHEN BS, HE YQ, LU GT, TANG DJ, VASSE J, ARLAT M, TANG JL. AvrACXcc8004, a type III effector with a leucine-rich repeat domain from Xanthomonas campestris pathovar campestris confers avirulence in vascular tissues of Arabidopsis thaliana ecotype col-0[J]. Journal of Bacteriology, 2008, 190(1):343-355.
    [31] JIMÉNEZ-GUERRERO I, PÉREZ-MONTAÑO F, Da SILVA GM, WAGNER N, SHKEDY D, ZHAO M, PIZARRO L, BAR M, WALCOTT R, SESSA G, PUPKO T, BURDMAN S. Show me your secret(ed) weapons:a multifaceted approach reveals a wide arsenal of type III-secreted effectors in the cucurbit pathogenic bacterium Acidovorax citrulli and novel effectors in the Acidovorax genus[J]. Molecular Plant Pathology, 2020, 21(1):17-37.
    [32] CUNNAC S, BOUCHER C, GENIN S. Characterization of the cis-acting regulatory element controlling HrpB-mediated activation of the type III secretion system and effector genes in Ralstonia solanacearum[J]. Journal of Bacteriology, 2004, 186(8):2309-2318.
    [33] GARG RP, HUANG J, YINDEEYOUNGYEON W, DENNY TP, SCHELL MA. Multicomponent transcriptional regulation at the complex promoter of the exopolysaccharide I biosynthetic operon of Ralstonia solanacearum[J]. Journal of Bacteriology, 2000, 182(23):6659-6666.
    [34] WEI HL, ZHANG W, COLLMER A. Modular study of the type III effector repertoire in Pseudomonas syringae pv. tomato DC3000 reveals a matrix of effector interplay in pathogenesis[J]. Cell Reports, 2018, 23(6):1630-1638.
    [35] GUO M, TIAN F, WAMBOLDT Y, ALFANO JR. The majority of the type III effector inventory of Pseudomonas syringae pv. tomato DC3000 can suppress plant immunity[J]. Molecular Plant-Microbe Interactions:MPMI, 2009, 22(9):1069-1080.
    [36] TRAORE SM, ECKSHTAIN-LEVI N, MIAO JM, CASTRO SPARKS A, WANG ZB, WANG KR, LI Q, BURDMAN S, WALCOTT R, WELBAUM GE, ZHAO BY. Nicotiana species as surrogate host for studying the pathogenicity of Acidovorax citrulli, the causal agent of bacterial fruit blotch of cucurbits[J]. Molecular Plant Pathology, 2019, 20(6):800-814.
    [37] JENCZMIONKA NJ, MAIER FJ, LÖSCH AP, SCHÄFER W. Mating, conidiation and pathogenicity of Fusarium graminearum, the main causal agent of the head-blight disease of wheat, are regulated by the MAP kinase gpmk1[J]. Current Genetics, 2003, 43(2):87-95.
    [38] 马英, 赵冬梅, 杨志辉, 朱杰华. RxLR基因PITG-14788和PITG-19831促进致病疫霉对本氏烟的侵染[C]//中国植物病理学会. 中国植物病理学会2019年学术年会论文集. 北京:中国农业科学技术出版社, 2019:240. MA Y, ZHAO DM, YANG ZH, ZHU JH. The RxLR genes PITG-14788 and PITG-19831 promote the infection of Phytophthora infestans on Nicotiana benthamiana[C]//Chinese Society for Plant Pathology. Proceedings of the 2019 Annual Conference of the Chinese Society for Plant Pathology. Beijing:China Agriculture Science and Technology Press, 2019:240(in Chinese).
    [39] JELENSKA J, YAO N, VINATZER BA, WRIGHT CM, BRODSKY JL, GREENBERG JT. A J domain virulence effector of Pseudomonas syringae remodels host chloroplasts and suppresses defenses[J]. Current Biology:CB, 2007, 17(6):499-508.
    [40] CANONNE J, MARINO D, JAUNEAU A, POUZET C, BRIÈRE C, ROBY D, RIVAS S. The Xanthomonas type III effector XopD targets the Arabidopsis transcription factor MYB30 to suppress plant defense[J]. The Plant Cell, 2011, 23(9):3498-3511.
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MA Boya, LI Yingying, Youlituzi Naibi, CHEN Baoqiang, LIU Jun. Identification and functional analysis of the type III effector AopBF1 in Acidovorax citrulli[J]. Microbiology China, 2024, 51(1): 189-208

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  • Received:May 22,2023
  • Adopted:June 19,2023
  • Online: January 02,2024
  • Published: January 20,2024
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