Receptor-like protein kinases RPK1 and BAK1 sequentially form complexes with the cytoplasmic kinase OST1 to regulate ABA-induced stomatal closure

Regulation of plant water status occurs via abscisic acid (ABA)-induced stomatal closure. Open Stomata 1 (OST1) is a critical ABA signaling component regulating this process in guard cells. We previously reported that BRI1-associated receptor kinase 1 (BAK1) positively regulates ABA-induced stomatal closure by interacting with and phosphorylating OST1. Here, using Arabidopsis, we show that the receptor-like protein kinase 1 (RPK1), previously known to be induced by ABA, is a positive ABA-signaling component in guard cell movement, and interacts with OST1. ABA-inducible expression patterns were observed in RPK1 and OST1, but not in BAK1. We investigated the underlying mechanisms by which the RPK1-OST1 and BAK1-OST1 complexes interact in stomatal guard cells by monitoring the complex formation continuously using fluorescence resonance energy transfer analyses. We found that the BAK1-OST1 complex was formed earlier than the RPK1-OST1 complex in response to ABA. In vitro and semi-in vivo kinase assays revealed that a transphosphorylation event occurred in the RPK1-OST1 complex, which differs from that in the BAK1-OST1 complex, wherein only OST1 phosphorylation occurred via BAK1. ABA-insensitive 1 (ABI1) only dephosphorylated OST1 in the BAK1-OST1 complex, but dephosphorylated both RPK1 and OST1 proteins in the RPK1-OST1 complex. Our results suggest that there are multiple coordinated ABA signaling systems to regulate stomatal movement.