大丽轮枝菌黑色素合成相关基因与微菌核形成的关系
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摘要
大丽轮枝菌是一种重要的土传植物病原真菌,以休眠结构微菌核作为初始接种体,可侵染660多种植物引致黄萎病。微菌核是致密的多细胞结构,表面附着大量的DHN黑色素。许多报道指出,在微菌核发育过程中,传统的DHN黑色素合成途径中有5种催化酶编码基因Vd PKS、Vd T4HR、Vd SCD、Vd T3HR和Vd LAC均被诱导表达,但这些基因与微菌核形成的关系目前尚无报道。本研究通过基因敲除技术,系统研究了传统DHN黑色素合成通路上这5种关键酶编码基因及一种缩链催化酶编码基因Vayg1在大丽轮枝菌黑色素合成及微菌核形成中的作用。结果表明,大丽轮枝菌DHN黑色素合成需要Vayg1基因的参与,且Vayg1和Vd T3HR基因还参与微菌核的形成过程。因此,Vayg1基因和Vd T3HR基因可作为黄萎病防治的新靶标。
The soil-born plant fungal pathogen Verticillium dahliae, with its resting structure of microsclerotia as primary inoculum, can cause Verticillium wilt on more than 660 plant species. The well-developed microsclerotia are thick-walled structures, where DHN melanin particles are deposited. Many studies suggest that expressions of five genes, Vd PKS, VdT4HR, Vd SCD, VdT3HR and Vd LAC, encoding five enzymes involved in DHN melanin biosynthesis were induced during microsclerotium development. However, their role in microsclerotium formation remains unclear. In the current study, melanin deficient mutants generated by deletion of these five genes and the Vayg1 gene, which is responsible for chain-length shortening of a precursor,were used to systemically elucidate their role on melanin production and microsclerotium formation in V. dahliae. The results showed that Vayg1 is required for DHN melanin production in V. dahliae. Vayg1 and Vd T3 HR are involved in microsclerotium formation, indicating that these two enzymes would be potential targets for Verticillium wilt management.
The soil-born plant fungal pathogen Verticillium dahliae, with its resting structure of microsclerotia as primary inoculum, can cause Verticillium wilt on more than 660 plant species. The well-developed microsclerotia are thick-walled structures, where DHN melanin particles are deposited. Many studies suggest that expressions of five genes, Vd PKS, VdT4HR, Vd SCD, VdT3HR and Vd LAC, encoding five enzymes involved in DHN melanin biosynthesis were induced during microsclerotium development. However, their role in microsclerotium formation remains unclear. In the current study, melanin deficient mutants generated by deletion of these five genes and the Vayg1 gene, which is responsible for chain-length shortening of a precursor,were used to systemically elucidate their role on melanin production and microsclerotium formation in V. dahliae. The results showed that Vayg1 is required for DHN melanin production in V. dahliae. Vayg1 and Vd T3 HR are involved in microsclerotium formation, indicating that these two enzymes would be potential targets for Verticillium wilt management.
引文
Bayry J,Beaussart A,Dufrêne YF,Sharma M,Bansal K,Kniemeyer O,Aimanianda V,Brakhage AA,Kaveri SV,Kwon-Chung KJ,LatgéJP,Beauvais A,2014.Surface structure characterization of Aspergillus fumigatus conidia mutated in the melanin synthesis pathway and their human cellular immune response.Infection and Immunity,82(8):3141-3153
Beauvais A,Fontaine T,Aimanianda V,LatgéJP,2014.Aspergillus cell wall and biofilm.Mycopathologia,178:371-377
Bell AA,Puhalla JE,Tolmsoff WJ,Stipanovic RD,1976.Use of mutants to establish(+)-scytalone as an intermediate in melanin biosynthesis.Canadian Journal of Microbiology,22:787-799
Bell AA,Wheeler MH,1986.Biosynthesis and functions of fungal melanins.Annual Review of Phytopathology,24(1):411-451
