Department of Systems Biology, College of Life Science and Biotechnology, Yonsei University, Seoul 120-749, Korea.
Plant Physiol. 2011 Dec;157(4):2240-57. doi: 10.1104/pp.111.185595. Epub 2011 Oct 10.
The ubiquitin (Ub)-26S proteasome pathway is implicated in various cellular processes in higher plants. AtAIRP1, a C3H2C3-type RING (for Really Interesting New Gene) E3 Ub ligase, is a positive regulator in the Arabidopsis (Arabidopsis thaliana) abscisic acid (ABA)-dependent drought response. Here, the AtAIRP2 (for Arabidopsis ABA-insensitive RING protein 2) gene was identified and characterized. AtAIRP2 encodes a cytosolic C3HC4-type RING E3 Ub ligase whose expression was markedly induced by ABA and dehydration stress. Thus, AtAIRP2 belongs to a different RING subclass than AtAIRP1 with a limited sequence identity. AtAIRP2-overexpressing transgenic (35S:AtAIRP2-sGFP) and atairp2 loss-of-function mutant plants exhibited hypersensitive and hyposensitive phenotypes, respectively, to ABA in terms of seed germination, root growth, and stomatal movement. 35S:AtAIRP2-sGFP plants were highly tolerant to severe drought stress, and atairp2 alleles were more susceptible to water stress than were wild-type plants. Higher levels of drought-induced hydrogen peroxide production were detected in 35S:AtAIRP2-sGFP as compared with atairp2 plants. ABA-inducible drought-related genes were up-regulated in 35S:AtAIRP2-sGFP and down-regulated in atairp2 progeny. The positive effects of AtAIRP2 on ABA-induced stress genes were dependent on SNF1-related protein kinases, key components of the ABA signaling pathway. Therefore, AtAIRP2 is involved in positive regulation of ABA-dependent drought stress responses. To address the functional relationship between AtAIRP1 and AtAIRP2, FLAG-AtAIRP1 and AtAIRP2-sGFP genes were ectopically expressed in atairp2-2 and atairp1 plants, respectively. Constitutive expression of FLAG-AtAIRP1 and AtAIRP2-sGFP in atairp2-2 and atairp1 plants, respectively, reciprocally rescued the loss-of-function ABA-insensitive phenotypes during germination. Additionally, atairp1/35S:AtAIRP2-sGFP and atairp2-2/35S:FLAG-AtAIRP1 complementation lines were more tolerant to dehydration stress relative to atairp1 and atairp2-2 single knockout plants. Overall, these results suggest that AtAIRP2 plays combinatory roles with AtAIRP1 in Arabidopsis ABA-mediated drought stress responses.
泛素（Ub）-26S 蛋白酶体途径参与高等植物的各种细胞过程。在拟南芥中，AtAIRP1 是一种 C3H2C3 型 RING（代表真正有趣的新基因）E3 Ub 连接酶，是 ABA 依赖性干旱响应的正调节剂。在这里，鉴定并表征了 AtAIRP2（拟南芥 ABA 不敏感 RING 蛋白 2）基因。AtAIRP2 编码一种细胞质 C3HC4 型 RING E3 Ub 连接酶，其表达受 ABA 和脱水胁迫的显著诱导。因此，AtAIRP2 属于与 AtAIRP1 不同的 RING 亚类，其序列同一性有限。在种子萌发、根生长和气孔运动方面，过表达 AtAIRP2 的转基因（35S：AtAIRP2-sGFP）和 atairp2 功能丧失突变体植物表现出对 ABA 的超敏和低敏表型。35S：AtAIRP2-sGFP 植物对严重干旱胁迫高度耐受，而 atairp2 等位基因比野生型植物更容易受到水分胁迫。与 atairp2 植物相比，在 35S：AtAIRP2-sGFP 中检测到更高水平的干旱诱导过氧化氢产生。ABA 诱导的耐旱相关基因在 35S：AtAIRP2-sGFP 中上调，在 atairp2 后代中下调。AtAIRP2 对 ABA 诱导的应激基因的正向作用依赖于 SNF1 相关蛋白激酶，这是 ABA 信号通路的关键组成部分。因此，AtAIRP2 参与正向调控 ABA 依赖性干旱应激反应。为了解 AtAIRP1 和 AtAIRP2 之间的功能关系，分别在 atairp2-2 和 atairp1 植物中外源表达 FLAG-AtAIRP1 和 AtAIRP2-sGFP 基因。在 atairp2-2 和 atairp1 植物中组成型表达 FLAG-AtAIRP1 和 AtAIRP2-sGFP 分别在萌发过程中挽救了 ABA 不敏感表型的功能丧失。此外，与 atairp1/35S：AtAIRP2-sGFP 和 atairp2-2/35S：FLAG-AtAIRP1 互补系相比，atairp1 和 atairp2-2 单敲除植物对脱水胁迫更具耐受性。总体而言，这些结果表明 AtAIRP2 与 AtAIRP1 一起在拟南芥 ABA 介导的干旱应激反应中发挥组合作用。
