Takahashi Yohei, Ebisu Yuta, Shimazaki Ken-Ichiro
Department of Biology, Faculty of Science, Kyushu University 744 Motooka, Fukuoka 819-0395, Japan
Department of Biology, Faculty of Science, Kyushu University 744 Motooka, Fukuoka 819-0395, Japan.
Plant Physiol. 2017 Jun;174(2):815-822. doi: 10.1104/pp.16.01825. Epub 2017 Apr 24.
The plant hormone abscisic acid (ABA) confers drought tolerance in plants through stomatal closure and regulation of gene expression. The complex consisting of the ABA receptor PYRABACTIN RESISTANCE/REGULATORY COMPONENTS OF ABA RECEPTOR (PYR/RCAR), type 2C protein phosphatase (PP2C), and SNF1-related protein kinase 2 (SnRK2) has a key role in ABA signaling. Basic helix-loop-helix (bHLH) transcriptional activator ABA-RESPONSIVE KINASE SUBSTRATE1 (AKS1, also known as FBH3) is released from DNA by phosphorylation-induced monomerization in response to ABA in guard cells. Here we reconstituted the release of AKS1 from DNA via the ABA signaling core complex in vitro. We first obtained evidence to confirm that AKS1 is an endogenous substrate for SnRK2s. Phosphorylation of AKS1 and activation of SnRK2 showed the same time course in response to ABA in guard cells. AKS1 was bound to SnRK2.6 in vivo. Three ABA-responsive SnRK2s (SnRK2.2/SRK2D, SnRK2.3/SRK2I, and SnRK2.6/SRK2E/OST1) phosphorylated AKS1 in vitro, and the phosphorylation was eliminated by the triple mutation of SnRK2s in plants. We reconstituted the AKS1 phosphorylation in vitro via the signaling complex containing the ABA receptor PYR1, a PP2C, HYPERSENSITIVE TO ABA1 (HAB1), and a protein kinase, SnRK2.6 in response to ABA We further reconstituted the release of AKS1 from the target gene of () via the complex in response to ABA These results demonstrate that AKS1 provides a link between the signaling complex and ABA-responsive genes and furnish evidence for a minimal signaling mechanism from ABA perception to DNA.
植物激素脱落酸（ABA）通过气孔关闭和基因表达调控赋予植物耐旱性。由ABA受体PYRABACTIN RESISTANCE/ABA受体调节成分（PYR/RCAR）、2C型蛋白磷酸酶（PP2C）和SNF1相关蛋白激酶2（SnRK2）组成的复合物在ABA信号传导中起关键作用。碱性螺旋-环-螺旋（bHLH）转录激活因子ABA应答激酶底物1（AKS1，也称为FBH3）在保卫细胞中响应ABA时通过磷酸化诱导的单体化从DNA上释放。在这里，我们在体外通过ABA信号核心复合物重建了AKS1从DNA上的释放。我们首先获得证据证实AKS1是SnRK2s的内源性底物。在保卫细胞中，AKS1的磷酸化和SnRK2的激活在响应ABA时显示出相同的时间进程。AKS1在体内与SnRK2.6结合。三种ABA应答性SnRK2s（SnRK2.2/SRK2D、SnRK2.3/SRK2I和SnRK2.6/SRK2E/OST1）在体外使AKS1磷酸化，并且该磷酸化在植物中通过SnRK2s的三突变而消除。我们通过包含ABA受体PYR1、PP2C、对ABA敏感1（HAB1）和蛋白激酶SnRK2.6的信号复合物在体外重建了响应ABA的AKS1磷酸化。我们进一步通过该复合物在响应ABA时重建了AKS1从（）的靶基因上的释放。这些结果表明，AKS1在信号复合物和ABA应答基因之间提供了联系，并为从ABA感知到DNA的最小信号机制提供了证据。