Brandt WH,1964.Morphogenesis in Verticillium:effects of light and ultraviolet radiation on microsclerotia and melanin.Botany,42(8):1017-1023
Duressa D,Anchieta A,Chen DQ,Klims A,Garcia-Pedrajas MD,Dobinson KF,Klosterman SJ,2013.RNA-seq analyses of gene expression in the microsclerotia of Verticillium dahliae.BMC Genomics,14(1):607
Eisenman HC,Casadevall A,2012.Synthesis and assembly of fungal melanin.Applied Microbiology and Biotechnology,93(3):931-940
Fan R,Klosterman SJ,Wang C,Subbarao KV,Xu X,Shang W,Hu X,2017.Vayg1 is required for microsclerotium formation and melanin production in Verticillium dahliae.Fungal Genetics and Biology,98:1-11
Fradin EF,Thomma BPHJ,2006.Physiology and molecular aspects of Verticillium wilt diseases caused by V.dahliae and V.albo-atrum.Molecular Plant Pathology,7(2):71-86
Fujii I,Mori Y,Watanabe A,Kubo Y,Tsuji G,Ebizuka Y,2000.Enzymatic synthesis of 1,3,6,8-tetrahydroxynaphthalene solely from malonyl coenzyme A by a fungal iterative type I polyketide synthase PKS1.Biochemistry,39(30):8853-8858
Fujii I,Yasuoka Y,Tsai HF,Chang YC,Kwon-Chung KJ,Ebizuka Y,2004.Hydrolytic polyketide shortening by Ayg1p,a novel enzyme involved in fungal melanin biosynthesis.Journal of Biological Chemistry,279(43):44613-44620
Green RJ,1981.An overview in fungal wilt diseases of plants.Academic Press,New York.1-20
Jian GL,Zou YF,Ma C,2003.Epidemiology and management of Verticillium wilt on cotton.Chinese Cotton,30(3):13-14(in Chinese)
Klimes A,Amyotte SG,Grant S,Kang S,Dobinson KF,2008.Microsclerotia development in Verticillium dahliae:regulation and differential expression of the hydrophobin gene VDH1.Fungal Genetics and Biology,45(12):1525-1532
Klimes A,Dobinson KF,2006.A hydrophobin gene,VDH1,is involved in microsclerotial development and spore viability in the plant pathogen Verticillium dahliae.Fungal Genetics and Biology,43(4):283-294
Klosterman SJ,Atallah ZK,Vallad GE,Subbarao KV,2009.Diversity,pathogenicity,and management of Verticillium species.Annual Review of Phytopathology,47(1):39-62
Liu XT,Song XX,Guo JC,1998.Studies and advances on cotton Verticillium wilt.Cotton Science,10(1):6-13(in Chinese)
Nosanchuk JD,Casadevall A,2003.The contribution of melanin to microbial pathogenesis.Cellular Microbiology,5(4):203-223
Paz Z,García-Pedrajas MD,Andrews DL,Klosterman SJ,Baeza-Monta?ez L,Gold SE,2011.One step construction of Agrobacterium-recombination-ready-plasmids(OSCAR),an efficient and robust tool for ATMT based gene deletion construction in fungi.Fungal Genetics and Biology,48(7):677-684
Pegg GF,Brady BL,2002.Verticillium wilts.CABI Publishing,Wallingford.57-60
Polacheck I,Rosenberger RF,1977.Aspergillus nidulans mutant lacking alpha-(1,3)-glucan,melanin,and cleistothecia.Journal of Bacteriology,132(2):650-656
Subbarao KV,Hubbard JC,Greathead AS,Spencer GA,1997.Verticillium wilt.The American Phytopathological Society(APS)Press,St Paul,Minnesota.26-27
Tsai HF,Fujii I,Watanabe A,Wheeler MH,Chang YC,Yasuoka Y,Ebizuka T,Kwon-Chung KJ,2001.Pentaketide melanin biosynthesis in Aspergillus fumigatus requires chain-length shortening of a heptaketide precursor.Journal of Biological Chemistry,276(31):29292-29298
