Large-scale gene disruption in Magnaporthe oryzae identifies MC69, a secreted protein required for infection by monocot and dicot fungal pathogens.
Large-scale gene disruption in Magnaporthe oryzae identifies MC69, a secreted protein required for infection by monocot and dicot fungal pathogens.
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DOI：
10.1371/journal.ppat.1002711
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发表时间：
2012
期刊：
影响因子：
6.7
通讯作者：
Terauchi R
中科院分区：
文献类型：
--
作者：
Saitoh H;Fujisawa S;Mitsuoka C;Ito A;Hirabuchi A;Ikeda K;Irieda H;Yoshino K;Yoshida K;Matsumura H;Tosa Y;Win J;Kamoun S;Takano Y;Terauchi R
为了寻找稻瘟病菌的毒力效应基因，我们对稻瘟病菌侵染早期表达的78个可能的分泌蛋白的基因进行了大规模的靶向干扰。大多数基因的中断不会影响米曲霉的生长、分生孢子或致病性。一个例外是MC69基因。Mc69突变体在水稻和大麦上表现出严重的稻瘟病症状减轻，表明mc69对稻瘟病菌的致病力具有重要作用。Mc69突变体没有表现出腐生生长和分生孢子的变化。对mc69突变体的侵染行为的显微分析表明，mc69对于附着胞的形成是必不可少的。然而，mc69突变体在水稻叶鞘附着胞形成后没有形成侵染菌丝，这表明mc69在与寄主植物的互作中起着关键作用。MC69编码一个由54个氨基酸组成的含有信号肽的蛋白质。活细胞成像表明，荧光标记的MC69没有转移到水稻细胞质中。在成熟的MC69中对两个保守的半胱氨酸残基(Cys36和Cys46)进行定点突变，可在不影响其分泌的情况下损害MC69的功能，提示二硫键在MC69致病功能中的重要性。此外，MC69同源基因的缺失降低了黄瓜炭疽病菌对黄瓜和烟草叶片的致病力。我们的结论是，MC69是一种分泌型致病蛋白，通常是两种不同的植物病原真菌侵染单子叶植物和双子叶植物时所必需的。稻瘟病菌是水稻中最具破坏性的真菌病害。米曲霉分泌大量的效应蛋白，其中包括几种已知可被激活天然免疫的寄主抗性蛋白识别的无毒蛋白。然而，到目前为止，除了最近发现的一种效应蛋白--分泌型LysM蛋白1(SLp1)外，在水稻分枝杆菌中还没有鉴定出毒力活动所需的效应物。我们对米曲霉的候选效应进行了大规模的干扰分析，发现了一种由真菌在感染过程中分泌的小蛋白MC69。当缺乏MC69时，穿透宿主细胞后的致病性严重降低。此外，炭疽菌中MC69基因的缺失降低了其在寄主植物黄瓜和烟草中的致病力。因此，MC69在子囊菌中是保守的，对建立亲和性是至关重要的。这是首次报道在单子叶和双子叶病原真菌中都是致病所必需的单一分泌蛋白。MC69如何对致病性或致病力起作用尚不清楚，但它可能是真菌成为病原体所必需的，也可能是作用于植物目标分子的经典效应器。
To search for virulence effector genes of the rice blast fungus, Magnaporthe oryzae, we carried out a large-scale targeted disruption of genes for 78 putative secreted proteins that are expressed during the early stages of infection of M. oryzae. Disruption of the majority of genes did not affect growth, conidiation, or pathogenicity of M. oryzae. One exception was the gene MC69. The mc69 mutant showed a severe reduction in blast symptoms on rice and barley, indicating the importance of MC69 for pathogenicity of M. oryzae. The mc69 mutant did not exhibit changes in saprophytic growth and conidiation. Microscopic analysis of infection behavior in the mc69 mutant revealed that MC69 is dispensable for appressorium formation. However, mc69 mutant failed to develop invasive hyphae after appressorium formation in rice leaf sheath, indicating a critical role of MC69 in interaction with host plants. MC69 encodes a hypothetical 54 amino acids protein with a signal peptide. Live-cell imaging suggested that fluorescently labeled MC69 was not translocated into rice cytoplasm. Site-directed mutagenesis of two conserved cysteine residues (Cys36 and Cys46) in the mature MC69 impaired function of MC69 without affecting its secretion, suggesting the importance of the disulfide bond in MC69 pathogenicity function. Furthermore, deletion of the MC69 orthologous gene reduced pathogenicity of the cucumber anthracnose fungus Colletotrichum orbiculare on both cucumber and Nicotiana benthamiana leaves. We conclude that MC69 is a secreted pathogenicity protein commonly required for infection of two different plant pathogenic fungi, M. oryzae and C. orbiculare pathogenic on monocot and dicot plants, respectively. Magnaporthe oryzae causes the most devastating fungal disease in rice. M. oryzae secretes a plethora of effector proteins, including several avirulence proteins which are known to be recognized by host resistance proteins activating innate immunity. However, the effectors that are required for virulence activity have not been identified in M. oryzae to date except for an effector protein, Secreted LysM Protein 1 (Slp1) that was recently identified. We performed a large-scale disruption analysis of M. oryzae effector candidates and identified a small protein MC69, which is secreted by the fungus during infection. When MC69 is absent, pathogenicity is severely reduced after penetration into the host cells. Furthermore, deletion of the MC69 orthologous gene in Colletotrichum orbiculare reduced its pathogenicity in the host plants cucumber and Nicotiana benthamiana. Thus, MC69 is conserved in ascomycete fungi and is crucial for establishing compatibility. This is the first report of a single secreted protein that is indispensable for pathogenicity in both monocot and dicot pathogenic fungi. How MC69 contributes to pathogenicity or virulence is unknown but it could be required for the fungus to be a pathogen or might be a classical effector that acts on plant target molecules.
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影响因子：
11.6
作者：
Mosquera, Gloria;Giraldo, Martha C.;Valent, Barbara
通讯作者：
Valent, Barbara
影响因子：
11.4
作者：
Jia, Y;McAdams, SA;Valent, B
通讯作者：
Valent, B
影响因子：
30.5
作者：
Ausubel, FM
通讯作者：
Ausubel, FM
DOI：
10.1126/science.1171647
发表时间：
2009-05-08
期刊：
Science (New York, N.Y.)
影响因子：
--
作者：
Boller T;He SY
通讯作者：
He SY
影响因子：
14.9
作者：
Ceroni, Alessio;Passerini, Andrea;Vullo, Alessandro;Frasconi, Paolo
通讯作者：
Frasconi, Paolo