Identifying the molecular transducers of retrograde redox signaling and unravelling compartmental redox dynamics in the diatom Phaeodactylum tricornutum

• Applicant: Professor Dr. Bernard Lepetit
• Subject Area: Plant Physiology
• Term: since 2022
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 515451811

Diatoms are microalgae that belong to the most important primary producers on earth. They contain so-called complex plastids acquired by secondary endosymbiosis from a red alga. Consequently, their plastid envelope is composed of four membranes, instead of two in green algae and vascular plants. A decade ago, we identified the existence of light dependent retrograde redox signaling in the diatom Phaeodactylum tricornutum, i.e. the regulation of nuclear gene expression by a stimulus originating from the plastid and induced by changes in light intensity. We identified the redox state of the plastoquinone pool (PQRS) to be at the onset of this signaling chain. Since then, no further details regarding retrograde redox signaling in diatoms have been published. Moreover, the dynamics of other redox components such as the pH, ATP and H2O2 - all directly dependent on light conditions and potentially being influenced by and influencing themselves retrograde signaling - are completely unknown in diatoms. In this project, I aim to reveal molecular components involved in retrograde redox signaling in P. tricornutum and thus obtain first insights in the signaling chain originating from the plastid and ending in the nucleus (Task 1). I also aim to elucidate the compartmental redox dynamics of pH, ATP and H2O2 depending on light intensity, with a specific emphasis on the impact of the photosynthetic light reaction and its interplay with mitochondrial respiration (Task 2). Together with the dynamic regulation of the photosynthetic light reaction, which I am working on in other projects, the results of this project will enable me to understand photophysiology in a much more holistic context and allow me establishing new models regarding light acclimation in diatoms but also beyond in other taxa. To set the stage for these objectives, we have done extensive pre-work in the past. We have performed multiple transcriptomic analyses under changing light conditions and by manipulating the photosynthetic electron transfer chain. Out of four major groups of genes regulated by different triggers identified this way, I selected two groups, namely genes being regulated i) by the PQRS, and ii) by a signal generated at the acceptor side of photosystem I, likely H2O2. In Task 1, for six promoters of these genes, respectively, the respective binding transcription factors will be identified using yeast-1-hybrid assays. These factors will then be knocked out in P. tricornutum by CRISPR-Cas and their impact on retrograde signaling will be verified afterwards. We also established fluorescent proteins sensors for pH, ATP and H2O2 in different compartments in P. tricornutum. In Task 2, we will produce additional sensor lines targeting all relevant compartments and afterwards elucidate the compartmental redox dynamics under changing light conditions without and with photosynthetic and respiratory impairments induced by various pharmacological treatments.

• DFG Programme: Research Grants
• International Connection: Belgium
• Cooperation Partner: Professor Dr. Lieven De Veylder