摘要
辣椒疫霉菌(Phytophthora capsici Leonian)是一种普遍存在的土壤兼性寄生菌,在世界范围内广泛引起辣椒疫病,是严重危害辣椒生产的重要病原之一。世界上种植辣椒的地区,由于辣椒疫病的发生严重影响了辣椒的产量和质量,造成了巨大的经济损失,对环境也造成了一定的影响。
近年来,辣椒生产国开展了许多关于抗疫病育种和抗病性分析的研究,对抗病性生理生化机制,抗病性遗传,抗病基因的定位与克隆以及抗病性鉴定和抗病育种方面进行了大量研究但是进展缓慢。目前辣椒疫霉的防治还是以化学药剂防治为主,农药残留等问题仍然给辣椒生产、人类健康和环境安全带来了巨大威胁。因此,寻找探索辣椒疫霉菌重要的致病因子及其致病机制成为人们研究的重点。
研究表明,在植物病原菌与寄主互作过程中,大多数植物病原会产生一系列调控植物生化、生理、形态学特征的效应分子来侵染寄主或调节植物的防卫反应。皱缩坏死(CRN)蛋白就是一类效应分子——胞质效应蛋白,在病原菌与寄主互作过程中通过附着胞和吸器等特殊的结构进入植物活细胞,调节植物的防卫反应。皱缩坏死(CRN)基因是Torto等人于2003年首次在马铃薯致病疫霉(Phytophthora infestans)胞外蛋白cDNA数据库中发现的,其在农杆菌介导的瞬时表达体系中导致本氏烟产生皱缩坏死(crinkling and necrosis)的症状反应。
CRN效应蛋白做为一类新的效应蛋白,目前只是在疫霉中发现了这类基因,对这类基因的研究还不多。在寄主与病原菌互作过程中,皱缩坏死蛋白的致病机制、进入细胞机制、作用靶标等都不清楚,所以成为人们研究的热点。
本研究以实验室保存的高致病性辣椒疫霉菌株SD33为实验材料,对辣椒疫霉中的皱缩坏死基因进行了以下研究:
(1)根据文献中报道的CRN基因的保守序列,我们从辣椒疫霉基因组全序列中筛选CRN-like序列,然后再NCBI-BLAST在线比对,确认筛选的基因属于CRN基因家族;
(2)从本实验室保存的高致病性辣椒疫霉菌株基因组中,成功克隆了两个CRN基因,PcCRN1和PcCRN2,并在线比对和进行生物信息学分析;
(3)为了了解PcCRN1和PcCRN2在辣椒疫霉菌中的致病性和保守序列LQLFLAK和WL在致病中的作用,对两个基因的保守位点进行突变,利用pGR106构建PcCRN1和PcCRN2及保守位点突变基因的PVX表达载体,通过农杆菌介导的瞬时表达进行体外功能检测;PcCRN1和PcCRN2在农杆菌中瞬时表达后均可产生褪绿症状;这两个基因保守位点突变前后在本氏烟中瞬时表达也均有差异,说明PcCRN1和PcCRN2在辣椒疫霉菌中起作用,保守位点的突变对其致病性也有影响;
(4)利用pHam34载体构建PcCRN1和PcCRN2全长基因的沉默载体,通过PEG介导的原生质体共转化技术获得PcCRN1和PcCRN2基因沉默的转化子,通过RT-PCR和qRT-PCR技术检验并接种验证辣椒疫霉中CRN的功能。
通过本研究,我们可以看出CRN基因在辣椒疫霉菌的致病过程中起作用,沉默转化子的成功筛选也证明了PEG介导的原生质体共转化技术在P. capsici中可以实施,为辣椒疫霉的分子致病机制研究提供了一种分子生物学技术。
Phytophthora capsici Leonian, a ubiquitous soil facultative parasite, can cause pepper blight disease in the world-wide and which is one of the important pathogens in reducing pepper production. In pepper-growing areas around the world, the widespread occurrence of the pepper blight disease seriously affects the yield and quality of pepper, and causes enormous economic losses as well as environmental damage in natural ecosystems.
