Abstract
SnRK2.6 (SUCROSE NONFERMENTING 1-RELATED PROTEIN KINASE2.6) has been characterized as a molecular switch for the intracellular abscisic acid (ABA) signal-transduction pathway. Normally, SnRK2.6 is kept in an "off" state, forming a binary complex with protein phosphatase type 2Cs (PP2Cs). Upon stressful conditions, SnRK2.6 turns into an "on" state by its release from PP2Cs and then phosphorylation at Ser175. However, how the "on" and "off" states for SnRK2.6 are fine-tuned, thereby controlling the initiation and braking processes of ABA signaling, is still largely unclear. SnRK2.6 activity was tightly regulated through protein post-translational modifications (PTM), such as persulfidation and phosphorylation. Taking advantage of molecular dynamics simulations, our results showed that Cys131/137 persulfidation on SnRK2.6 induces destabilized binding and weakened interactions between SnRK2.6 and HAB1 (HYPERSENSITIVE TO ABA1), an important PP2C family protein. This unfavorable effect on the association of the SnRK2.6-HAB1 complex suggests that persulfidation functions are a positive regulator of ABA signaling initiation. In addition, Ser267 phosphorylation in persulfidated SnRK2.6 renders a stable physical association between SnRK2.6 and HAB1, a key characterization for SnRK2.6 inhibition. Rather than Ser175, HAB1 cannot dephosphorylate Ser267 in SnRK2.6, which implies that the retained phosphorylation status of Ser267 could ensure that the activated SnRK2.6 reforms the binary complex to cease ABA signaling. Taken together, our findings expand current knowledge concerning the regulation of persulfidation and phosphorylation on the state transition of SnRK2.6 and provide insights into the fine-tuned mechanism of ABA signaling.