Systematically programmed adaptive evolution revealed the potential role of carbon and nitrogen pathways during lipid accumulation in Chlamydomonas reinhardtii.

Background The concept of adaptive evolution implies underlying genetic mutations conferring a selective advantage to an organism under particular environmental conditions. Thus, a flow cytometry-based strategy was used to study the adaptive evolution in Chlamydomonas reinhardtii wild type CC124 and a starchless mutant sta6-1 cells, with respect to lipid metabolism under nitrogen (N) deplete and replete conditions. Results The successive sorting and regeneration of top 25000 high-lipid content cells of CC124 and sta6-1 combined with nitrogen starvation led to the generation of a new population with improved lipid content when compared to the original populations, ~175% and ~50% lipid increase in sta6-1 and CC124, respectively. During adaptive evolution period, the major fatty acid components observed in cells were C16:0, C16:1, C18:0, and C18:1-3, and elemental analysis revealed that cellular carbon to nitrogen ratio increased at the end of adaptive evolution period. In order to gain an insight into highly stimulated intracellular lipid accumulation in CC124 and sta6-1 resulting from the adaptive evolution, proteomics analyses of newly generated artificial high-lipid content populations were performed. Functional classifications showed the heightened regulation of the major chlorophyll enzymes and the enzymes involved in carbon fixation and uptake including chlorophyll-ab-binding proteins, and Rubisco activase. The key control protein (periplasmic L-amino acid oxidase (LAO1)) of carbon-nitrogen integration was specifically over-expressed. Glutathione-S-transferases and esterase, the enzymes involved in lipid-metabolism and lipid-body associated proteins, were also induced during adaptive evolution. Conclusions Adaptive evolution results demonstrate the potential role of photosynthesis in terms of carbon partitioning/flux/fixation and carbon/nitrogen metabolism during lipid accumulation in microalgae and this strategy can be a new tool to develop C. reinhardtii strains and other microalgal strains with desired phenotypes such as high lipid accumulation. [ABSTRACT FROM AUTHOR]