FOR 964 - TP Z2: Central proteomic platform for the Research Unit

Basic data for this project

Description

In plant cells, the increase in cytoplasmic calcium Ca2+ represents a frequent response to various abiotic and biotic stresses. After changes in cytosolic Ca2+ signatures have been sensed, phoshorylation cascades appear to represent general downstream responses in Ca2+-dependent signal transduction. To elucidate Ca2+ dependent protein phosphorylation, identify Ca2+ dependent phosphoproteins, map phosphorylation sites and quantify phosphorylation, a phosphoproteomics approach will be applied. Mass spectrometric analysis will be performed from tryptic digests of whole root and leaf extracts from wild plants, expressing aeqourin, where a cytosolic calcium transient is induced by calmodulin antagonists and blocked by lanthanides. Changes in Ca2+ will be visualized by a calcium imaging system. We further aim to identify phosphorylation targets of Ca2+-dependent kinases CIPK23. Here we will also take advantage of cipk23 T-DNA insertion lines and will analyze the phosphoprotein status under potassium deficiency in wild-type and mutant plants. To absolutely quantitate the phosphorylation status of the phosphoproteins of interest, proteotypic peptide profiling will be performed. We envision that the outlined strategy will shed light on the role of Ca2+-dependent protein phosphorylation in Ca2+-dependent signal transduction pathways.