Laboratory of Cell Function and Structure, Graduate School of Agricultural Science, Faculty of Agriculture, Kobe University, Kobe 657-8501, Japan.
Genetics. 2013 Nov;195(3):845-55. doi: 10.1534/genetics.113.155978. Epub 2013 Aug 26.
The long terminal repeat retrotransposon, Magnaporthe gypsy-like element (MAGGY), has been shown to be targeted for cytosine methylation in a subset of Magnaporthe oryzae field isolates. Analysis of the F1 progeny from a genetic cross between methylation-proficient (Br48) and methylation-deficient (GFSI1-7-2) isolates revealed that methylation of the MAGGY element was governed by a single dominant gene. Positional cloning followed by gene disruption and complementation experiments revealed that the responsible gene was the DNA methyltransferase, MoDMT1, an ortholog of Neurospora crassa Dim-2. A survey of MAGGY methylation in 60 Magnaporthe field isolates revealed that 42 isolates from rice, common millet, wheat, finger millet, and buffelgrass were methylation proficient while 18 isolates from foxtail millet, green bristlegrass, Japanese panicgrass, torpedo grass, Guinea grass, and crabgrass were methylation deficient. Phenotypic analyses showed that MoDMT1 plays no major role in development and pathogenicity of the fungus. Quantitative polymerase chain reaction analysis showed that the average copy number of genomic MAGGY elements was not significantly different between methylation-deficient and -proficient field isolates even though the levels of MAGGY transcript were generally higher in the former group. MoDMT1 gene sequences in the methylation-deficient isolates suggested that at least three independent mutations were responsible for the loss of MoDMT1 function. Overall, our data suggest that MoDMT1 is not essential for the natural life cycle of the fungus and raise the possibility that the genus Magnaporthe may be losing the mechanism of DNA methylation on the evolutionary time scale.
长末端重复逆转录转座子，稻曲菌类反转录元件（MAGGY），已被证明是在一组稻曲菌田间分离物中被胞嘧啶甲基化所靶向的。对来自甲基化能力强（Br48）和甲基化缺陷（GFSI1-7-2）分离物之间遗传杂交的 F1 后代的分析表明，MAGGY 元件的甲基化受一个单一的显性基因控制。通过位置克隆，然后进行基因敲除和互补实验，发现负责的基因是 DNA 甲基转移酶，MoDMT1，Neurospora crassa Dim-2 的同源物。对 60 个稻曲菌田间分离物中 MAGGY 甲基化的调查显示，来自水稻、谷子、小麦、珍珠粟和象草的 42 个分离物是甲基化能力强的，而来自黍、狗尾草、日本看麦娘、梯牧草、Guinea 草和稗草的 18 个分离物是甲基化缺陷的。表型分析表明，MoDMT1 在真菌的发育和致病性中没有起主要作用。定量聚合酶链反应分析表明，即使在前者中 MAGGY 转录本的水平通常更高，但在甲基化缺陷和甲基化能力强的田间分离物中基因组 MAGGY 元件的平均拷贝数没有显著差异。甲基化缺陷分离物中的 MoDMT1 基因序列表明，至少有三个独立的突变导致 MoDMT1 功能丧失。总的来说，我们的数据表明 MoDMT1 对真菌的自然生命周期不是必需的，并提出了 Magnaporthe 属可能在进化时间尺度上失去 DNA 甲基化机制的可能性。
