Functional and molecular characterization of maize open stomata 1 protein kinase

Plant growth and productivity are compromised by environmental stresses such as pathogens, extreme temperatures, drought and high salinity. Being sessile organisms, plants had to develop different physiologic and biochemical strategies to cope with these potential harmful situations. Drought in particular is one of the major environmental stresses that plants are forced to endure during their life cycle. The adaptation to water deficit is controlled by a cascade of molecular networks that start with the perception of water shortage which leads to increases in the ABA levels. Even though the ABA signalling model is well described for Arabidopsis, little is known for other plant species. With this thesis we proposed to increase the knowledge of maize response to drought, focusing on a maize kinase of the SnRK2 family ? ZmSnRK2.8/ZmOST1 ? which is highly homologous to the Arabidopsis OST1. We divided our work on three chapters, namely the biochemical characterization of ZmOST1, the functional characterization of ZmOST1 and the study of ZmOST1 regulation. 1) ZmOST1 biochemical characterization With Chapter 1 we characterized ZmOST1 at the biochemical level, making parallels with the Arabidopsis system whenever pertinent. We found a very close biochemical relationship between the maize and Arabidopsis kinases that suggests a conserved mechanism of plant responses to ABA and drought stress and point to the potential use of this kinase in improvement programs of drought tolerance in crops. 2) ZmOST1 functional characterization With Chapter 2 we described ZmOST1 as a functional kinase that is activated by different osmotic stresses and that is able to complement the Arabidopsis ost1? 2 mutant with effects on stomata closure. We also present a transcription factor of the NAC superfamily (ZmSNAC1) as a novel cognate substrate of ZmOST1. Under abiotic stresses ZmOST1 is capable of phosphorylating this transcription factor with further implications on stomata regulation. 3) ZmOST1 regulation With the results presented in Chapter 3 a larger picture of ABA signalling appears that implicates new partners on ZmOST1 regulation, specifically the CK2 kinase and the proteasome degradation. Ample evidence is shown suggesting CK2 phosphorylation is implicated in ABA signalling by affecting ZmOST1 localization, protein levels, protein degradation and interaction with PP2C phosphatases. At the plant level, overexpressing ZmOST1 mutagenized on the CK2 loci of phosphorylation grants several potential beneficial traits that could prove important for crop biotechnology, such as higher protein levels, better protein stability, enhanced phosphorylation activity and better stomata regulation. Working model: Taken the results presented in this thesis together, we propose a change in the current ABA signalling model. First we believe that there is an important role for CK2 in ABA sensing and SnRK2 activation that could affect the binding of the kinase to the PP2C phosphatises and regulate SnRK2 through degradation. Second, we propose that, apart from the always off and transiently fast on/off modes of SnRK2 activity, there is a third always on mechanism in which the kinase is fully detached from the phosphatase.