稻瘟病菌假定核糖体生成因子MoRei1功能研究

doi:10.3785/j.issn.1008-9209.2021.06.153

浙江大学学报（农业与生命科学版）

2022, Vol. 48

Issue (4): 443-452 DOI: 10.3785/j.issn.1008-9209.2021.06.153

植物保护

稻瘟病菌假定核糖体生成因子MoRei1功能研究

汤帅(

),徐喆,吕务云,童琪,肖宇,王政逸(

)

浙江大学生物技术研究所，水稻生物学国家重点实验室，杭州 310058

Functional analysis of the putative ribosome biogenesis factor MoRei1in Magnaporthe oryzae

Shuai TANG(

),Zhe XU,Wuyun Lü,Qi TONG,Yu XIAO,Zhengyi WANG(

)

State Key Laboratory of Rice Biology, Institute of Biotechnology, Zhejiang University, Hangzhou 310058, China

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摘要：

为深入探究稻瘟病菌假定核糖体生成因子MoRei1的生物学功能，通过敲除MoREI1基因、分析ΔMorei1突变体表型、鉴定基因互补及其互作蛋白功能等对其进行了研究。结果表明：敲除MoREI1基因后，稻瘟病菌ΔMorei1突变体的有性生殖能力丧失，附着胞形成率显著下降，致病力减弱；ΔMorei1突变体对细胞壁和氧化胁迫因子的敏感性增强，附着胞发育过程中糖原的移动和降解速率下降。稻瘟病菌MoREI1基因可恢复酿酒酵母REI1基因缺失突变体的部分表型，说明MoRei1与酵母核糖体生成因子Rei1在功能上具有同源性，并可能在核糖体生成过程中起类似的作用。稻瘟病菌MoRei1蛋白与预测的核输出因子MoAlb1间存在物理互作，且MoREI1基因缺失可影响稻瘟病菌MoAlb1蛋白的亚细胞定位。上述研究结果对阐明MoRei1在稻瘟病菌形态分化和致病性中的作用及其机制有重要意义。

关键词： 稻瘟病菌; 核糖体生成因子MoRei1; 功能分析; MoAlb1蛋白

Abstract:

To further explore the biological roles of MoRei1in the rice blast fungus Magnaporthe oryzae, we carried out MoREI1 gene deletion, phenotypic analysis of ΔMorei1 mutants, gene complementation and identification of its interaction proteins. The results showed that the ΔMorei1 mutants were defective in sexual reproduction, appressorium formation and pathogenicity. Deletion of MoREI1 led to significant increase of sensitivity to cell wall damaging and oxidative stress agents, and delay of glycogen mobilization and degradation during appressorium development. Moreover, the MoREI1 gene could partially complement the defects of Saccharomyces cerevisiae Δrei1 mutants, indicating that MoRei1 is a functional homolog of yeast Rei1, and may share a similar role in ribosome biogenesis. MoRei1 physically interacts with MoAlb1, a predicted nuclear export factor. Interestingly, deletion of MoREI1 resulted in the impairment of nuclear localization of MoAlb1. These results lay a foundation to clarify the roles of MoRei1 in fungal morphogenesis and pathogenicity in M. oryzae.

Key words: Magnaporthe oryzae ribosome biogenesis factor MoRei1 functional analysis MoAlb1 protein

收稿日期: 2021-06-15 出版日期: 2022-09-03

CLC:

S 432.1

基金资助: 国家自然科学基金项目(31770153)

通讯作者: 王政逸 E-mail: 21816071@zju.edu.cn;zhywang@zju.edu.cn

作者简介: 汤帅（https://orcid.org/0000-0001-6724-8850），E-mail：21816071@zju.edu.cn

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引用本文:

汤帅,徐喆,吕务云,童琪,肖宇,王政逸. 稻瘟病菌假定核糖体生成因子MoRei1功能研究[J]. 浙江大学学报（农业与生命科学版）, 2022, 48(4): 443-452.

Shuai TANG,Zhe XU,Wuyun Lü,Qi TONG,Yu XIAO,Zhengyi WANG. Functional analysis of the putative ribosome biogenesis factor MoRei1in Magnaporthe oryzae. Journal of Zhejiang University (Agriculture and Life Sciences), 2022, 48(4): 443-452.

链接本文:

https://www.zjujournals.com/agr/CN/10.3785/j.issn.1008-9209.2021.06.153 或 https://www.zjujournals.com/agr/CN/Y2022/V48/I4/443

表 1 本研究所用引物

图1 不同真菌Rei1同源蛋白质的系统进化分析

图2 酵母野生型BY4741、突变体Δrei1 及回补菌株Δreil/MoREI1在不同温度下的生长情况阴性对照：在酿酒酵母突变体菌株YBR267W中导入空载质粒pYES2。

图3 稻瘟病菌野生型Guy11与ΔMorei1-22 突变体的有性生殖测定

图4 MoREI1 基因缺失对稻瘟病菌致病性、附着胞形成及侵染过程的影响A. 水稻叶片喷雾分生孢子5 d后病斑形成情况；B. 附着胞形态（24 h）；C. 附着胞形成率统计（24 h）；D. 侵入菌丝类型（AP：附着胞；IH：侵入菌丝）；E. 大麦表皮穿透实验及各类型侵染菌丝所占比例（48 h）。**表示在P＜0.01水平差异有统计学意义。

图5 稻瘟病菌野生型菌株Guy11与ΔMorei1-22 突变体在含细胞壁及氧化胁迫因子条件下的生长情况A. 各测试菌株在胁迫培养基上的生长情况；B. 各测试菌株的生长抑制率。CM：完全培养基；CR：刚果红。*表示在P＜0.05水平差异有统计学意义，下同。

图6 MoREI1 基因缺失对稻瘟病菌附着胞形成过程中糖原移动及降解速率的影响A. 分生孢子染色情况；B. 各测试菌株分生孢子内的糖原转移情况统计；C. 各测试菌株附着胞内的糖原消耗情况统计。

图7 MoRei1互作蛋白的鉴定阳性对照：酿酒酵母野生型菌株BY4741中导入质粒pGADT7+pGBKT7-53；阴性对照：酿酒酵母野生型菌株BY4741中导入质粒pGADT7+pGBKT7-lam。AD：激活域； BD： DNA结合域。

图8 MoREI1 基因缺失对MoAlb1细胞核定位的影响

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