Identification and analysis of genes involved in tetrapyrrole metabolism and plastid-derived ROS-mediated signaling in Chlamydomonas reinhardtii

• Applicant: Professor Dr. Bernhard Grimm
• Subject Area: Plant Physiology
• Term: from 2016 to 2022
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 290763515

Because of the ability to synthesize chlorophyll in the dark, the green alga Chlamydomonas reinhardtii evolved a number of diverse regulatory mechanisms of tetrapyrrole biosynthesis (TBS), which may not be present or are difficult to study in angiosperms, e.g. due to the photosensitizing effect of TBS intermediates in light, although a great majority of other features discovered so far is similar. Additionally, the ability of C. reinhardtii to grow heterotrophically gives an opportunity to explore TBS or photosynthetic processes, which are not possible to study in plants, because their pigment biosynthesis mutants would be lethal. Therefore, C. reinhardtii presents an appropriate model to study TBS, retrograde signaling and related processes. We intend to take advantage of these characteristics of C. reinhardtii and apply different genetic and biochemical approaches (i) to analyze a specific set of mutants available in our group and a newly generated collection of insertional mutants with a high potential of genetic lesions in genes involved in regulation and catalysis of TBS, and (ii) to examine a possible cross-talk of different singlet-oxygen-mediated retrograde signaling pathways related to TBS. The mutants preselected in the search for TBS mutants will be examined for their levels of intermediates and endproducts of TBS, which are strong indication for impaired TBS. It is expected that novel genes can be identified, which encode either new subunits of TBS enzymes or regulatory factors which may show diversified algal control mechanisms of TBS at the transcriptional and posttranslational level in comparison to their homologous counterparts in the plants. The cause of a given phenotype resulting from mutagenesis will be confirmed by identification of the insertion site and the affected genes, followed by the rescue attempt with the wild-type copy of a given gene. The function of the encoded proteins in TBS and photosynthesis will be elucidated by biochemical and genetic methods. Subsequently, the impact of deregulated expression of these newly identified proteins will be explored on the entire pathway at the transcriptional and metabolic level. For the second part of the proposal, the strategy proposed to search for 1O2-inducible signaling components finally includes several steps in a workflow that ends with the screen for insertion mutants by means of a selected promoter-reporter constructs, which is induced by 1O2-accumulation.

• DFG Programme: Research Grants