For decades, pepper-producing countries have carried out a lot of researches about breeding and disease resistance, and have performed extensive work on physiological and biochemical mechanisms of disease resistance, genetic resistance, resistance gene mapping and cloning and disease resistance breeding but the progress is slow. Now, the prevention of P. capsici rely on chemical control, pesticide residues and other problems still threaten the pepper production, human health and environmental safety. Therefore, the search to explore important virulence factor and pathogenic mechanisms of P. capsici become the focus of the study.
The results show that in the plant pathogen and host interaction process, a diverse number of plant pathogenic microbes can manipulate biochemical, physiological, and morphological processes of their host plants through delivering a series of effector molecules that can either infect parasitic colonization or modulate plant defense responses. In 2003, Torto et al. discovered CRN1 and CRN2 from Pex (Phytophthora extracellular protein) cDNAs that triggered crinkling and necrosis of leaves (hence their moniker, crinklers or CRN proteins) when overexpressed in Nicotiana benthamiana transient expression assays used a vector derived from PVX (Potato virus X).
Currently, the CRN protein, as a new member in effector family, is only found in Phytophthora and there is few research on this protein family. In the interaction process of host and pathogen, the mechanism of CRN protein, the mechanism of enter cellular and the host target(s) and so on have known little, so it become a research hot spot.
In this study, the highly pathogenic strain of Phytophthora capsici SD33 saved in our lab was used as experimental material, and the work we did for the crinkling and necrosis genes was as follows:
(1) CRN-like sequences were collected from JGI database according to the conserved motifs reported and then blasted in NCBI database.
(2) Two CRN sequences, PcCRN1 and PcCRN2, were isolated from phytophthora capsici genome and analyzed using the NCBI online to confirm that both sequences were homologous to CRNs from nonredundant database.
(3) To examine the pathogenicity of PcCRN1 and PcCRN2 in P. capsici and the function of conserved motifs WL and LQLFLAK, a series of mutant constructs were generated and agrobacterium-mediated transient expression were used to assay in vitro. The PcCRN1 and PcCRN2 transient expression in Agrobacterium can produce chlorotic symptoms, and the symptoms of conserved motif-mutations before and after were different. This suggested that PcCRN1 and PcCRN2 play a role in pathogenicity and the conserved motif also had an impact on its pathogenicity.
(4) With pHam34 vector and PEG-mediated protoplast cotransformation techniques were implemented to produce transformants and RT-PCR and qRT-PCR technology were used to test and verify the CRN function in P. capsici.
Through this study, we can see the function of CRN in the pathogenesis of P. capsici, and we proved that PEG-mediated protoplast co-transformation technology can be implemented in P. capsici and it was determined that stable transformation successfully made a molecular tool to exploit the pathogenic mechanism of P. capsici.
引文
蔡艳,薛泉宏,陈占全,常显波,司美茹,孙小凤,来航线.青海高原东部土壤辣椒疫霉生防菌的初步筛选.西北农业学报,2007. 16: 241-244.
丁建成,戚仁德,顾江涛,黄立方,王贺胜,刘爱卿.辣椒疫病发生原因与防治对策.安徽农业科学,2001. 29:651-652.
贾菊生.新疆辣椒疫病及防治研究.植物病理学报,1992. 22: 257-262.
马辉刚.云南辣椒疫病菌种的鉴定.云南农业大学学报,1988. 3: 127-130.
欧雄常,柳凤,詹儒林,胡汉桥,何红,李信申,张小媛,赵艳龙.拮抗辣椒疫病菌的红树内生细菌筛选及RS261菌株鉴定.微生物学通报,2009. 36:175-180.
任光驰,马平虎,王延杰,张敬泽.甘肃辣椒疫病的发生与防治研究.植物保护,1990. 5: 16-17.
沈崇尧,王有琪,田林,沈克功,李延群.甘肃省辣椒疫病病原菌鉴定及生物学特性研究.云南农业大学学报,1990. 5: 72-77.
王志田,崔元瑜,杨华,何疆.新疆辣椒疫病菌鉴定及生物学特性研究.新疆农业科学,1993. 4:164-167.
许志刚.《普通植物病理学》第三版.中国农业出版社,2002.
