Abscisic acid induces rapid subnuclear reorganization in guard cells.

While the phytohormone abscisic acid (ABA) is well established as a regulator of gene transcription and ion channel activity (Rock, 2000; Assmann and Wang 2001; Schroeder et al., 2001; Finkelstein et al., 2002), recent identification of RNA-binding proteins ABH1 (ABA hypersensitive), SAD1 (supersensitive to ABA and drought), and HYL1 (hyponastic leaves), whose mutation confers an ABA-hypersensitive phenotype (Lu and Fedoroff, 2000; Hugouvieux et al., 2001, 2002; Xiong et al., 2001), suggests that ABA may also play important roles in posttranscriptional RNA processing. Here we show that ABA dynamically regulates subnuclear architecture in guard cells, promoting the partitioning of a heterogeneous nuclear ribonucleoprotein (hnRNP)-type protein, AKIP1, into discrete subnuclear bodies or speckles via a process that has both Ca-dependent and Ca-independent steps and requires an active transcriptional machinery. hnRNPs are mRNA-protein complex proteins (mRNP proteins) that bind RNA and affect its metabolism (Krecic and Swanson, 1999; Dreyfuss et al., 2002). Members of the hnRNP family are involved in all aspects of RNA metabolism, including pre-mRNA splicing, mRNA localization, mRNA stability, nuclear export of mRNA, and translational control (Dreyfuss et al., 1996, 2002; Krecic and Swanson, 1999; Mili et al., 2001; Reed and Magni, 2001). Binding of hnRNPs with nascent RNA transcripts results in the formation of ribonucleoprotein complexes (RNPs). RNPs are highly dynamic, and it has been demonstrated in mammalian cells that hnRNPs bind or dissociate from the target RNA at various stages of mRNA maturation until a distinct mRNP is formed and translocated from the nucleus to the cytoplasm for translation initiation (Dreyfuss et al., 2002). In plants, little is known about the function of hnRNPs (Albà and Pagès, 1998; Lorković et al., 2000; Lambermon et al., 2002; Lorković and Barta, 2002). We reported previously (Li et al., 2002) that AKIP1 is an hnRNP-like RNA binding protein from broad bean (Vicia faba). AKIP1 exhibits greatest sequence homology to mammalian hnRNP A/B and D proteins and has two RNA-recognition motifs. AKIP1 is phosphorylated by guard cell AAPK, an ABA-activated, Ca-independent protein kinase involved in ABA-regulation of stomatal closure and plasma membrane anion channel activity (Li and Assmann, 1996; Li et al., 2000). Upon phosphorylation, AKIP1 becomes competent to bind dehydrin mRNA, which encodes a class of stress-protectant proteins (Close, 1996, 1997). ABA treatment of guard cells expressing AKIP1-green fluorescent protein (GFP) also results in rapid subnuclear clustering of AKIP1-GFP. In this report, we examine the signal transduction machinery involved in this phenomenon.