Functional Genomics of Protein Phosphorylation in Arabidopsis Chloroplasts

点击翻译按钮获取中文摘要

Protein phosphorylation is a major mode of regulation of metabolism, gene expression, and cell architecture. Chloroplast or mitochondrial localization is predicted or has been found for 152 protein kinases (OPKs, organellar protein kinases) in Arabidopsis thaliana. The goal of this project is to define systematically targets and physiological role of phosphorylation mediated by OPKs by a combined approach, including (i) intracellular localization of the 50 OPKs with disputable organelle targeting by transient expression of RFP-fusions, (ii) mRNA expression profiling of OPKs for a number of organelle-relevant conditions, (iii) identification of knock-out mutants for 75 OPKs and their characterization with respect to key organellar functions in a gauntlet-type approach, (iv) identification of substrates of OPKs by differential proteomics (2-D gel analyses of 32P-labelled organellar proteins combined with mass spectrometry for wild-type [WT] versus OPK knock-outs). (v) For selected OPKs, transgenic plants (overexpressors and/or gain-offunction mutants) will be generated, and interactors will be identified by systematic two-hybrid screens.