草莓胶孢炭疽菌CFEM候选效应子的生物信息学鉴定及其侵染过程中的转录分析
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摘要
病原真菌通常分泌效应子到寄主组织中调控寄主的生理过程,从而有利于其侵染。CFEM(common in several fungal extracellular membrane)蛋白是真菌所独有的,且与致病性密切相关。本研究利用Pfam数据库对草莓胶孢炭疽菌全基因组进行搜索,鉴定获得22个CFEM蛋白。对CFEM蛋白的信号肽、跨膜结构域和亚细胞定位进行分析,结果表明仅有8个CFEM蛋白为分泌蛋白。对CFEM分泌蛋白在不同侵染阶段的转录情况进行转录组学及RT-PCR分析,结果显示8个CFEM蛋白在侵染后不同时期均有表达。其中,1个CFEM分泌蛋白于附着胞形成期特异表达,2个于活体寄生阶段特异表达,2个于死体寄生阶段特异表达。综合上述分析结果,预测这8个分泌蛋白可能为草莓胶孢炭疽菌的效应子。本研究为深入解析植物病原真菌CFEM效应子提供了理论依据。
Plant pathogens secrete effector proteins into host tissues to regulate host physiological processes and promote its infection.Common in several fungal extracellular membrane(CFEM)proteins is unique to fungi and plays an important role in pathogenesis.In our study,we obtained22 CFEM proteins of Colletotrichum gloeosporioides by searching Pfam database.Then,we conducted a bioinformatics analysis on signal peptide,transmembrane domain and protein subcellular localization of these 22 CFEM proteins.The results showed that only 8 of them were secreted proteins.The transcriptome and RT-PCR analysis of these 8 CFEM proteins at different stages of infection were performed.Among them,one protein was expressed in in planta appressorium stage,two were expressed in biotrophic stage and one was expressed in necrotrophic stage.Collectively,the 8 CFEM proteins were predicted to be the candidate effectors of C.gloeosporioides.Our findings provide a theoretical basis for in-depth analysis for the function of CFEM effectors in plant pathogenic fungi.
Plant pathogens secrete effector proteins into host tissues to regulate host physiological processes and promote its infection.Common in several fungal extracellular membrane(CFEM)proteins is unique to fungi and plays an important role in pathogenesis.In our study,we obtained22 CFEM proteins of Colletotrichum gloeosporioides by searching Pfam database.Then,we conducted a bioinformatics analysis on signal peptide,transmembrane domain and protein subcellular localization of these 22 CFEM proteins.The results showed that only 8 of them were secreted proteins.The transcriptome and RT-PCR analysis of these 8 CFEM proteins at different stages of infection were performed.Among them,one protein was expressed in in planta appressorium stage,two were expressed in biotrophic stage and one was expressed in necrotrophic stage.Collectively,the 8 CFEM proteins were predicted to be the candidate effectors of C.gloeosporioides.Our findings provide a theoretical basis for in-depth analysis for the function of CFEM effectors in plant pathogenic fungi.
引文
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[8]何芬.橡胶树炭疽病菌效应蛋白基因CgLysM的RNAi突变体构建及其蛋白的亚细胞定位[D].海口:海南大学,2014.
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[13]Horbach R,Navarro-Quesada A R,Knogge W,et al.When and how to kill a plant cell:Infection strategies of plant pathogenic fungi[J].Journal of Plant Physiology,2011,168(1):51-62.
[14]Koeck M,Hardham A R,Dodds P N.The role of effectors of biotrophic and hemibiotrophic fungi in infection[J].Cellular Microbiology,2011,13(12):1849-1857.
[15]Dodds P N,Rafiqi M,Gan P H P,et al.Effectors of biotrophic fungi and oomycetes:pathogenicity factors and triggers of host resistance[J].New Phytologist,2009,183(4):993-1000.
[16]Kamoun S.The secretome of plant-associated fungi and oomycetes[M]∥Deising H B.Plant relationships.Berlin:Springer,2009:173-180.
[17]Vleeshouwers V G,Oliver R P.Effectors as tools in disease resistance breeding against biotrophic,hemibiotrophic,and necrotrophic plant pathogens[J].Molecular Plant-Microbe Interactions:MPMI,2014,27(3):196-206.
[18]Weissman Z,Kornitzer D.A family of Candida cell surface haembinding proteins involved in haemin and haemoglobin-iron utilization[J].Molecular Microbiology,2004,53(4):1209-1220.
[19]Vaknin Y,Shadkchan Y,Levdansky E,et al.The three Aspergillus fumigatus,CFEM-domain GPI-anchored proteins(CfmAC)affect cell-wall stability but do not play a role in fungal virulence[J].Fungal Genetics&Biology,2014,63(1):55-64.
[20]Ling Jian,Zeng Feng,Cao Yuexia,et al.Identification of a class of CFEM proteins containing a new conserved motif in Fusarium oxysporum[J].Physiological&Molecular Plant Pathology,2015,89:41-48.
