Our Model System
is the unicellular green alga Chlamydomonas reinhardtii (Chlamy). Chlamy has been used since many decades as a model system to understand various aspects of cell and molecular biology. All three genomes (nuclear, chloroplast and mitochondrial) are sequenced and can be transformed. Tools for genome engineering (CRISPR/Cpf1) and synthetic biology are established and a library of ~60.000 indexed mutants is available. The organism has a generation time of ~8 h and can be grown on simple media to high densities.
Our high-tech toys
Of course we have all standard equipment available for molecular biology and protein biochemistry work. On top of that, we have some high-tech toys, including two Biostream photobioreactors. These allow the controlled growth of microalgal cultures in turbidostat mode over weeks. We can apply various light, temperature, and CO2 regimes and monitor pH, dissolved O2, and O2/CO2 in the exhaust gas. Moreover, we have an ABSciex Triple-TOF 6600 mass spectrometer coupled to an ekspert nanoLC 425, which we use primarily for the quantitative analysis of proteome dynamics and protein-protein interactions (see Center for MS Analytics).
I. The chloroplast as a hub for acclimation (Within TRR 175 and SPP1927)
All organisms face changes in environmental conditions. This is particularly challenging for plants which as sessile organisms cannot escape environmental changes. Within the TRR 175 we investigate the role of the chloroplast as a hub for the sensing and signalling of changes in temperature and light. In this context, we investigate how changes in temperature and light affect chloroplast protein homeostasis. Specifically, we ask how challenges of chloroplast protein homeostasis are sensed and signalled to the nucleus and what kind of responses are implemented to ensure homeostasis.
II. The role of VIPP1/2 in thylakoid membrane biogenesis (Within FOR2092)
Vesicle-inducing proteins in plastids (VIPPs) have been shown to be essential for the biogenesis of thylakoid membranes, as in vipp knockout mutants no thylakoid membranes are formed. We hypothesize that VIPPs play a role in organizing lipid microdomains in thylakoid membranes similar to the eisosomes in yeast plasma membranes. Eisosomes provide a specific lipid environment that is required for the proper functioning of membrane transporters. In thylakoid membranes, such a specific lipid environment might be important for the proper function of the Sec and TAT translocases, the Alb3 integrase as well as for other transporters in the thylakoid and envelope membranes. The goal of this project is to test this hypothesis.
III. Identification and characterization of factors involved in PSII biogenesis and repair (Within BioComp)
While the function of photosystem (PS)II is rather well understood, only few of the protein factors that drive its biosynthesis and assembly have been identified. The ability of PSII to oxidize water comes along with its vulnerability to light-induced damage, i.e. the D1 core subunit, requiring that PSII is continuously being repaired. Also PSII repair requires a plethora of protein factors that drive the disassembly of PSII (super)complexes, the targeted degradation of damaged D1, its replacement by de novo synthesized D1, and the reassembly of the PSII (super)complexes. In this project we wish to improve our understanding of how PSII is assembled during de novo biogenesis and how it is repaired after high light exposure.
IV. Dissection of the heat stress response in Chlamydomonas
The insufficient ability of many crop plants to acclimate to severe heat stress has a significant impact on crop yield safety. This problem might be solved by transgenic approaches, given that the mechanisms underlying heat stress acclimation responses are understood. Chlamy bears several advantages that make it easier to study fundamental aspects of the plant heat stress in this microalga rather than in land plants. First, gene families of proteins involved in the heat stress response (e.g. heat shock factors and molecular chaperones) are much smaller in Chlamy than in land plants. Second, Chlamy can be grown for weeks under highly controlled conditions in photobioreactors, facilitating competition experiments with mutant collections. The goal of this project is to understand the fundamental principles of the heat stress response in plant systems.
V. Investigation of the mechanisms by which the HSP70A promoter activates transgene expression
Transgene silencing is frequently observed in eukaryotic systems and may be mediated by epigenetic mechanisms. This is particularly the case in Chlamy and therefore epigenetic (trans)gene silencing is easy to study in this organism. Approaches undertaken so far aimed only at the identification of factors involved in transgene silencing, whereas no information exists on factors involved in transgene activation. We have found that transgene expression in Chlamy becomes activated when the promoter of the HSP70A gene is fused upstream of transgene-driving promoters. The mechanism underlying HSP70A promoter-mediated transgene activation might be applied to generally facilitate transgenic approaches in eukaryotes. Hence, the goal of this project is to understand the molecular mechanisms by which transgene silencing in Chlamy (and other organisms?) may be overcome.
VI. Synthetic biology combined with systems biology for microalgal biotechnology
The use of microalgae for biotechnology has become routine in recent years. This because they are carbon negative, grow rapidly to high densities on cheap media, are completely safe for human health, and are amenable to genetic engineering. Recent developments for Chlamy, like the establishment of robust genome engineering protocols, expression strains, insertional mutant libraries and a MoClo-based toolkit comprising 119 genetic modules, has widely opened the doors for microalgal biotechnology. In this project we engineer Chlamy for the production and secretion of biotechnologically important proteins (see also our iGEM team).