水稻miR444b.2调控稻瘟病抗性及分蘖

MiR444b.2 regulates resistance to Magnaporthe oryzae and tillering in rice

microRNA(miRNA)是一类长度在20~24 bp的保守小RNA分子,广泛存在于真核生物中,参与调控生长发育及抗逆、抗病等反应。在前期研究中,我们对比感病和抗病水稻材料在接种稻瘟病菌前后miRNA的累积量,发现miR444b.2的累积量变化在抗病和感病材料间存在显著差异,但其在水稻对稻瘟病抗性中的作用尚不清楚。本文进一步对水稻感病和抗病材料接种稻瘟病菌,通过检测不同时间点miR444b.2及其靶基因的累积量来分析miR444b.2与水稻稻瘟病抗性的关系;同时,构建过表达miR444b.2的转基因水稻材料,对转基因材料进行了抗性分析及农艺性状分析。结果表明,在稻瘟病菌开始侵染水稻的24 h内,随着侵染时间的增加,miR444b.2在感病材料中的累积量也随之增加,而在抗病材料中的累积量则逐渐降低;相反,miR444b.2的靶基因中,MADS盒家族基因在感病材料中的转录受到抑制或变化不大,在抗病材料中的表达明显增加,与miR444b.2的累积量变化趋势相反,提示miR444b.2可能通过调节MADS盒家族基因的表达量来调控稻瘟病抗性。而且,过表达miR444b.2的转基因植株变得更加感病,主要表现为:与对照相比,叶片发病病斑数量增多、病斑面积增大、防御相关基因转录水平变低;而miR444b.2的靶基因在过表达miR444b.2的转基因植株中的转录水平显著降低。此外,miR444b.2过表达植株的分蘖数较对照减少。这些结果说明,miR444b.2通过调控MADS盒家族基因来负调控稻瘟病抗性及调节水稻分蘖。

MicroRNAs （miRNA）, a class of conservative small RNA molecules in eukaryotes, are involved in regulation of biological growth and stress-induced responses. In our previous study, we found that the accumula- tion of miR444b.2 in susceptible and resistant rice accessions was significantly different in response to Magna- porthe oryzae infection. However, the roles of miR544b.2 in rice immunity against M. oryzae were unclear. Here, we found that miR444b.2 negatively regulates rice defense against M. oryzae. First, we performed a time course analysis on the accumulation of miR^4~b.2 and the expression level of the target genes in susceptible and resistant rice accessions upon M. oryzae infection. Then we obtained transgenic rice lines over-expressing miR54~b.2 through the constitutive 35S promoter. With these transgenic rice lines, we further analyzed the effect of over-expression of miR444b.2 on disease resistance and morphological phenotypes of rice plants. Our results showed that within the first 24 hours of M. oryzae infection, the accumulation of miR444b.2 was increased in the susceptible accession LTH, but was gradually reduced in the resistant accession IRBLkm-Ts. In contrast, the ex- pression of the target MADS-box family genes of miR444b.2 was reversely correlated with the accumulation of miR444b.2, indicating that miRa^4：~b.2 might regulate rice immunity against M. oryzae by down-regulating the expression of MADS-box genes. Moreover, over-expression of miR444b.2 led to hyper-susceptibility to the viru- lent M. oryzae strain Guyll in transgenic rice lines, as indicated by higher number and bigger size of disease le- sions than that in the control plants. Consistently, the transcriptional levels of defense-related genes were lower in the transgenic lines over-expressing miR^AAb.2 than those of the control plants. In addition, the tiller number of miR~：~a.b.2 transgenic lines was less than that of control plants, which was attributable to the suppression of the tiller-determinant MADS-box family genes. Taken together, our data suggest that miR444b.2 negatively regu- lates rice immunity against M. oryzae and the tiller number via suppressing the MADS-box family genes.