Plants use different signalling pathways to respond to external stimuli. Intracellular signalling via calcium-dependent protein kinases (CDPKs) or mitogen-activated protein kinases (MAPKs) present two major pathways that are widely used to react to a changing environment. Both CDPK and MAPK pathways are known to be involved in the signalling of abiotic and biotic stresses in animal, yeast and plant cells. Here, we show the essential function of the CDPK CPK3 (At4g23650) for salt stress acclimation in Arabidopsis thaliana, and test crosstalk between CPK3 and the major salt-stress activated MAPKs MPK4 and MPK6 in the salt stress response. CPK3 kinase activity was induced by salt and other stresses after transient overexpression in Arabidopsis protoplasts, but endogenous CPK3 appeared to be constitutively active in roots and leaves in a strictly Ca2+-dependent manner. cpk3 mutants show a salt-sensitive phenotype comparable with mutants in MAPK pathways. In contrast to animal cells, where crosstalk between Ca2+ and MAPK signalling is well established, CPK3 seems to act independently of those pathways. Salt-induced transcriptional induction of known salt stress-regulated and MAPK-dependent marker genes was not altered, whereas post-translational protein phosphorylation patterns from roots of wild type and cpk3 plants revealed clear differences. A significant portion of CPK3 was found to be associated with the plasma membrane and the vacuole, both depending on its N-terminal myristoylation. An initial proteomic study led to the identification of 28 potential CPK3 targets, predominantly membrane-associated proteins.

Starch-rich crops form the basis of our nutrition, but plants have still to yield all their secrets as to how they make this vital substance. Great progress has been made by studying both crop and model systems, and we approach the point of knowing the enzymatic machinery responsible for creating the massive, insoluble starch granules found in plant tissues. Here, we summarize our current understanding of these biosynthetic enzymes, highlighting recent progress in elucidating their specific functions. Yet, in many ways we have only scratched the surface: much uncertainty remains about how these components function together and are controlled. We flag-up recent observations suggesting a significant degree of flexibility during the synthesis of starch and that previously unsuspected non-enzymatic proteins may have a role. We conclude that starch research is not yet a mature subject and that novel experimental and theoretical approaches will be important to advance the field.

14-3-3 proteins play an important role in the regulation of many cellular processes. The Arabidopsis vacuolar two-pore K(+) channel 1 (TPK1) interacts with the 14-3-3 protein GRF6 (GF14-λ). Upon phosphorylation of the putative binding motif in the N-terminus of TPK1, GRF6 binds to TPK1 and activates the potassium channel. In order to gain a deeper understanding of this 14-3-3-mediated signal transduction, we set out to identify the respective kinases, which regulate the phosphorylation status of the 14-3-3 binding motif in TPK1. Here, we report that the calcium-dependent protein kinases (CDPKs) can phosphorylate and thereby activate the 14-3-3 binding motif in TPK1. Focusing on the stress-activated kinase CPK3, we visualized direct and specific interaction of TPK1 with the kinase at the tonoplast in vivo. In line with its proposed role in K(+) homeostasis, TPK1 phosphorylation was found to be induced by salt stress in planta, and both cpk3 and tpk1 mutants displayed salt-sensitive phenotypes. Molecular modeling of the TPK1-CPK3 interaction domain provided mechanistic insights into TPK1 stress-regulated phosphorylation responses and pinpointed two arginine residues in the N-terminal 14-3-3 binding motif in TPK1 critical for kinase interaction. Taken together, our studies provide evidence for an essential role of the vacuolar potassium channel TPK1 in salt-stress adaptation as a target of calcium-regulated stress signaling pathways involving Ca(2+), Ca(2+)-dependent kinases, and 14-3-3 proteins.