稻瘟病菌分泌蛋白MoCDIE2诱导植物细胞死亡的功能分析

doi:10.13560/j.cnki.biotech.bull.1985.2019-0606

摘要/Abstract

摘要： 由稻瘟病菌（Magnaporthe oryzae）引起的稻瘟病是全球最严重的植物真菌病害之一。稻瘟病菌通过分泌效应蛋白进入与植物相互作用界面或转运到植物细胞内,抑制寄主植物的免疫防卫反应,使病原菌成功侵染。通过农杆菌介导的异源表达策略,筛选到能引起非寄主植物烟草细胞死亡的候选效应蛋白MoCDIE2（Cell Death-Inducing Effector）。序列分析表明：MoCDIE2基因编码一个蓖麻毒素B凝集素蛋白;系统发育树构建结果表明MoCDIE2同源蛋白保守存在于丝状真菌中;利用基因敲除的方法获得MoCDIE2的敲除突变体,结果表明MoCDIE2的敲除突变体在菌丝生长和致病性方面与野生型菌株Guy11没有明显差异。

关键词: 稻瘟病菌, MoCDIE2, 基因敲除, 蓖麻毒素B凝集素蛋白

Abstract: Rice blast,caused by the fungus Magnaporthe oryzae,is one of the most destructive plant diseases around the world. M. oryzae secretes many effector proteins into the interface or plant cells to suppress host immune defense response,thus helping the fungus to infect plant successfully. Using agrobacterium-mediated transient expression strategy,we obtained one putative effector protein MoCDIE2（Cell Death-Inducing Effector）that induces cell death in non-host plant Nicotiana benthamiana. Sequence alignment result indicated that MoCDIE2 gene encoded a ricin B lectin protein. Phylogenetic tree demonstrated that MoCDIE2 protein was conserved in many filamentous fungi. We used gene knock out method to generate MoCDIE2 knock out mutant,and the result showed that there were no significant differences between wild type strain Guy11 and mutants of MoCDIE2 knockout in mycelia growth and pathogenicity.

Key words: Magnaporthe oryzae, MoCDIE2, gene knockout, ricin B lectin protein

吴航, 李智强, 刘文德. 稻瘟病菌分泌蛋白MoCDIE2诱导植物细胞死亡的功能分析[J]. 生物技术通报, 2020, 36(1): 15-22.

WU Hang, LI Zhi-qiang, LIU Wen-de. Functional Analysis of Secreted Protein MoCDIE2 Inducing Plant Cell-Death in Magnaporthe oryzae[J]. Biotechnology Bulletin, 2020, 36(1): 15-22.

