Xie Zhen, Zhang Zhong-Lin, Zou Xiaolu, Yang Guangxiao, Komatsu Setsuko, Shen Qingxi J
Department of Biological Sciences, University of Nevada, Las Vegas, NV 89154, USA.
Plant J. 2006 Apr;46(2):231-42. doi: 10.1111/j.1365-313X.2006.02694.x.
Gibberellins (GA) promote while abscisic acid (ABA) inhibits seed germination and post-germination growth. To address the cross-talk of GA and ABA signaling, we studied two rice WRKY genes (OsWRKY51 and OsWRKY71) that are ABA-inducible and GA-repressible in embryos and aleurone cells. Over-expression of these two genes in aleurone cells specifically and synergistically represses induction of the ABA-repressible and GA-inducible Amy32b alpha-amylase promoter reporter construct (Amy32b-GUS) by GA or the GA-inducible transcriptional activator, GAMYB. The physical interactions of OsWRKY71 proteins themselves and that of OsWRKY71 and OsWRKY51 are revealed in the nuclei of aleurone cells using bimolecular fluorescence complementation (BiFC) assays. Although OsWRKY51 itself does not bind to the Amy32b promoter in vitro, it interacts with OsWRKY71 and enhances the binding affinity of OsWRKY71 to W boxes in the Amy32b promoter. The binding activity of OsWRKY71 is abolished by deleting the C-terminus containing the WRKY domain or substituting the key amino acids in the WRKY motif and the zinc finger region. However, two of these non-DNA-binding mutants are still able to repress GA induction by enhancing the binding affinity of the wild-type DNA-binding OsWRKY71 repressors. In contrast, the third non-DNA-binding mutant enhances GA induction of Amy32b-GUS, by interfering with the binding of the wild-type OsWRKY71 or the OsWRKY71/OsWRKY51 repressing complex. These data demonstrate the synergistic interaction of ABA-inducible WRKY genes in regulating GAMYB-mediated GA signaling in aleurone cells, thereby establishing a novel mechanism for ABA and GA signaling cross-talk.
赤霉素（GA）促进种子萌发和萌发后的生长，而脱落酸（ABA）则抑制这一过程。为了研究GA和ABA信号转导的相互作用，我们研究了两个水稻WRKY基因（OsWRKY51和OsWRKY71），它们在胚胎和糊粉层细胞中受ABA诱导、受GA抑制。在糊粉层细胞中过表达这两个基因，会特异性且协同地抑制GA或GA诱导的转录激活因子GAMYB对ABA抑制型和GA诱导型Amy32bα-淀粉酶启动子报告构建体（Amy32b-GUS）的诱导。利用双分子荧光互补（BiFC）分析，在糊粉层细胞的细胞核中揭示了OsWRKY71蛋白自身以及OsWRKY71与OsWRKY51之间的物理相互作用。虽然OsWRKY51自身在体外不与Amy32b启动子结合，但它与OsWRKY71相互作用，并增强了OsWRKY71与Amy32b启动子中W盒的结合亲和力。通过缺失包含WRKY结构域的C末端或替换WRKY基序和锌指区域中的关键氨基酸，OsWRKY71的结合活性被消除。然而，这些非DNA结合突变体中的两个仍能够通过增强野生型DNA结合OsWRKY71阻遏物的结合亲和力来抑制GA诱导。相反，第三个非DNA结合突变体通过干扰野生型OsWRKY71或OsWRKY71/OsWRKY51阻遏复合物的结合，增强了Amy32b-GUS的GA诱导。这些数据证明了ABA诱导的WRKY基因在调节糊粉层细胞中GAMYB介导的GA信号转导中的协同相互作用，从而建立了一种ABA和GA信号转导相互作用的新机制。