Wei F,Fan R,Dong H,Shang W,Xu X,Zhu H,Yang J,Hu X,2015.Threshold microsclerotial inoculum for cotton Verticillium wilt determined through wet-sieving and real-time quantitative PCR.Phytopathology,105(2):220-229
Wheeler MH,Abramczyk D,Puckhaber LS,Naruse M,Ebizuka Y,Fujii I,Szaniszlo PJ,2008.New biosynthetic step in the melanin pathway of Wangiella(Exophiala)dermatitidis:evidence for 2-acetyl-1,3,6,8-tetrahydroxynaphthalene as a novel precursor.Eukaryotic Cell,7(10):1699-1711
Xiong D,Wang Y,Ma J,Klosterman SJ,Xiao S,T ian C,2014.Deep m RNA sequencing reveals stage-specific transcriptome alterations during microsclerotia development in the smoke tree vascular wilt pathogen,Verticillium dahliae.BMC Genomics,15(1):324
Xu L,Zhu LF,Zhang XL,2012.Research on resistance mechanism of cotton to Verticillium wilt.Acta Agronomica Sinica,38(9):1553-1560(in Chinese)
Zhang CL,2004.Occurrence and management of wilt disease on potato.Plant Doctor,17(5):6(in Chinese)
Zhang JW,2009.Occurrence and management of wilt disease on eggplant.Jilin Vegetables,3:62-63(in Chinese)
Zhu HQ,Feng ZL,Yin ZX,Li ZF,Zhao LH,Shi YQ,Yang JR,2012.Pathogenicity differentiation and ISSR fingerprint analysis of cotton Verticillium dahliae in China.Acta Phytopathologica Sinica,42(3):225-235(in Chinese)
简桂良,邹亚飞,马存,2003.棉花黄萎病连年流行的原因及对策.中国棉花,30(3):13-14
刘学堂,宋晓轩,郭金城,1998.棉花黄萎病菌的研究及最新进展.棉花学报,10(1):6-13
徐理,朱龙付,张献龙,2012.棉花抗黄萎病机制研究进展.作物学报,38(9):1553-1560
张成礼,2004.马铃薯黄萎病的发生与防治.植物医生,17(5):6
张继伟,2009.茄子黄萎病发生及综合防治技术.吉林蔬菜,3:62-63
朱荷琴,冯自力,尹志新,李志芳,赵丽红,师勇强,杨家荣,2012.我国棉花黄萎病菌致病力分化及ISSR指纹分析.植物病理学报,42(3):225-235
Beauvais A,Fontaine T,Aimanianda V,LatgéJP,2014.Aspergillus cell wall and biofilm.Mycopathologia,178:371-377
Bell AA,Puhalla JE,Tolmsoff WJ,Stipanovic RD,1976.Use of mutants to establish(+)-scytalone as an intermediate in melanin biosynthesis.Canadian Journal of Microbiology,22:787-799
Bell AA,Wheeler MH,1986.Biosynthesis and functions of fungal melanins.Annual Review of Phytopathology,24(1):411-451
Brandt WH,1964.Morphogenesis in Verticillium:effects of light and ultraviolet radiation on microsclerotia and melanin.Botany,42(8):1017-1023
Duressa D,Anchieta A,Chen DQ,Klims A,Garcia-Pedrajas MD,Dobinson KF,Klosterman SJ,2013.RNA-seq analyses of gene expression in the microsclerotia of Verticillium dahliae.BMC Genomics,14(1):607
Eisenman HC,Casadevall A,2012.Synthesis and assembly of fungal melanin.Applied Microbiology and Biotechnology,93(3):931-940
Fan R,Klosterman SJ,Wang C,Subbarao KV,Xu X,Shang W,Hu X,2017.Vayg1 is required for microsclerotium formation and melanin production in Verticillium dahliae.Fungal Genetics and Biology,98:1-11
Fradin EF,Thomma BPHJ,2006.Physiology and molecular aspects of Verticillium wilt diseases caused by V.dahliae and V.albo-atrum.Molecular Plant Pathology,7(2):71-86
Fujii I,Mori Y,Watanabe A,Kubo Y,Tsuji G,Ebizuka Y,2000.Enzymatic synthesis of 1,3,6,8-tetrahydroxynaphthalene solely from malonyl coenzyme A by a fungal iterative type I polyketide synthase PKS1.Biochemistry,39(30):8853-8858
Fujii I,Yasuoka Y,Tsai HF,Chang YC,Kwon-Chung KJ,Ebizuka Y,2004.Hydrolytic polyketide shortening by Ayg1p,a novel enzyme involved in fungal melanin biosynthesis.Journal of Biological Chemistry,279(43):44613-44620
Green RJ,1981.An overview in fungal wilt diseases of plants.Academic Press,New York.1-20
Jian GL,Zou YF,Ma C,2003.Epidemiology and management of Verticillium wilt on cotton.Chinese Cotton,30(3):13-14(in Chinese)
