Improved lipid productivity in Nannochloropsis gaditana in nitrogen-replete conditions by selection of pale green mutants

Abstract

Background: Nannochloropsis gaditana is a photosynthetic unicellular microalgae considered one of the most interesting marine algae to produce biofuels and food additive due to its rapid growth rate and high lipid accumulation. Although microalgae are attractive platforms for solar energy bioconversion, the overall efficiency of photosynthesis is reduced due to the steep light gradient in photobioreactors. Moreover, accumulation of lipids in microalgae for biofuels production is usually induced in a two-phase cultivation process by nutrient starvation, with additional time and costs associated. In this work, a biotechnological approach was directed for the isolation of strains with improved light penetration in photobioreactor combined with increased lipids productivity. Results: Mutants of Nannochloropsis gaditana were obtained by chemical mutagenesis and screened for having both a reduced chlorophyll content per cell and increased affinity for Nile red, a fluorescent dye which binds to cellular lipid fraction. Accordingly, one mutant, called e8, was selected and characterized for having a 30% reduction of chlorophyll content per cell and an almost 80% increase of lipid productivity compared to WT in nutrient-replete conditions, with C16:0 and C18:0 fatty acids being more than doubled in the mutant. Whole-genome sequencing revealed mutations in 234 genes in e8 mutant among which there is a non-conservative mutation in the dgd1 synthase gene. This gene encodes for an enzyme involved in the biosynthesis of DGDG, one of the major lipids found in the thylakoid membrane and it is thus involved in chloroplast biogenesis. Lipid biosynthesis is strongly influenced by light availability in several microalgae species, including Nannochloropsis gaditana: reduced chlorophyll content per cell and more homogenous irradiance in photobioreactor is at the base for the increased lipid productivity observed in the e8 mutant. Conclusions: The results herein obtained presents a promising strategy to produce algal biomass enriched in lipid fraction to be used for biofuel and biodiesel production in a single cultivation process, without the additional complexity of the nutrient starvation phase. Genome sequencing and identification of the mutations introduced in e8 mutant suggest possible genes responsible for the observed phenotypes, identifying putative target for future complementation and biotechnological application.

Comparative Proteomics Reveals Evidence of Enhanced EPA Trafficking in a Mutant Strain of Nannochloropsis oculata

“…Identifying protein changes that contribute to membrane lipid composition remains challenging because a relatively small number of membrane proteins have been sequenced and studied in algae ( Garibay-Hernández et al, 2017 ). MGDG and DGDG are the major lipids of the photosynthetic membrane, where MGDG is synthesized in the chloroplast ( Dolch et al, 2017 ), and DGDG is synthesized in both chloroplast and ER ( Cecchin et al, 2020 ). The formation of membrane polar lipids is intrinsically linked to photosynthesis, for example, there is evidence to suggest that the light-harvesting complex may stabilize the MGDG component in thylakoids membranes ( Han et al, 2017 ).…”

Overexpression of the Transcription Factor AtLEC1 Significantly Improved the Lipid Content of Chlorella ellipsoidea

“…Based on the transcriptome and lipidome of C. reinhardtii under heat stress, a phospholipase A2 homolog and the DAG acyltransferase gene DGTT1 were identified ( Légeret et al, 2016 ). Cecchin et al (2020) sequenced the nuclear and organelle genomes of C. vulgaris 211/11P by combining next-generation sequencing and optical mapping of isolated DNA molecules and identified 10,724 nuclear genes, 121 chloroplast genes and 48 mitochondrial genes. LEC1 is an important TF, and its function in microalgae has not been elucidated.…”

Random Mutagenesis as a Promising Tool for Microalgal Strain Improvement towards Industrial Production

“…mutants ( Table 1 ). A similar approach was also used for other species, such as Nannochloropsis gaditana, Nannochloropsis oceanica and C. reinhardtii ( Table S1 ) [ 132 , 133 , 134 ]. Furthermore, a FACS-based selection using BODIPY staining showed success in the isolation of high-fatty-acid-producing mutants of Microchloropsis salina (syn.…”