[21]Stephenson S A,Hatfield J,Rusu A G,et al.CgDN3:An essential pathogenicity gene of Colletotrichum gloeosporioides necessary to avert a hypersensitive-like response in the host Stylosanthes guianensis[J].Molecular Plant-Microbe Interactions,2000,13(9):929-941.
[22]Heijne G V.The signal peptide[J].Journal of Membrane Biology,1990,115(3):195-201.
[23]Presti L L,Lanver D,Schweizer G,et al.Fungal effectors and plant susceptibility[J].Annual Review of Plant Biology,2015,66:513-545.
[2]Hadwiger L A,George N.Agrios,Plant Pathology[M].5th ed.Amsterdam:Elsevier Academic Press,2005.
[3]Stergiopoulos I,Wit P J G M D.Fungal effector proteins[J].Annual Review of Phytopathology,2009,47(1):233-263.
[4]Jones J D G,Dangl J L.The plant immune system[J].Nature,2006,444(7117):323-329.
[5]Kamoun S.A catalogue of the effector secretome of plant pathogenic oomycetes[J].Annual Review of Phytopathology,2006,44(1):41-60.
[6]Gan P,Ikeda K,Irieda H,et al.Comparative genomic and transcriptomic analyses reveal the hemibiotrophic stage shift of Colletotrichum,fungi[J].New Phytologist,2013,197(4):1236-1249.
[7]汪倩,何朝族,罗红丽.橡胶树胶孢炭疽病菌CgBASP2基因敲除突变体构建及其致病力分析[J].热带生物学报,2015,6(1):41-46.
[8]何芬.橡胶树炭疽病菌效应蛋白基因CgLysM的RNAi突变体构建及其蛋白的亚细胞定位[D].海口:海南大学,2014.
[9]Kulkarni R D,Kelkar H S,Dean R A.An eight-cysteine-containing CFEM domain unique to a group of fungal membrane proteins[J].Trends in Biochemical Sciences,2003,28(3):118-121.
[10]Zhang Zhenna,Wu Qinyi,Zhang Guizhi,et al.Systematic analyses reveal uniqueness and origin of the CFEM domain in fungi[J].Scientific Reports,2015,5:13032.
[11]井忠英,王国梁,刘文德.禾谷炭疽菌CFEM效应子的生物信息学鉴定与转录分析[J].植物保护,2016,42(1):81-86.
[12]Mendgen K,Hahn M.Plant infection and the establishment of fungal biotrophy[J].Trends in Plant Science,2002,7(8):352-356.
[13]Horbach R,Navarro-Quesada A R,Knogge W,et al.When and how to kill a plant cell:Infection strategies of plant pathogenic fungi[J].Journal of Plant Physiology,2011,168(1):51-62.
[14]Koeck M,Hardham A R,Dodds P N.The role of effectors of biotrophic and hemibiotrophic fungi in infection[J].Cellular Microbiology,2011,13(12):1849-1857.
[15]Dodds P N,Rafiqi M,Gan P H P,et al.Effectors of biotrophic fungi and oomycetes:pathogenicity factors and triggers of host resistance[J].New Phytologist,2009,183(4):993-1000.
[16]Kamoun S.The secretome of plant-associated fungi and oomycetes[M]∥Deising H B.Plant relationships.Berlin:Springer,2009:173-180.
[17]Vleeshouwers V G,Oliver R P.Effectors as tools in disease resistance breeding against biotrophic,hemibiotrophic,and necrotrophic plant pathogens[J].Molecular Plant-Microbe Interactions:MPMI,2014,27(3):196-206.
[18]Weissman Z,Kornitzer D.A family of Candida cell surface haembinding proteins involved in haemin and haemoglobin-iron utilization[J].Molecular Microbiology,2004,53(4):1209-1220.
[19]Vaknin Y,Shadkchan Y,Levdansky E,et al.The three Aspergillus fumigatus,CFEM-domain GPI-anchored proteins(CfmAC)affect cell-wall stability but do not play a role in fungal virulence[J].Fungal Genetics&Biology,2014,63(1):55-64.
[20]Ling Jian,Zeng Feng,Cao Yuexia,et al.Identification of a class of CFEM proteins containing a new conserved motif in Fusarium oxysporum[J].Physiological&Molecular Plant Pathology,2015,89:41-48.
[21]Stephenson S A,Hatfield J,Rusu A G,et al.CgDN3:An essential pathogenicity gene of Colletotrichum gloeosporioides necessary to avert a hypersensitive-like response in the host Stylosanthes guianensis[J].Molecular Plant-Microbe Interactions,2000,13(9):929-941.
[22]Heijne G V.The signal peptide[J].Journal of Membrane Biology,1990,115(3):195-201.
[23]Presti L L,Lanver D,Schweizer G,et al.Fungal effectors and plant susceptibility[J].Annual Review of Plant Biology,2015,66:513-545.