Agricultural Biotechnology Research Center, Academia Sinica, Taipei 115, Taiwan.
Plant Cell Physiol. 2010 Dec;51(12):2119-31. doi: 10.1093/pcp/pcq170. Epub 2010 Nov 9.
Here we report on the characterization of rice osa-miR827 and its two target genes, OsSPX-MFS1 and OsSPX-MFS2, which encode SPX-MFS proteins predicted to be implicated in phosphate (Pi) sensing or transport. We first show by Northern blot analysis that osa-miR827 is strongly induced by Pi starvation in both shoots and roots. Hybridization of osa-miR827 in situ confirms its strong induction by Pi starvation, with signals concentrated in mesophyll, epidermis and ground tissues of roots. In parallel, we analyzed the responses of the two OsSPX-MFS1 and OsSPX-MFS2 gene targets to Pi starvation. OsSPX-MFS1 mRNA is mainly expressed in shoots under sufficient Pi supply while its expression is reduced on Pi starvation, revealing a direct relationship between induction of osa-miR827 and down-regulation of OsSPX-MFS1. In contrast, OsSPX-MFS2 responds in a diametrically opposed manner to Pi starvation. The accumulation of OsSPX-MFS2 mRNA is dramatically enhanced under Pi starvation, suggesting the involvement of complex regulation of osa-miR827 and its two target genes. We further produced transgenic rice lines overexpressing osa-miR827 and T-DNA knockout mutant lines in which the expression of osa-miR827 is abolished. Compared with wild-type controls, both target mRNAs exhibit similar changes, their expression being reduced and increased in overexpressing and knockout lines, respectively. This suggests that OsSPX-MFS1 and OsSPX-MFS2 are both negatively regulated by osa-miR827 abundance although they respond differently to external Pi conditions. We propose that this is a complex mechanism comprising fine tuning of spatial or temporal regulation of both targets by osa-miR827.
在这里，我们报告了水稻 osa-miR827 及其两个靶基因 OsSPX-MFS1 和 OsSPX-MFS2 的特征，它们编码的 SPX-MFS 蛋白被预测参与磷酸盐（Pi）感应或运输。我们首先通过 Northern blot 分析表明，在地上部和根部，osa-miR827 强烈地被 Pi 饥饿诱导。osa-miR827 的原位杂交证实了其强烈的 Pi 饥饿诱导，信号集中在叶肉、表皮和根的地组织中。与此同时，我们分析了两个 OsSPX-MFS1 和 OsSPX-MFS2 基因靶标的对 Pi 饥饿的反应。在充足 Pi 供应下，OsSPX-MFS1 mRNA 主要在地上部表达，而在 Pi 饥饿时其表达减少，这揭示了 osa-miR827 的诱导与 OsSPX-MFS1 的下调之间存在直接关系。相比之下，OsSPX-MFS2 对 Pi 饥饿的反应则截然相反。在 Pi 饥饿下，OsSPX-MFS2 mRNA 的积累显著增强，表明 osa-miR827 及其两个靶基因的复杂调控。我们进一步生成了过表达 osa-miR827 的转基因水稻系和 T-DNA 敲除突变系，其中 osa-miR827 的表达被消除。与野生型对照相比，两个靶标 mRNA 都表现出相似的变化，它们的表达分别在过表达和敲除系中减少和增加。这表明 OsSPX-MFS1 和 OsSPX-MFS2 都受到 osa-miR827 丰度的负调控，尽管它们对外部 Pi 条件的反应不同。我们提出，这是一个复杂的机制，包括通过 osa-miR827 对两个靶标进行时空调节的精细调整。