易图永,谢丙炎,张宝玺,高必达.辣椒疫病防治研究进展.中国蔬菜,2002. 5: 52-55.
郑小波.疫霉菌及其研究技术.北京:中国农业出版社,1997. 10-13.
周启明,李林英,杨淑华.辣椒疫病的调查研究.中国蔬菜,1984. 1: 40-43.
Abramovitch RB, Martin GB. Strategies used by bacterial pathogens to suppress plant defenses. Curr Opin Plant Biol, 2004. 7: 356-364.
Allen RL, Bittner-Eddy PD, Grenville-Briggs LJ, Meitz JC, Rehmany AP, Rose LE, Beynon JL. Host-parasite coevolutionary conflict between Arabidopsis and downy mildew. Science, 2004. 306: 1957-1960.
Armstrong MR, Whisson SC, Pritchard L, Bos JI,Venter E, Avrova AO, Rehmany AP, Bo¨hme U, Brooks K, Cherevach I, Hamlin N, White B, FraserA, Lord A, Quail M A, Churcher C, Hall N, Be An ancestral oomycete locus contains late blight avirulence gene Avr3a, encoding a protein that is recognized in the host cytoplasm. Proc Natl Acad Sci USA, 2005. 102: 7766-7771.
Bailey BA, Jennings JC, Anderson JD. The 24-kDa protein from Fusarium oxysporum f.sp. erythroxyli: occurrence in related fungi and the effect of growth medium on its production. Can J Microbiol, 1997.43: 45-55.
Bendtsen JD, Nielsen H, von Heijne G, and Brunak S. Improved prediction of signal peptides: SignalP 3.0. J Mol Biol, 2004. 340: 783-795.
Birch PR, Boevink PC, Gilroy EM, Hein I, Pritchard L, and Whisson SC. Oomycete RXLR effectors: delivery, functional redundancy and durable disease resistance. Curr Opin Plant Biol, 2008. 11: 373-379.
Bishop JG, Ripoll DR, Bashir S, Damasceno CM, Seeds JD, Rose JK. Selection on Glycine beta-1, 3-endoglucanase genes differentially inhibited by a Phytophthora glucanase inhibitor protein. Genetics, 2004. 169: 1009-1019.
Bittner-Eddy PD, Allen RL, Rehmany AP, Birch P, Beynon JL. Use of suppression subtractive hybridization to identify downy mildew genes expressed during infection of Arabidopsis thaliana. Mol Plant Pathol, 2003.4: 501-507.
Bos JIB, Armstrong M, Whisson SC, Torto T, Ochwo M, Birch PRJ, Kamoun S. Intraspecific comparative genomics to identify avirulence genes from Phytophthora. New Phytol, 2003.159: 63-72.
Bos JI, Kanneganti TD, Young C, Cakir C, Huitema E, Win J, Armstrong M R, Birch P R, and Kamoun S. The C-terminal half of Phytophthora infestans RXLR effector AVR3a is sufficient to trigger R3a-mediated hypersensitivity and suppress INF1-induced cell death in Nicotiana benthamiana. Plant J, 2006. 48: 165-176.
Brunner F, Rosahl S, Lee J, Rudd JJ, Geiler C, Kauppinen S, Rasmussen G, Scheel D and Nurnberger T. Pep-13, a plant defense inducing pathogen-associated pattern from Phytophthora transglutaminases. EMBO J, 2002. 21: 81-88.
Damasceno CM, Bishop JG, Ripoll DR, Win J, Kamoun S, and Rose JK. Structure of the glucanase inhibitor protein (GIP) family from Phytophthora species suggests coevolution with plant endobeta- 1, 3-glucanases. Mol Plant Microbe Interact, 2008. 21: 820-830.
Dou D, Kale SD, Wang X, Chen Y, Wang Q, Wang X, Jiang RHY, Arredondo FD, Anderson R, Thakur P, McDowell JM, Wang YC, Tyler BM. Conserved C-terminal motifs required for avirulence and suppression of cell death by Phytophthora sojae effector Avr1b. Plant Cell, 2008.20: 1118-1133.
