Identification and characterisation of state transition mutants in Arabidopsis thaliana

Abstract

The regulation of optimal utilisation of absorbed light energy and avoidance of oxidative damage constitute a major problem for all oxygen evolving cells. Adaptations to transient changes are achieved by regulation of photochemistry: In order to balance the absorbed energy distribution between PS II and PS I redox-regulated phosphorylation of the LHC antenna results in reversible coupling with the photosystem cores known as LHC state transitions. So far neither the LHC-kinase and -phosphatase nor any redox sensors and regulators of a putative regulation cascade have been identified. In this context we established a molecular genetic approach, targeting nuclear encoded genes that are essential for functional state transitions. The approach includes the construction of tag-induced knock out-mutants of Chlamydomonas reinhardtii and, together with purchased random T-DNA- and UV-mutants from Arabidopsis thaliana, a screening step by fluorescence video imaging for clones that are defective in state transitions. For the subsequent identification of insert-flanking gene regions we established the procedure of ligation-mediated suppression PCR for Chlamydomonas. By doing so we identified several tagged genes of interest. The characterisation of their gene products made it feasible to suggest that some of them are linked to the regulation process of state transitions. Additionally, we did a systematic analysis of the fully sequenced Arabidopsis genome to identify possible LHC kinase candidates. As a result we localised a gene encoding for a potential thylakoid-associated kinase. A T-DNA mutant with a defect in this gene has been isolated from the T-DNA library in Madison (USA) for further analysis.