Exploiting the microalga Nannochloropsis oceanica for biotechnological use

Over the last two decades, research on microalgae has experienced a spectacular upsurge due to the organisms’ high growth rates, undemanding cultivation and interesting cellular composition, making them an ideal sustainable resource. However, despite great advances, microalgal productivities and their exploitation as cell factories still lag behind our aspirations. In this work, we aimed to improve the applicability of species from the microalgal genus Nannochloropsis, which are attractive for industrial purposes due to their cellular lipid profile and robust growth. In addition, Nannochloropsis has an intriguing evolutionary history as secondary endosymbiont and presents an unusual pigment composition, making it an interesting organism for the study of photosynthesis. Chapter 1 entails a general review over the fascinating world of (micro-) algae and their use by humans with a focus on the genus of Nannochloropsis. Chapter 2 presents a study on the regulation of photosynthesis and photoprotection of Nannochloropsis oceanica by modulating its XC through the overexpression of ZEP1, VDE, or both enzymes simultaneously. The accumulation of ZEP1 was shown to reduce XC and NPQ activation and accelerate their relaxation. These modifications made the algal cells more sensitive to chemically induced ROS and light excess. The overexpression of VDE resulted in the enhanced activation of the XC and NPQ, but also slowed down NPQ relaxation, yet, it resulted in improved survival under high light. The concurrent accumulation of both enzymes caused faster XC and NPQ activation and relaxation. In sum, this study reveals that the content of VDE and ZEP1 are the rate-limiting factors of the XC and highlights the significance of this cycle to balance photosynthesis and protection in Nannochloropsis. Chapter 3 puts forward a detailed analysis of the growth performance and adaptation of the N. oceanica lines with a differently modulated XC generated in chapter 2. In tubular PBRs, the overall acceleration of xantcycle kinetics resulted in improved growth. Moreover, VDE accumulation expanded the alga’s tolerance to high light. These phenotypes were confirmed in experiments in flat-panel PBRs. Chapter 4 closer explores the enzymatic mechanism of the xanthophyll cycle. From analyses of the xanthophyll pool of ZEP1-/VDE-accumulating cells during light and dark phases in chapter 2, we generated a computational model of the XC that respects biochemical enzyme properties. By testing different mechanistic possibilities, we showed that the three xanthophylls in the cycle are likely interconverted by VDE and ZEP1 in a stepwise manner with the intermediate release of antheraxanthin. In addition, we determined the activity of ZEP1 after different illumination periods and found that the enzyme is downregulated under light. Chapter 5 outlines the investigation of protein secretion in N. oceanica. Using a homology- based approach, we could determine three signal peptides that can target proteins for secretion. The characterization of the secretory efficiency under different growth conditions showed that it is highest under optimal light and temperature and ample CO2 supply. The investigation of the exportation pathway by confocal imaging disclosed that secretory proteins are presumably translocated to the chloroplast membrane system of N. oceanica and subsequently trafficked into the periplasmic space between the cell membrane and the cell wall. Taken together, this thesis investigates physiological and photosynthetic features of the microalga Nannochloropsis that can serve as leverage points for strain improvement. At that, it illustrates the interconnectedness of fundamental research as a basis for biotechnological application.