Abstract Plants under combined stresses exhibit a prominent shift in molecular responses compared with plants exposed to the same stresses independently. Profiling responses to individual and combined stressors at the gene expression level have identified several genes with intersecting responses to these stressors. However, the upstream regulators at the intersection of plant responses to individual and combined stresses are not known. Here, using the transcriptome of Arabidopsis thaliana under individual and combined drought and Pseudomonas syringae infection, we identified several genes whose expression overlaps between individual and combined stresses. To study the key regulator of such an overlapping gene, we predicted that the expression of 1-Pyrroline-5-carboxylate synthase 1 (AtP5CS1) is regulated by ath-miR164c at post-transcriptional level. Our results from the stem-loop RT-PCR based expression analysis revealed significant downregulation of ath-miR164c in response to P. syringae infection under both well-irrigated (pathogen only) and drought stress (combined stress) conditions. Furthermore, an Arabidopsis loss-of-function mutant of the miRNA ath-miR164c exhibited resistance to pathogen infection under combined stress, unlike the wild-type plants, implicating the role of ath-miR164c in regulating plant immunity. AtP5CS1 gene expression and proline accumulation were enhanced in the ath-miR164c mutant plants relative to the wild-type plants, demonstrating that ath-miR164c regulates AtP5CS1 of the proline biosynthesis pathway, which was also validated by 5’RLM-RACE results. This miRNA-mediated modulation of AtP5CS1 gene expression under combined stress fills crucial gaps in identifying the key convergent players in the current understanding of plant stress responses.

中文翻译：

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ath-miR164c 通过调节脯氨酸代谢影响植物对干旱和细菌感染联合胁迫的反应

摘要 与单独暴露于相同胁迫下的植物相比，在复合胁迫下的植物表现出显着的分子响应变化。在基因表达水平上分析对个体和组合压力源的反应已经确定了几个对这些压力源具有交叉反应的基因。然而，植物对个体和组合胁迫响应的交叉点的上游调节器是未知的。在这里，使用拟南芥在单独和联合干旱和丁香假单胞菌感染下的转录组，我们确定了几个基因，其表达在个体和联合胁迫之间重叠。为了研究这种重叠基因的关键调节因子，我们预测 1-Pyrroline-5-carboxylate synthase 1 (AtP5CS1) 的表达受 ath-miR164c 在转录后水平的调控。我们基于茎环 RT-PCR 的表达分析结果显示，在良好灌溉（仅病原体）和干旱胁迫（联合胁迫）条件下，ath-miR164c 显着下调以响应丁香假单胞菌感染。此外，与野生型植物不同，miRNA ath-miR164c 功能丧失的拟南芥突变体在复合胁迫下表现出对病原体感染的抗性，这表明 ath-miR164c 在调节植物免疫中的作用。与野生型植物相比，ath-miR164c 突变植物中的 AtP5CS1 基因表达和脯氨酸积累增强，表明 ath-miR164c 调节脯氨酸生物合成途径的 AtP5CS1，这也得到了 5'RLM-RACE 结果的验证。这种在联合胁迫下 miRNA 介导的 AtP5CS1 基因表达调节填补了目前对植物胁迫反应理解中关键趋同参与者的鉴定空白。