bHLH transcription factors facilitating K+ uptake during stomatal opening are repressed by abscisic acid through phosphorylation. Arabidopsis thaliana

Stomata open in response to light and close following exposure to abscisic acid (ABA). They regulate gas exchange between plants and atmosphere, allowing plants to adapt to changing environmental conditions. ABA binding to receptors initiates a signaling cascade that involves protein phosphorylation. Here we show that ABA induced phosphorylation of three basic helix-loop-helix (bHLH) transcription factors, called AKSs (ABA-RESPONSIVE KINASE SUBSTRATES; AKS1, AKS2, AKS3), in Arabidopsis guard cells, and that they facilitated stomatal opening through the transcription of genes encoding inwardly-rectifying K+ channels. aks1aks2-1 double mutant plants showed decreases in light-induced stomatal opening, K+ accumulation in response to light, activity of inwardly-rectifying K+ channels, and transcription of genes encoding major inwardly-rectifying K+ channels. Overexpression of POTASSIUM CHANNEL IN ARABIDOPSIS THALIANA 1 (KAT1), which encodes a major inwardly-rectifying K+ channel in guard cells, rescued the phenotype of aks1aks2-1 plants. AKS1 bound directly to the promoter of KAT1, an interaction that was attenuated after ABA-induced phosphorylation. The ABA agonist pyrabactin induced phosphorylation of AKSs. Our results demonstrate that the AKS family of bHLH transcription factors facilitates stomatal opening through transcription of genes encoding inwardly-rectifying K+ channels, and that ABA suppresses the activity of inwardly-rectifying K+ channel activity by triggering the phosphorylation of these transcription factors. Overall design: To find the affect of AKS1 and AKS2 transcription factors on gene expression, Arabidopsis guard cell protoplasts from wild type and aks1aks2-1 mutant were compared. Three independent experiments were performed.

气孔（stomata）会响应光照而张开，并在暴露于脱落酸（abscisic acid, ABA）后关闭。它们介导植物与大气间的气体交换，帮助植物适应不断变化的环境条件。脱落酸与其受体结合后会启动涉及蛋白质磷酸化的信号级联反应。本研究发现，在拟南芥保卫细胞中，脱落酸可诱导三种名为AKS（ABA-RESPONSIVE KINASE SUBSTRATES；AKS1、AKS2、AKS3）的碱性螺旋-环-螺旋（basic helix-loop-helix, bHLH）转录因子发生磷酸化，且这些转录因子可通过促进编码内向整流钾通道（inwardly-rectifying K+ channels）的基因的转录来介导气孔张开。aks1aks2-1双突变体植株的光诱导气孔张开、光响应钾离子积累、内向整流钾通道活性以及主要内向整流钾通道编码基因的转录水平均出现降低。过表达在保卫细胞中编码主要内向整流钾通道的拟南芥钾通道1（POTASSIUM CHANNEL IN ARABIDOPSIS THALIANA 1, KAT1）可挽救aks1aks2-1植株的表型。AKS1可直接结合KAT1的启动子区域，该相互作用在脱落酸诱导的磷酸化后会被减弱。ABA激动剂吡巴芬（pyrabactin）可诱导AKS发生磷酸化。本研究结果表明，AKS家族的碱性螺旋-环-螺旋转录因子可通过促进编码内向整流钾通道的基因的转录来介导气孔张开，而脱落酸可通过触发这些转录因子的磷酸化来抑制内向整流钾通道的活性。实验设计概述：为探究AKS1与AKS2转录因子对基因表达的调控作用，本研究对比了野生型与aks1aks2-1突变体拟南芥的保卫细胞原生质体（guard cell protoplasts）的基因表达情况，共开展了三次独立重复实验。