Targets of calcium dependent protein kinases in Arabidopsis

This project is based on the results of a previous FWF-funded project on "Calcium dependent protein kinases in Arabidopsis". Continuing the work started in that project we aim here at identification of molecular targets of Calcium dependent protein kinases (CPKs), and establishing their biological function in the regulation of plant development and stress response. Based on our present data we will focus mainly on CPK3 (At4g23650) and on posttranslational modification of Arabidopsis basic leucine zipper (bZIP) transcription factors of the subgroup C as putative CPK targets. We will analyze the impact of their phosphorylation on regulation of general metabolism and in response pathways triggered by environmental changes. Different approaches will be used to study transcription factor phosphorylation and functional consequences in close cooperation with the groups of Klaus Harter in Tübingen and Wolfgang Dröge-Laser in Göttingen. Functional studies on bZIP mediated gene expression, sugar signalling, and metabolic adjustments will be performed in collaboration with Sjef Smeekens in Utrecht. The general aim of our network proposal is to establish the role of selected protein kinases in regulation of plant metabolism in response to environmental changes.

Acquisition of energy and regulation of its use by different metabolic pathways is the fundamental basis for live on earth. Plants use photosynthesis to generate the energy rich compounds needed to sustain their growth and development thereby providing the basis for human nutrition. The modulation of their metabolism in response to changes in the environment is of fundamental importance for efficient growth and productivity. Central signal transduction pathways regulate different responses, which allow plants to survive unfavorable environmental conditions. This includes certain defense responses such as the regulation of ion transporters or the synthesis of small compounds acting as osmolytes under salt stress conditions, and also the regulation of their energy usage. These reactions are mediated by signaling pathways which involve different protein kinases acting as molecular switches in cellular metabolism by attaching a phosphate group to proteins. This could then lead to a different activity of the phosphorylated protein or a change in its localization within a cell. Many external stimuli do also generate calcium-signals in the cell, where a sudden increase in the concentration of free Ca2+ ions leads to the activation of calcium-dependent protein kinases (CDPKs). In this project we identified that such a CDPK is required for salt-stress acclimation in the model plant Arabidopsis thaliana. The kinase is localized in the nucleus and also at cellular membranes. Membrane-localization of the kinase was dependent on the attachment of myristic acid, a C14 fatty acid, at the N-terminus of the kinase. We could also show that this protein modification is also used to anchor several other CDPKs at cellular membranes (Benetka et al. 2008). The major function of the kinase during salt-stress acclimation seems to be the regulation of different membrane proteins, which is most likely required for adaptation of the ion homeostasis (Mehlmer et al. 2010). The function of the kinase in the nucleus remained open as we could not observe significant changes in gene expression. In addition we identified also targets of this kinase in the cytosol, which are involved in the regulation of root development in response to external stimuli (Rietz et al. 2010) or in the regulation of nitrogen assimilation (Lambeck et al. 2010). References: Benetka W, Mehlmer N, Sammer M, Neumüller R, Maurer-Stroh S, Koranda M, Knoblich J, Teige M, and Eisenhaber F. (2008) Experimental testing of predicted myristoylation targets involved in asymmetric cell division and calcium-dependent signalling. Cell Cycle 7: 3709-3719. Mehlmer N, Wurzinger B, Stael S, Hofman-Rodrigues D, Csaszar E, Pfister B, Bayer, R, and Teige M (2010) The Ca2+-dependent protein kinase CPK3 is required for MAPK-independent salt-stress acclimation in Arabidopsis. The Plant Journal 63: 484-498. Rietz S, Dermendjiev G, Oppermann E, Tafesse FG, Holk A, Parker JE, Teige M, and Scherer GE. (2010) Roles of Arabidopsis Patatin-Related Phospholipases A in Root Development Are Related to Auxin Responses and Phosphate Deficiency. Mol Plant 3:524-538. Lambeck I, Chi JC, Krizowski S, Mueller S, Mehlmer N, Teige M, Fischer K, and Schwarz G (2010) Kinetic analysis of 14-3-3 inhibited Arabidopsis thaliana nitrate reductase. Biochemistry, in press. PMID: 20690630