Dou D, Kale SD,Wang X, Jiang RHY, Bruce NA, Arredondo FD, Zhang X, Tyler BM. RXLR-mediated entry of Phytophthora sojae effector Avr1b into soybean cells does not require pathogen-encoded machinery. Plant Cell, 2008b. 20: 1930-1947.
Ellis JG, Rafiqi M, Gan P, Chakrabarti A, and Dodds PN. Recent progress in discovery and functional analysis of effector proteins of fungal and oomycete plant pathogens. Curr Opin Plant Biol, 2009. 4: 399-405.
Gaulin E, Jauneau A, Villalba F, Rickauer M, Esquerr′e-Tugay′e M-T, Bottin A. The CBEL glycoprotein of Phytophthora parasitica var. nicotianae is involved in cell wall deposition and adhesion to cellulosic substrates. J Cell Sci, 2002. 115: 4565-4575.
Hammond-Kosack KE, Jones JDG. Plant disease resistance genes. Annu Rev Plant Physiol Plant Mol Biol, 1997. 48: 575-607.
Haas BJ, Kamoun S, Zody MC, Jiang RH, Handsaker RE, Cano LM, Grabherr M, Kodira CD, Raffaele S, Torto-Alalibo T, Bozkurt TO, Ah-Fong AM, Alvarado L, Anderson VL, Armstrong MR, Avrova A, Baxter L, Beynon J, Boevink PC, Bollmann SR, Bos JI, Bulone V, Cai G, Cakir C, Carrington JC, Chawner M, Conti L, Costanzo S, Ewan R, Fahlgren N, Fischbach MA, Fugelstad J, Gilroy EM, Gnerre S, Green PJ, Grenville-Briggs LJ, Griffith J, Grunwald NJ, Horn K, Horner NR, Hu CH, Huitema E, Jeong DH, Jones AM, Jones JD, Jones RW, Karlsson EK, Kunjeti SG, Lamour K, Liu Z, Ma L, Maclean D, Chibucos MC, McDonald H, McWalters J, Meijer HJ, Morgan W, Morris PF, Munro CA, O’Neill K, Ospina-Giraldo M, Pinzon A, Pritchard L, Ramsahoye B, Ren Q, Restrepo S, Roy S, Sadanandom A, Savidor A, Schornack S, Schwartz DC, Schumann UD, Schwessinger B, Seyer C, Sharpe T, Silvar C, Song J, Studholme DJ, Sykes S, Thines M, van de Vondervoort PJ, Phuntumart V,Wawra S,Weide R, Win J, Young C, Zhou S, Fry W, Meyers BC, van West P, Ristaino J, Govers F, Birch P,Whisson SC, Judelson HS, and Nusbaum C. Genome sequence and comparative analysis of the Irish potato famine pathogen Phytophthora infestans. Nature, 2009. 461: 393-398. Hogenhout SA, Van der Hoorn RA, Terauchi R and Kamoun S. Emerging concepts in effector biology of plant-associated organisms. Mol Plant Microbe Interact, 2009. 22: 115-122.
Hord MJ, Ristaino J B. Effects of physical and chemical factors on the germination of oospores of Phytophthora capsici in vitro. Phytopathology, 1991. 81: 1541-1546. Huitema E, Bos JIB, Tian M, Win J, Waugh ME, Kamoun S. Linking sequence to phenotype in Phytophthora-plant interactions. Trends Microbiol, 2004. 12: 193-200.
Huitema E, Vleeshouwers VGAA, Cakir C, Kamoun S, Govers F. Differences in intensity and specificity of hypersensitive response induction in Nicotiana spp. by INF1, INF2A, and INF2B of Phytophthora infestans. Mol Plant Microbe Interact, 2005. 18: 183-193.
Hwang BK, de Cock AW AM, Bahnweg G, Prell HH, Heitefuse R. Restriction fragment length polymorphisms of mitochondrial DNA among Phytophthora capsici isolates from pepper(Capsicum annuum). Syst Appl Microbiol, 1991. 14: 111-116.
Jiang RH, Tripathy S, Govers F, and Tyler BM. RXLR effector reservoir in two Phytophthora species is dominated by a single rapidly evolving superfamily with more than 700 members. Proc Natl Acad Sci USA, 2008. 105: 4874-4879.
