亚甲基四氢叶酸还原酶参与调控稻瘟病菌的生长、发育和致病性

 稻瘟病菌(Magnaporth oryzae)引起的稻瘟病是水稻上一种毁灭性病害，每年给全球水稻生产造成10%～30%的产量损失，严重威胁着全球的粮食生产安全。从分子水平探究该病菌的致病机制，对于挖掘潜在的控制稻瘟病的药剂靶标具有重要的理论和实践意义。本研究在稻瘟病菌中鉴定到2个分别与酿酒酵母亚甲基四氢叶酸还原酶Met12和Met13同源的蛋白，命名为MoMet12和MoMet13。MoMet12和MoMet13分别含有690和631个氨基酸，两者的一致性为32%。对这2个蛋白编码基因分别进行敲除突变，发现ΔMomet12和ΔMomet13突变体在CM营养丰富培养基上生长减慢、气生菌丝减少。ΔMomet13在MM基本培养基上不能生长，在SDC和OM营养饥饿培养基上生长显著减慢，气生菌丝稀薄。ΔMomet12在MM、SDC和OM培养基上生长均显著减慢;对突变体进行产孢和致病性测定发现，ΔMomet13突变体在CM和SDC培养基上均丧失了产孢能力，而ΔMomet12在CM培养基上产孢量显著下降，在SDC上产孢能力丧失;ΔMomet13突变体侵染菌丝扩展和致病力显著下降，ΔMomet12突变体致病力与野生型比较无明显变化。加入外源的甲硫氨酸可恢复ΔMomet12和ΔMomet13突变体在不同培养基上的生长和产孢缺陷，且能部分恢复ΔMomet13突变体的致病能力。上述结果表明，MoMet12和MoMet13通过参与甲硫氨酸的生物合成，从而控制稻瘟病菌的生长和无性繁殖。MoMet13同时是一个重要的致病相关因子，参与病菌侵染菌丝的生长和致病过程。

Methylene tetrahydrofolate reductase regulates the growth,development and pathogenicity of the rice blast fungus Magnaporthe oryzae

Rice blast, caused by Magnaporthe oryzae, is one of the most devastating fungal diseases of rice worldwide. Clarification of pathogenic mechanisms of the pathogen will help to find the potential target sites for new fungicide, and therefore the effective way for disease control. In this study, two methylene tetrahydrofolate reductases MoMet12 and MoMet13 in M. oryzae, orthologous to Saccharomyces cerevisiae Met12 and Met13 were identified, which has 691 and 632 amino acids, respectively, and shares 32% identity. Deletion of Mo-MET12 and MoMET13 resulted in decreased vegetative growth and aerial hyphae formation on CM media. ΔMomet13 was auxotrophic on MM and significantly decreased in growth rate and formation of aerial hyphae on OM and SDC media. The growth rate of ΔMomet12 was also dramatically decreased on MM、SDC and OM media. Further analysis revealed that ΔMomet13 lost the ability to produce conidia on both CM and SDC media, and ΔMomet12 significantly reduced conidiation on CM and was unable to produce conidia on SDC media. Infection assay showed that ΔMomet13 was decreased in virulence and had a defect in infectious hyphal growth. Exogenous methionine could restore the growth and conidiation defects of ΔMomet12 and ΔMomet13 mutants, and also partially rescue the virulence of ΔMomet13. These results indicated that MoMet12 and MoMet13 have crucial roles in vegetative growth and asexual development by modulating the biosynthesis of methionine, and MoMet13 is a key pathogenicity-related factor involved in infectious hyphal growth and virulence.