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

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.

Retraction Note: Increased biomass productivity in green algae by tuning non-photochemical quenching.

This paper has been retracted.

The function of LHCBM4/6/8 antenna proteins in Chlamydomonas reinhardtii.

In eukaryotic autotrophs, photosystems are composed of a core moiety, hosting charge separation and electron transport reactions, and an antenna system, enhancing light harvesting and photoprotection. In Chlamydomonas reinhardtii, the major antenna of PSII is a heterogeneous trimeric complex made up of LHCBM1-LHCBM9 subunits. Despite high similarity, specific functions have been reported for several members including LHCBM1, 2, 7, and 9. In this work, we analyzed the function of LHCBM4 and LHCBM6 gene products in vitro by synthesizing recombinant apoproteins from individual sequences and refolding them with pigments. Additionally, we characterized knock-down strains in vivo for LHCBM4/6/8 genes. We show that LHCBM4/6/8 subunits could be found as a component of PSII supercomplexes with different sizes, although the largest pool was free in the membranes and poorly connected to PSII. Impaired accumulation of LHCBM4/6/8 caused a decreased LHCII content per PSII and a reduction in the amplitude of state 1-state 2 transitions. In addition, the reduction of LHCBM4/6/8 subunits caused a significant reduction of the Non-photochemical quenching activity and in the level of photoprotection.

Increased biomass productivity in green algae by tuning non-photochemical quenching.

Photosynthetic microalgae have a high potential for the production of biofuels and highly valued metabolites. However, their current industrial exploitation is limited by a productivity in photobioreactors that is low compared to potential productivity. The high cell density and pigment content of the surface layers of photosynthetic microalgae result in absorption of excess photons and energy dissipation through non-photochemical quenching (NPQ). NPQ prevents photoinhibition, but its activation reduces the efficiency of photosynthetic energy conversion. In Chlamydomonas reinhardtii, NPQ is catalyzed by protein subunits encoded by three lhcsr (light harvesting complex stress related) genes. Here, we show that heat dissipation and biomass productivity depends on LHCSR protein accumulation. Indeed, algal strains lacking two lhcsr genes can grow in a wide range of light growth conditions without suffering from photoinhibition and are more productive than wild-type. Thus, the down-regulation of NPQ appears to be a suitable strategy for improving light use efficiency for biomass and biofuel production in microalgae.

Non-photochemical quenching and xanthophyll cycle activities in six green algal species suggest mechanistic differences in the process of excess energy dissipation.

In the present study the non-photochemical quenching (NPQ) of four biofilm-forming and two planktonic green algae was investigated by fluorescence measurements, determinations of the light-driven proton gradient and determination of the violaxanthin cycle activity by pigment analysis. It was observed that, despite the common need for efficient photoprotection, the structural basis of NPQ was heterogeneous in the different species. Three species, namely Chlorella saccharophila, Chlorella vulgaris and Bracteacoccus minor, exhibited a zeaxanthin-dependent NPQ, while in the three other species, Tetracystis aeria, Pedinomonas minor and Chlamydomonas reinhardtii violaxanthin de-epoxidation was absent or unrelated to the establishment of NPQ. Acclimation of the algae to high light conditions induced an increase of the NPQ activity, suggesting that a significant part of the overall NPQ was rather inducible than constitutively present in the green algae. Comparing the differences in the NPQ mechanisms with the phylogenetic position of the six algal species led to the conclusion that the NPQ heterogeneity observed in the present study was not related to the phylogeny of the algae but to the environmental selection pressure. Finally, the difference in the NPQ mechanisms in the different species is discussed within the frame of the current NPQ models.