State Key Laboratory of Genetic Engineering and Institute of Plant Biology, School of Life Sciences, Fudan University, Shanghai 200433, People's Republic of China;
College of Life Science and Technology, Huazhong Agricultural University, Wuhan 430070, People's Republic of China;
Proc Natl Acad Sci U S A. 2014 Jul 1;111(26):9651-6. doi: 10.1073/pnas.1316717111. Epub 2014 Jun 13.
Arabidopsis glycogen synthase kinase 3 (GSK3)-like kinases have versatile functions in plant development and in responding to abiotic stresses. Although physiological evidence suggested a potential role of GSK3-like kinases in abscisic acid (ABA) signaling, the underlying molecular mechanism was largely unknown. Here we identified members of Snf1-related kinase 2s (SnRK2s), SnRK2.2 and SnRK2.3, that can interact with and be phosphorylated by a GSK3-like kinase, brassinosteroid insensitive 2 (BIN2). bin2-3 bil1 bil2, a loss-of-function mutant of BIN2 and its two closest homologs, BIN2 like 1 (BIL1) and BIN2 like 2 (BIL2), was hyposensitive to ABA in primary root inhibition, ABA-responsive gene expression, and phosphorylating ABA Response Element Binding Factor (ABF) 2 fragment by in-gel kinase assays, whereas bin2-1, a gain-of-function mutation of BIN2, was hypersensitive to ABA, suggesting that these GSK3-like kinases function as positive regulators in ABA signaling. Furthermore, BIN2 phosphorylated SnRK2.3 on T180, and SnRK2.3(T180A) had decreased kinase activity in both autophosphorylation and phosphorylating ABFs. Bikinin, a GSK3 kinase inhibitor, inhibited the SnRK2.3 kinase activity and its T180 phosphorylation in vivo. Our genetic analysis further demonstrated that BIN2 regulates ABA signaling downstream of the PYRABACTIN RESISTANCE1/PYR1-LIKE/REGULATORY COMPONENTS OF ABA RECEPTORS receptors and clade A protein phosphatase 2C but relies on SnRK2.2 and SnRK2.3. These findings provide significant insight into the modulation of ABA signaling by Arabidopsis GSK3-like kinases.
拟南芥糖原合酶激酶 3（GSK3）样激酶在植物发育和应对非生物胁迫中具有多种功能。尽管生理证据表明 GSK3 样激酶在脱落酸（ABA）信号转导中可能发挥作用，但潜在的分子机制在很大程度上尚不清楚。在这里，我们鉴定了 Snf1 相关激酶 2（SnRK2）成员 SnRK2.2 和 SnRK2.3，它们可以与 GSK3 样激酶油菜素内酯不敏感 2（BIN2）相互作用并被其磷酸化。bin2-3 bil1 bil2，BIN2 和其两个最接近的同源物 BIN2 样 1（BIL1）和 BIN2 样 2（BIL2）的功能丧失突变体，对 ABA 抑制主根伸长、ABA 响应基因表达以及在凝胶激酶测定中磷酸化 ABA 反应元件结合因子（ABF）2 片段的敏感性降低，而 BIN2 的功能获得性突变体 bin2-1 对 ABA 则敏感，这表明这些 GSK3 样激酶在 ABA 信号转导中作为正调控因子发挥作用。此外，BIN2 在 T180 处磷酸化 SnRK2.3，而 SnRK2.3（T180A）在自身磷酸化和磷酸化 ABFs 方面的激酶活性均降低。Bikinin，一种 GSK3 激酶抑制剂，在体内抑制 SnRK2.3 激酶活性及其 T180 磷酸化。我们的遗传分析进一步表明，BIN2 调节 PYRABACTIN RESISTANCE1/PYR1-LIKE/ABA 受体调节元件受体和 clade A 蛋白磷酸酶 2C 下游的 ABA 信号转导，但依赖于 SnRK2.2 和 SnRK2.3。这些发现为拟南芥 GSK3 样激酶调节 ABA 信号转导提供了重要的见解。
