Under drought stress, reactive oxygen species (ROS) overaccumulate as a secondary stress that impairs plant performance and thus severely reduces crop yields. The mitigation of ROS levels under drought stress is therefore crucial for drought tolerance. MicroRNAs (miRNAs) are critical regulators of plant development and stress responses. However, the complex molecular regulatory mechanism by which they function during drought stress, especially in drought-triggered ROS scavenging, is not fully understood. Here, we report a newly identified drought-responsive miRNA, miR164g, in the wild apple species Malus sieversii and elucidate its role in apple drought tolerance. Our results showed that expression of miR164g is significantly inhibited under drought stress and it can specifically cleave transcripts of the transcription factor MsNAC022 in M. sieversii. The heterologous accumulation of miR164g in Arabidopsis thaliana results in enhanced sensitivity to drought stress, while overexpression of MsNAC022 in Arabidopsis and the cultivated apple line ‘GL-3’ (Malus domestica Borkh.) lead to enhanced tolerance to drought stress by raising the ROS scavenging enzymes activity and related genes expression levels, particularly PEROXIDASE (MsPOD). Furthermore, we showed that expression of MsPOD is activated by MsNAC022 in transient assays. Interestingly, Part1 (P1) region is the key region for the positive regulation of MsPOD promoter by MsNAC022, and the different POD expression patterns in M. sieversii and M. domestica is attributed to the specific fragments inserted in P1 region of M. sieversii. Our findings reveal the function of the miR164g-MsNAC022 module in mediating the drought response of M. sieversii and lay a foundation for breeding drought-tolerant apple cultivars.

中文翻译：

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miR164g-MsNAC022 作为一种新的模块通过苹果活性氧清除系统的转录调控来介导干旱反应

在干旱胁迫下，活性氧 (ROS) 作为次生胁迫过度积累，会损害植物性能，从而严重降低作物产量。因此，干旱胁迫下活性氧水平的降低对于耐旱性至关重要。MicroRNAs (miRNAs) 是植物发育和应激反应的关键调节因子。然而，它们在干旱胁迫期间发挥作用的复杂分子调控机制，特别是在干旱引发的 ROS 清除中，尚不完全清楚。在这里，我们报告了野生苹果品种 Malus sieversii 中新发现的干旱响应 miRNA，miR164g，并阐明了它在苹果耐旱性中的作用。我们的研究结果表明，在干旱胁迫下，miR164g的表达受到显着抑制，它可以特异性切割MsNAC022转录因子的转录本。sieversii. 拟南芥中 miR164g 的异源积累导致对干旱胁迫的敏感性增强，而 MsNAC022 在拟南芥和栽培苹果系 'GL-3' (Malus domestica Borkh.) 中的过表达通过提高 ROS 清除能力导致对干旱胁迫的耐受性增强酶活性和相关基因表达水平，特别是过氧化物酶（MsPOD）。此外，我们发现 MsPOD 的表达在瞬时测定中被 MsNAC022 激活。有趣的是，Part1（P1）区域是MsNAC022正向调控MsPOD启动子的关键区域，而M. sieversii和M. domestica中不同的POD表达模式归因于插入M. sieversii P1区域的特定片段。