Many microRNAs (miRNAs) are critical regulators of plant antiviral defense. However, little is known about how these miRNAs respond to virus invasion at the transcriptional level. We previously show that defense against Rice stripe virus (RSV) invasion entailed a reduction of miR528 accumulation in rice, alleviating miR528-mediated degradation of L-Ascorbate Oxidase (AO) mRNA and bolstering the antiviral activity of AO. Here we show that the miR528-AO defense module is regulated by the transcription factor SPL9. SPL9 displayed high-affinity binding to specific motifs within the promoter region of miR528 and activated the expression of miR528 gene in vivo. Loss-of-function mutations in SPL9 caused a significant reduction in miR528 accumulation but a substantial increase of AO mRNA, resulting in enhanced plant resistance to RSV. Conversely, transgenic overexpression of SPL9 stimulated the expression of miR528 gene, hence lowering the level of AO mRNA and compromising rice defense against RSV. Importantly, gain in RSV susceptibility did not occur when SPL9 was overexpressed in mir528 loss-of-function mutants, or in transgenic rice expressing a miR528-resistant AO. Taken together, the finding of SPL9-mediated transcriptional activation of miR528 expression adds a new regulatory layer to the miR528-AO antiviral defense pathway.

许多microRNA（miRNA）是植物抗病毒防御的关键调节剂。然而，关于这些miRNA如何在转录水平上对病毒入侵的反应知之甚少。我们以前表明防御水稻条纹病毒（RSV）入侵需要减少miR528在水稻中的积累，减轻miR528介导的L-抗坏血酸氧化酶（AO）mRNA降解并增强AO的抗病毒活性。在这里，我们显示了miR528-AO防御模块受转录因子SPL9调控。SPL9显示出与miR528启动子区域内特定基序的高亲和力结合，并激活了miR528基因在体内的表达。丧失功能的突变在SPL9引起miR528积累的显着减少，但AO mRNA的显着增加，导致植物对RSV的抗性增强。相反地，转基因过表达SPL9刺激miR528基因的表达，因此，降低的水平AO mRNA和损害针对RSV大米防御。重要的是，当没有发生增益RSV易感性SPL9中过表达mir528丧失功能突变体，或在转基因水稻表达miR528耐AO。综上，SPL9介导的miR528表达转录激活的发现为miR528-AO抗病毒防御途径增加了新的调控层。