Klimes A,Amyotte SG,Grant S,Kang S,Dobinson KF,2008.Microsclerotia development in Verticillium dahliae:regulation and differential expression of the hydrophobin gene VDH1.Fungal Genetics and Biology,45(12):1525-1532
Klimes A,Dobinson KF,2006.A hydrophobin gene,VDH1,is involved in microsclerotial development and spore viability in the plant pathogen Verticillium dahliae.Fungal Genetics and Biology,43(4):283-294
Klosterman SJ,Atallah ZK,Vallad GE,Subbarao KV,2009.Diversity,pathogenicity,and management of Verticillium species.Annual Review of Phytopathology,47(1):39-62
Liu XT,Song XX,Guo JC,1998.Studies and advances on cotton Verticillium wilt.Cotton Science,10(1):6-13(in Chinese)
Nosanchuk JD,Casadevall A,2003.The contribution of melanin to microbial pathogenesis.Cellular Microbiology,5(4):203-223
Paz Z,García-Pedrajas MD,Andrews DL,Klosterman SJ,Baeza-Monta?ez L,Gold SE,2011.One step construction of Agrobacterium-recombination-ready-plasmids(OSCAR),an efficient and robust tool for ATMT based gene deletion construction in fungi.Fungal Genetics and Biology,48(7):677-684
Pegg GF,Brady BL,2002.Verticillium wilts.CABI Publishing,Wallingford.57-60
Polacheck I,Rosenberger RF,1977.Aspergillus nidulans mutant lacking alpha-(1,3)-glucan,melanin,and cleistothecia.Journal of Bacteriology,132(2):650-656
Subbarao KV,Hubbard JC,Greathead AS,Spencer GA,1997.Verticillium wilt.The American Phytopathological Society(APS)Press,St Paul,Minnesota.26-27
Tsai HF,Fujii I,Watanabe A,Wheeler MH,Chang YC,Yasuoka Y,Ebizuka T,Kwon-Chung KJ,2001.Pentaketide melanin biosynthesis in Aspergillus fumigatus requires chain-length shortening of a heptaketide precursor.Journal of Biological Chemistry,276(31):29292-29298
Wei F,Fan R,Dong H,Shang W,Xu X,Zhu H,Yang J,Hu X,2015.Threshold microsclerotial inoculum for cotton Verticillium wilt determined through wet-sieving and real-time quantitative PCR.Phytopathology,105(2):220-229
Wheeler MH,Abramczyk D,Puckhaber LS,Naruse M,Ebizuka Y,Fujii I,Szaniszlo PJ,2008.New biosynthetic step in the melanin pathway of Wangiella(Exophiala)dermatitidis:evidence for 2-acetyl-1,3,6,8-tetrahydroxynaphthalene as a novel precursor.Eukaryotic Cell,7(10):1699-1711
Xiong D,Wang Y,Ma J,Klosterman SJ,Xiao S,T ian C,2014.Deep m RNA sequencing reveals stage-specific transcriptome alterations during microsclerotia development in the smoke tree vascular wilt pathogen,Verticillium dahliae.BMC Genomics,15(1):324
Xu L,Zhu LF,Zhang XL,2012.Research on resistance mechanism of cotton to Verticillium wilt.Acta Agronomica Sinica,38(9):1553-1560(in Chinese)
Zhang CL,2004.Occurrence and management of wilt disease on potato.Plant Doctor,17(5):6(in Chinese)
Zhang JW,2009.Occurrence and management of wilt disease on eggplant.Jilin Vegetables,3:62-63(in Chinese)
Zhu HQ,Feng ZL,Yin ZX,Li ZF,Zhao LH,Shi YQ,Yang JR,2012.Pathogenicity differentiation and ISSR fingerprint analysis of cotton Verticillium dahliae in China.Acta Phytopathologica Sinica,42(3):225-235(in Chinese)
简桂良,邹亚飞,马存,2003.棉花黄萎病连年流行的原因及对策.中国棉花,30(3):13-14
刘学堂,宋晓轩,郭金城,1998.棉花黄萎病菌的研究及最新进展.棉花学报,10(1):6-13
徐理,朱龙付,张献龙,2012.棉花抗黄萎病机制研究进展.作物学报,38(9):1553-1560
张成礼,2004.马铃薯黄萎病的发生与防治.植物医生,17(5):6
张继伟,2009.茄子黄萎病发生及综合防治技术.吉林蔬菜,3:62-63
朱荷琴,冯自力,尹志新,李志芳,赵丽红,师勇强,杨家荣,2012.我国棉花黄萎病菌致病力分化及ISSR指纹分析.植物病理学报,42(3):225-235