Arabidopsis ABA-activated kinase MAPKKK18 is regulated by protein phosphatase 2C ABI1 and the ubiquitin proteasome pathway

Abstract

Phosphorylation and dephosphorylation events play an important role in the transmission of the abscisic acid signal. Although SnRK2 protein kinases and group A PP2C-type phosphatases constitute a core ABA pathway, little is known about the interplay between MAP kinases and protein phosphatases 2C in the regulation of ABA pathways. In this study, an effort was made to elucidate the role of MKKK18 in relation to ABA signaling and response. The MKKK18 knockout lines showed more vigorous root growth, decreased abaxial stomatal index, and increased stomatal aperture under normal growth conditions, compared to the control WT Col-0 line. In addition to transcriptional regulation of the MKKK18 promoter by ABA, we demonstrated by using in vitro and in vivo kinase assays that the kinase activity of MKKK18 was regulated by ABA and in response to H2O2 treatment. Analysis of the cellular localization of MKKK18 showed that the active kinase was targeted specifically to the nucleus. Notably, we identified ABI1 PP2C as a MKKK18-interacting protein, and demonstrated that ABI1 inhibited its activity. Using a cell-free degradation assay, we also established that MKKK18 was unstable and was degraded by the proteasome pathway. The rate of MKKK18 degradation was delayed in the ABI1 knockout line. Overall, we provide evidence that ABI1 regulates activity and promotes proteasomal degradation of MKKK18.