参考文献

[1] Pennisi E.Armed and dangerous[J]. Science, 2010, 327(5967):804-805.
[2] Dagdas Y, Yoshino K, Dagdas G, et al.Septin-mediated plant cell invasion by the rice blast fungus, Magnaporthe oryzae[J]. Science, 2012, 336(6088):1590-1595.
[3] Xu J, Hamer J.MAP kinase and cAMP signaling regulate infection structure formation and pathogenic growth in the rice blast fungus Magnaporthe grisea[J]. Genes Dev, 1996, 10(21):2696-2706.
[4] Xu J, Staiger C, Hamer J.Inactivation of the mitogen-activated protein kinase Mps1 from the rice blast fungus prevents penetration of host cells but allows activation of plant defense responses[J]. Proc Natl Acad Sci USA, 1998, 95(21):12713-12718.
[5] Dixon K, Xu J, Smirnoff N, et al.Independent signaling pathways regulate cellular turgor during hyperosmotic stress and appressorium-mediated plant infection by Magnaporthe grisea[J]. Plant Cell, 1999, 11(10):2045-2058.
[6] Choi J, Kim Y, Kim S, et al.MoCRZ1, a gene encoding a calcineurin-responsive transcription factor, regulates fungal growth and pathogenicity of Magnaporthe oryzae[J]. Fungal Genet Biol, 2009, 46(3):243-254.
[7] Adachi K, Hamer J.Divergent cAMP signaling pathways regulate growth and pathogenesis in the rice blast fungus Magnaporthe grisea[J]. Plant Cell, 1998, 10(8):1361-1374.
[8] Chisholm S, Coaker G, Day B, et al.Host-microbe interactions:shaping the evolution of the plant immune response[J]. Cell, 2006, 124(4):803-814.
[9] Jones J, Dangl J.The plant immune system[J]. Nature, 2006, 444(7117):323-329.
[10] Zhang J, Zhou J.Plant immunity triggered by microbial molecular signatures[J]. Mol Plant, 2010, 3(5):783-793.
[11] Dodds P, Rathjen J.Plant immunity:towards an integrated view of plant-pathogen interactions[J]. Nat Rev Genet, 2010, 11(8):539-548.
[12] Boller T, Felix G.A renaissance of elicitors:perception of microbe-associated molecular patterns and danger signals by pattern-recognition receptors[J]. Annu Rev Plant Biol, 2009, 60:379-406.
[13] Jia Y, Mcadams S, Bryan G, et al.Direct interaction of resistance gene and avirulence gene products confers rice blast resistance[J]. EMBO J, 2000, 19(15):4004-4014.
[14] Park C, Chen S, Shirsekar G, et al.The Magnaporthe oryzae effector AvrPiz-t targets the ring E3 ubiquitin ligase APIP6 to suppress pathogen-associated molecular pattern-triggered immunity in rice[J]. Plant Cell, 2012, 24(11):4748-4762.
[15] Cunnac S, Lindeberg M, Collmer A.Pseudomonas syringae type III secretion system effectors:repertoires in search of functions[J]. Curr Opin Microbiol, 2009, 12(1):53-60.
[16] Lo PL, Lanver D, Schweizer G, et al.Fungal effectors and plant susceptibility[J]. Annu Rev Plant Biol, 2015, 66:513-545.
[17] Whisson S, Boevink P, Moleleki L, et al.A translocation signal for delivery of oomycete effector proteins into host plant cells[J]. Nature, 2007, 450(7166):115-118.
[18] Sweigard J, Carroll A, Kang S, et al.Identification, cloning, and characterization of PWL2, a gene for host species specificity in the rice blast fungus[J]. Plant Cell, 1995, 7(8):1221-1233.
[19] Kang S, Sweigard J, Valent B.The PWL host specificity gene family in the blast fungus Magnaporthe grisea[J]. Mol Plant Microbe Interact, 1995, 8(6):939-948.
[20] Ribot C, Cesari S, Abidi I, et al.The Magnaporthe oryzae effector AVR1-CO39 is translocated into rice cells independently of a fungal-derived machinery[J]. Plant J, 2013, 74(1):1-12.
[21] Wu J, Kou Y, Bao J, et al.Comparative genomics identifies the Magnaporthe oryzae avirulence effector AvrPi9 that triggers Pi9-mediated blast resistance in rice[J]. New Phytol, 2015, 206(4):1463-1475.
[22] Saitoh H, Fujisawa S, Mitsuoka C, et al.Large-scale gene disruption in Magnaporthe oryzae identifies MC69, a secreted protein required for infection by monocot and dicot fungal pathogens[J]. PLoS Pathog, 2012, 8(5):e1002711.
[23] Dean R, Talbot N, et al.The genome sequence of the rice blast fungus Magnaporthe grisea[J]. Nature, 2005, 434(7036):980-986.
[24] Yoshida K, Saitoh H, Fujisawa S, et al.Association genetics reveals three novel avirulence genes from the rice blast fungal pathogen Magnaporthe oryzae[J]. Plant Cell, 2009, 21(5):1573-1591.
[25] Brenner S, Johnson M, Bridgham J, et al.Gene expression analysis by massively parallel signature sequencing(MPSS)on microbead arrays[J]. Nat Biotechnol, 2000, 18(6):630-634.
[26] Nobuta K, Venu R, Lu C, et al.An expression atlas of rice mRNAs and small RNAs[J]. Nat Biotechnol, 2007, 25(4):473-477.
[27] Gowda M, Jantasuriyarat C, Dean R, et al.Robust-LongSAGE(RL-SAGE):a substantially improved LongSAGE method for gene discovery and transcriptome analysis[J]. Plant Physiol, 2004, 134(3):890-897.
[28] German M, Pillay M, Jeong D, et al.Global identification of microRNA-target RNA pairs by parallel analysis of RNA ends[J]. Nat Biotechnol, 2008, 26(8):941-946.
[29] Chen S, Songkumarn P, Venu R, et al.Identification and characterization of in-planta expressed secreted effector proteins from Magnaporthe oryzae that induce cell death in rice[J]. Mol Plant Microbe Interact, 2013, 26(2):191-202.
[30] Anderson RG, Deb D, Fedkenheuer K, et al.Recent progress in RXLR effector research[J]. Mol Plant Microbe Interact, 2015, 28(10):1063-1072.
[31] Li W, Wang B, Wu J, et al.The Magnaporthe oryzae avirulence gene AvrPiz-t encodes a predicted secreted protein that triggers the immunity in rice mediated by the blast resistance gene Piz-t[J]. Mol Plant Microbe Interact, 2009, 22(4):411-420.
[32] Liu L, Dong Y, Li J, et al.The apoptosis of mouse iymphocytes induced by ricin toxin[J]. China Animal Husbandry & Veterinary Medicine, 2009. 36(2):46-50.