Judelson HS, Tyler BM, and Michelmore RW. Transformation of the oomycete pathogen, Phytophthora infestans. Mol Plant Microbe Interact, 1991. 4: 602-607.
Kamoun S. Molecular genetics of pathogenic oomycetes. Eukaryot Cell, 2003. 2: 191-199.
Kamoun S. A catalogue of the effector secretome of plant pathogenic oomycetes. Annu Rev Phytopathol, 2006. 44: 41-60.
Kamoun S. Groovy times: filamentous pathogen effectors revealed. Curr Opin in Plant Biol, 2007.10: 358-365.
Kamoun S, Honee G,Weide R, Lauge R, Kooman-Gersmann M, De Groot K, Govers F, De Wit PJGM. The fungal gene Avr9 and the oomycete gene inf1 confer avirulence to potato virus X on tobacco. Mol Plant Microbe Interact, 1999.12: 459-462.
Kamoun S, Lindqvist H, Govers F. A novel class of elicitin-like genes from Phytophthora infestans. Mol Plant Microbe Interact, 1997.10: 1028-1030.
Kamoun S, Hraber P, Sobral B, Nuss D, Govers F. Initial assessement of gene diversity for the oomycete pathogen Phytophthora infestans based on expressed sequences. Fungal Genet Biol, 1999. 28: 94-106.
Kamoun S, Smart CD. Late blight of potato and tomato in the genomics era. Plant Dis, 2005.89: 692-699.
Kamoun S, van West P, de Jong AJ, de Groot K, Vleeshouwers V, Govers F. A gene encoding a protein elicitor of Phytophthora infestans is down-regulated during infection of potato. Mol Plant Microbe Interact, 1997. 10: 13-20.
Kamoun S, van West P, Vleeshouwers VG, de Groot KE, Govers F. Resistance of Nicotiana benthamiana to Phytophthora infestans is mediated by the recognition of the elicitor protein INF1. Plant Cell, 1998. 10: 1413-1426.
Kamoun S, Young M, Glascock C, Tyler BM. Extracellular protein elicitors from Phytophthora: host-specificity and induction of resistance to fungal and bacterial phytopathogens.Mol Plant Microbe Interact, 1993. 6: 15-25.
Kamoun S, van West P, Vleeshouwers VG, de Groot KE, and Govers, F. Resistance of Nicotiana benthamiana to Phytophthora infestans is mediated by the recognition of the elicitor protein INF1. Plant Cell, 1998. 10: 1413-1426.
Kanneganti TD, Huitema E, Cakir C, and Kamoun S. Synergistic interactions of the plant cell death pathways induced by Phytophthora infestans Nep1-like protein PiNPP1.1 and INF1 elicitin. Mol Plant Microbe Interact, 2006. 19: 854-863.
Kim ES, Hwang K. Virulence to Korea pepper cuhivars of Photophthora capsici from different geographic areas. Plant Dis, 1992. 76: 486-489.
Ko WH. Hormonal heterothallism and homothallism in Phytophthora. Ann Rev Phytopathol, 1988. 26: 57-73.
Leonian LH. Stem and fruit blight of peppers caused by Phytophthora capsici. Phytopathology, 1922. 12: 401-408.
Liu Z, Bos JIB, Armstrong M, Whisson SC, da Cunha LD, Torto-Alalibo T, Win J, Avrova AO, Wright F, Birch PRJ and Kamoun S. Patterns of diversifying selection in the phytotoxin-like scr74 gene family of Phytophthora infestans. Mol Biol Evol, 2005. 22: 659-672.
Liu T, Ye W, Ru Y, Yang X, Gu B, Tao K, Lu S, Dong S, Zheng X, Shan W, Wang Y, and Dou D. Two host cytoplasmic effectors are required for pathogenesis of Phytophthorasojae by suppression of host defenses. Plant Physiol, 2010. 155: 450-501.
McLeod A, Fry BA, Zuluaga-Duque AP, Meyers KL, and Fry WE. Toward improvements of oomycete transformation protocols. J Eukaryot Microbiol, 2008. 55: 103-109.
