Studies on morphogenesis of infection structure and signal network in plant pathogenic fungi - regulatory mechanisms of environmental response and gene expression

植物病原真菌侵染结构与信号网络形态建成研究——环境响应与基因表达调控机制

• 批准号: 12460024
• 负责人: KUDO Yasuyuki
• 金额: $ 3.97万
• 依托单位: KYOTO PREFECTURAL UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-12460024/
• 关键词: Colletotrichum; Infection structure; Appressoria; Morphogenesis; Signal transduction; Gene expression; Response to environment; Plant pathogenic fungus

1) Isolation and characterization of genes involved in morphogenesis of infection structure and signal networkThe C. lagenarium GMK1 gene, a homologue of the Saccharomyces cerevisiae FUS3/KSS1 MAP kinase genes, was shown to regulate conidial germination, appressorium formation, and invasive growth. In S. cerevisiae, Ste12p is known to be a transcriptional factor downstream of Fus3p/Kss1p MAP kinases. In this research, to evaluate the CMK1 MAP kinase pathway, the Ste12 homologue CST1 gene from C. lagenarium was isolated and characterized. cst1Δ strains were nonpathogenic on intact host leaves, but could form lesions when inoculated on wounded leaves. Conidia of the cst1Δ strains could germinate and form melanized appressoria on both host leaf surface and artificial cellulose membrane, but could not produce infectious hyphae from appressoria, suggesting that CST1 is essential for appressorium penetration in C. lagenarium. In addition, matured appressoria of the cst1Δ strains contained ex … More tremely low level of lipid droplets, compared with that of the wild type strain. Lipid droplets were abundant in conidia of the cst1Δ strains, but rapidly disappeared during appressorium formation. This misscheduled lipid degradation might be related to the failure of appressorium penetration in the cst1Δ strain.2) Establishment of Agrobacterium tumefaciens-mediated transformation as an experimental tool for random insertional mutagenesis in Colletotrichum lagenariumIn this research, Agrobacterium tumefociens-mediated transformation (AtMT) was applied to Colletotrichum lagenarium for random insertional mutagenesis. Optimal co-cultivation of C. lagenarium wild-type 104-T with pBIG2RHPH2, a binary vector carrying hygromycin-resistant gene cassette, introduced A. tumefaciens C58C1 led to good production of hygromycin-resistant transformants. The fungal genomic DNA segments flanking T-DNA could be identified from several pathogenicity or morphogenesis deficient mutants by thermal asymmetric interlaced-polymerase chain reaction (TAIL-PCR). Sequence from one of the mutants was identical with melanin biosynthesis gene PKS1 of C. lagenarium that we have previously characterized. These results indicated that AtMT could be an effective tool for tagging genes relevant to pathogenicity and morphogenesis in the plant pathogenic fungus C. lagenarium. Less

1)感染结构和信号网络形态发生相关基因的分离和鉴定Lagenarium GMK 1基因是酿酒酵母FUS 3/KSS 1 MAP激酶基因的同源物，被证明调节分生孢子萌发、附着胞形成和侵入生长。In S.已知Ste 12 p是Fus 3 p/Kss 1 p MAP激酶下游的转录因子。本研究利用C.分离并表征了Lagenarium。cst 1 Δ菌株对完整的寄主叶片无致病性，但对受伤的寄主叶片有致病性。Cst 1 Δ菌株的分生孢子在寄主叶片表面和人工纤维素膜上均能萌发并形成黑化附着胞，但不能从附着胞中产生侵染性菌丝，表明Cst 1是C.拉根纳瑞。此外，cst 1 Δ菌株的成熟附着胞含有ex ...更多信息 与野生型菌株相比，脂滴水平极低。cst 1 Δ菌株的分生孢子中含有丰富的脂滴，但在附着胞形成过程中迅速消失。cst 1 Δ菌株的脂质降解可能与附着胞穿透失败有关。2）根癌农杆菌介导的西瓜炭疽菌随机插入突变体的建立本研究利用根癌农杆菌介导的转化（Agrobacterium tumefociens-mediated transformation，AtMT）对西瓜炭疽菌进行随机插入突变。优化了C.将野生型104-T与携带潮霉素抗性基因盒的双元载体pBIG 2 RHPH 2连接，将A.根癌农杆菌C58 C1导致潮霉素抗性转化子的良好产生。利用热不对称交错聚合酶链反应（TAIL-PCR）技术，可以从几种致病性或形态发生缺陷突变体中鉴定出T-DNA侧翼的真菌基因组DNA片段。其中一株突变体的基因序列与C.我们之前描述过的拉格纳里。这些结果表明，AtMT可以作为标记植物病原真菌C.拉根纳瑞。少