B2/3-RAF mediates SnRK2 activation in ABA signaling in Arabidopsis

Z. Lin,  Y. Li,  Y. Wang,  P. Wang

Institute of Advanced Biotechnology and School of Life Sciences, Southern University of Science and Technology

The phytohormone abscisic acid (ABA) is crucial for plant responses to environmental challenges. The SNF1-regulated protein kinase 2s (SnRK2s) are key components in ABA-receptor coupled core signaling. It is rapidly phosphorylated and activated by ABA. Early work proved SnRK2 could activate itself by auto-phosphorylation, after ABA mediates SnRK2 release from its inhibitor complex PP2C-SnRK2. Recent studies have suggested that Raf-like protein kinases (RAFs) could phosphorylate SnRK2 and participate in ABA-triggered SnRK2 activation. Thus, how SnRK2 kinases are quickly activated during ABA signaling still needs to be clarified. Here, we used ATP analog and corresponding mutated SnRK2 to label the phosphorylation on SnRK2. The result showed that both B2/3 RAFs directly phosphorylate SnRK2.6 in the kinase activation loop. This trans-phosphorylation by RAFs stimulated SnRK2 activation, and the activated SnRK2s then intermolecularly trans-phosphorylate other SnRK2s that are not yet activated. No RAF kinases will fail the initiate of SnRK2 activation, and genetic evidence has proved that high-order Arabidopsis mutants lacking multiple B2 and B3 RAFs show ABA hyposensitivity and SnRK2 inactivation. So, our findings revealed a unique initiation and amplification mechanism of SnRK2 activation by which RAF participate in ABA signaling.