Miki D, Itoh R and Shimamoto K. RNA silencing of single and multiple members in a gene family of rice. Plant Physiol, 2005. 138: 1903-1913.
Morgan, W. and Kamoun, S. RXLR effectors of plant pathogenic oomycetes. Curr Opin Microbiol, 2007. 10: 332-338.
Nurnberger T, Nennstiel D, Jabs T, Sacks WR, Hahlbrock K, Scheel D. High affinity binding of a fungal oligopeptide elicitor to parsley plasma membranes triggers multiple defense responses. Cell, 1994. 78: 449-460.
Oh SK, Young C, Lee M, Oliva R, Bozkurt T, Cano LM, Win J, Bos JIB, Liu H-Y, van Damme M, Morgan W, Choi D, van der Vossen EAG, Vleeshouwers V, and Kamoun S. In planta expression screens of Phytophthora infestans RXLR effectors reveal diverse phenotypes including activation of the Solanum bulbocastanum disease resistance protein Rpi-blb2. Plant Cell, 2009. 21: 2928-2947.
Orsomando G, Lorenzi M, Ferrari E, de Chiara C, Spisni A, Ruggieri S. PcF protein from Phytophthora cactorum and its recombinant homologue elicit phenylalanine ammonia lyase activation in tomato. Cell Mol Life Sci, 2003. 60: 1470-1476.
Oliva R, Win J, Raffaele S, Boutemy L, Bozkurt TO, Chaparro-Garcia A, Segretin ME, Stam R, Schornack S, Cano LM, Damme M van, Huitema E, Thines M, Banfield M J, Kamoun S. Recent developments in effector biology of filamentous plant pathogens. Cell Micro, 2010. 12: 705-715.
Pemberton CL, Salmond GPC. The Nep1-like proteins a growing family of microbial elicitors of plant necrosis. Mol Plant Pathol, 2004. 5: 353-359.
Pemberton CL, Whitehead NA, Sebaihia M, Bell KS, Hyman LJ, et al. Novel quorum-sensing-controlled genes in Erwinia carotovora subsp. carotovora: identification of a fungal elicitor homologue in a soft-rotting bacterium. Mol Plant Microbe Interact, 2005. 18: 343-353.
Polach FJ, Webster PK. Identification of strains and inheritance of pathogenicity in Phytophthora capsici. Phytopathology, 1992. 62: 20-26.
Ponchet M, Panabieres F, Milat M-L, Mikes V, Montillet J-L, Suty L, Triantaphylides C, Tirilly Y, Blein JP. Are elicitins cryptograms in plant-Oomycete communications? Cell Mol Life Sci, 1999. 56: 1020-1047.
Punja ZK. Fungal Disease Resistance in Plants. Binghamton, NY: Food Products Press, 2004. 266.
Qutob D, Kamoun S, Gijzen M. Expression of a Phytophthora sojae necrosis-inducing protein occurs during transition from biotrophy to necrotrophy. Plant J, 2002. 32:361-373.
Qutob D, Tedman-Jones J, Dong S, Kuflu K, Pham H, Wang Y, Dou D, Kale SD, Arredondo FD, Tyler BM, and Gijzen M. Copy number variation and transcriptional polymorphisms of Phytophthora sojae RXLR effector genes Avr1a and Avr3a. PLoS ONE, 2009. 4: 5066.
Rawlings ND, Tolle DP, Barrett AJ. MEROPS: the peptidase database. Nucleic Acids Res, 2004. 32: 160-164.
Rehmany AP, Gordon A, Rose LE, Allen RL, Armstrong MR, Whisson SC, Kamoun S, Tyler BM, Birch PRJ, and Beynon JM. Differential recognition of highly divergent downy mildew avirulence gene alleles by RPP1 resistance genes from two Arabidopsis lines. Plant Cell, 2005. 17: 1839-1850.
Ricci P, Bonnet P, Huet J-C, Sallantin M, Beauvais-Cante F, Bruneteau M, Billard V, Michel G, Pernollet JC. Structure and activity of proteins from pathogenic fungi Phytophthora eliciting necrosis and acquired resistance in tobacco. Eur J Biochem, 1989.183: 555-563.
Ristaino, JB. Intraspecific variation among isolates of Phytophthora capsici from pepper and cucurbit fields in North Carolina. Phytopathology, 1990. 80: 1253-1259.
Rose JK, Ham KS, Darvill AG, Albersheim P. Molecular cloning and characterization of glucanase inhibitor proteins: coevolution of a counterdefense mechanism by plant pathogens. Plant Cell, 2002.14: 1329-1345.
Rose LE, Bittner-Eddy PD, Langley CH, Holub EB, Michelmore RW, Beynon JL. The maintenance of extreme amino acid diversity at the disease resistance gene, RPP13, in Arabidopsis thaliana. Genetics, 2004.166: 1517-1527.
Schornack S, van Damme M, Bozkurta O T, Canoa M L, Smokera M, Thines M, Gaulinc E, Kamoun S, and Huitema E. Ancient class of translocated oomycete effectors targets the host nucleus. Proc Natl Acad Sci, 2010. 107: 17421-17426.
Song J, Win J, Tian M, Schornack S, Kaschani F, Ilyas M, van der Hoorn RA, and Kamoun S. Apoplastic effectors secreted by two unrelated eukaryotic plant pathogens target the tomato defense protease Rcr3. Proc Natl Acad Sci USA, 2009. 106: 1654-1659.
Sun WX, Jia YJ, O’Neil NR, Feng BZH, Zhang XG. Gnentic diversity in Phytophthora capsici from eastern china. Can J Plant pathol, 2008. 30: 414-424.
Sun WX, Jia YJ, Feng BZ, O’Neill NR, Zhu XP, Xie BY, and Zhang XG. Functional analysis of Pcipg2 from the straminopilous plant pathogen Phytophthora capsici. Genesis, 2009. 47: 535-544.
Sacks W, Nurnberger T, Hahlbrock K, Scheel D. Molecular characterization of nucleotide sequences encoding the extracellular glycoprotein elicitor from Phytophthora megasperma. Mol Gen Genet, 1995. 246: 45-55.
Sasabe M, Takeuchi K, Kamoun S, Ichinose Y, Govers F, Toyoda K. Independent pathways leading to apoptotic cell death, oxidative burst and defense gene expression in response toelicitin in tobacco cell suspension culture. Eur J Biochem, 2000. 267: 5005-5013.
Sejalon-Delmas N, Villalba MF, Bottin A, Rickauer M, Dargent R, Esquerr′e-Tugay′e MT. Purification, elicitor activity, and cell wall localization of a glycoprotein from Phytophthora parasitica var. nicotianae, a fungal pathogen of tobacco. Phytopathology, 1997. 87: 899-909.
Shan W, Cao M, Leung D, Tyler BM. The Avr1b locus of Phytophthora sojae encodes an elicitor and a regulator required for avirulence on soybean plants carrying resistance gene Rps1b. Mol Plant Microbe Interact, 2004.17: 394-403.
Sun WX, Jia YJ, O’Neill NR, Feng BZ, Zhang XG.Genetic diversity in Phytophthora capsici from eastern China. Can J Plant Pathol, 2008.30: 414-424.
Tian M, Benedetti B, Kamoun S. A second Kazal-like protease inhibitor from Phytophthora infestans inhibits and interacts with the apoplastic pathogenesis-related protease P69B of tomato. Plant Physiol, 2005. 138: 1785-1793.
Tian M, Huitema E, da Cunha L, Torto-Alalibo T, Kamoun S. A Kazallike extracellular serine protease inhibitor from Phytophthora infestans targets the tomato pathogenesis-related protease P69B. J Biol Chem, 2004. 279: 26370-26377.
Tian M, Kamoun S. A two disulfide bridge Kazal domain from Phytophthora exhibits stable inhibitory activity against serine proteases of the subtilisin family. BMC Biochem, 2005. 6: 15.
Tian M, Win J, Song J, van der Hoorn R, van der Knaap E. and Kamoun S. A Phytophthora infestans cystatin-like protein targets a novel tomato papain-like apoplastic protease. Plant Physiol, 2007.143: 364-377.
Torto TA, Li SH, Styer A, Huitema E, Testa A, Gow NAR, van West P, and Kamoun S. EST mining and functional expression assays identify extracellular effector proteins from the plant pathogen phytophthora. Genome Res, 2003.13: 1675-1685.
Tyler BM. Entering and breaking: virulence effector proteins of oomycete plant pathogens. Cell Microbiol, 2009. 11: 13-20.
Tyler BM, Tripathy S, Zhang X, Dehal P, Jiang RH, Aerts A, Arredondo FD, Baxter L, Bensasson D, Beynon JL, Chapman J, Damasceno CM, Dorrance AE, Dou D, Dickerman AW, Dubchak IL, Garbelotto M, Gijzen M, Gordon SG, Govers F, Grunwald NJ, Huang W, Ivors KL, Jones RW, Kamoun S, Krampis K, Lamour KH, Lee MK, McDonald WH, Medina M, Meijer HJ, Nordberg EK, Maclean DJ, Ospina-Giraldo MD, Morris PF, Phuntumart V, Putnam NH, Rash S, Rose JK, Sakihama Y, Salamov AA, Savidor A, Scheuring CF, Smith BM, Sobral BW, Terry A, Torto-Alalibo TA, Win J, Xu Z, Zhang H, Grigoriev IV, Rokhsar DS, and Boore JL. Phytophthora genome sequences uncover evolutionary origins and mechanisms of pathogenesis. Science, 2006. 313: 1261-1266.
van der Hoorn RAL, Laurent F, Roth R, and De Wit PJGM. Agroinfiltration is a versatile tool that facilitates comparative analyses of Avr9/Cf-9-induced and Avr4/Cf-4-inducednecrosis. Mol Plant Microbe Interact, 2000. 16: 669-680.
van West P, Kamoun S, van’T Klooster JW, and Govers F. Internuclear gene silencing in Phytophthora infestans. 1999. Mol Cell, 3: 339-348.
Villalba Mateos F, Rickauer M, Esquerre-Tugaye MT. Cloning and characterization of a cDNA encoding an elicitor of Phytophthora parasitica var. nicotianae that shows cellulose-binding and lectin-like activities. Mol Plant Microbe Interact, 1997. 10: 1045-1053.
Vleeshouwers VGAA, Driesprong J-D, Kamphuis LG, Torto-Alalibo T, van‘t Slot, KAE, Govers F, Visser RGF, Jacobsen E, and Kamoun S. Agroinfection-based high throughput screening reveals specific recognition of INF elicitins in Solanum. Mol Plant Pathol, 2006. 7:499-510.
Whisson SC, Boevink PC, Moleleki L, Avrova AO, Morales JG, Gilroy EM, Armstrong MR, Grouffaud S, van West P, Chapman S, Hein I, Toth IK, Pritchard L, and Birch PR. A translocation signal for delivery of oomycete effector proteins into host plant cells. Nature, 2007. 450: 115-118.
Wu CH, Yan HZ, Liu LF, Liou RF. Functional characterization of a gene family encoding polygalacturonases in Phytophthora parasitica. Mol Plant Microbe Interact, 2008. 21: 480-489.
Win J, Kanneganti TD, Torto-Alalibo T, Kamoun S. Computational and comparative analyses of 150 full-length cDNA sequences from the oomycete plant pathogen Phytophthora infestans. Fungal Genet Biol, 2006. 43: 20-33.
Win J, Morgan W, Bos J, Krasileva KV, Cano LM, Chaparro-Garcia A, Ammar R, Staskawicz BJ, and Kamoun S. Adaptive evolution has targeted the C-terminal domain of the RXLR effectors of plant pathogenic oomycetes. Plant Cell, 2007.19: 2349-2369.
Xiang T, Zong N, Zhang J, Chen J, Chen M, Zhou J. BAK1 Is Not a Target of the Pseudomonas syringae Effector AvrPto. Mol Plant Microbe Interact, 2011. 24: 100-107.
Xing W, Zou Y, Liu Q, Liu J, Luo X, Huang Q, Chen S, Zhu L, Bi R, Hao Q, Wu JW, Zhou JM & Chai J. The structural basis for activation of plant immunity by bacterial effector protein AvrPto. Nature, 2007. 449: 243-247.