Identification of a chloroplast fatty acid exporter protein, CmFAX1, and triacylglycerol accumulation by its overexpression in the unicellular red alga Cyanidioschyzon merolae

Triacylglycerols (TAGs) produced by microalgae are a promising biofuel resource, but low productivity is still a problem for practical production. Understanding the underlying molecular mechanisms of TAG production is essential to improve productivity, which remains mostly in a primitive stage. Recently, a nuclear gene encoding a chloroplast inner membrane protein, AtFAX1, which is involved in the export of fatty acids (FAs) from the chloroplast, was identified and analyzed in Arabidopsis thaliana. In this study, we identified and analyzed a nuclear-encoded homolog of AtFAX1, named CmFAX1, from the unicellular red alga Cyanidioschyzon merolae. CmFAX1 was localized around the chloroplast envelope membrane in C. merolae, and heterologous expression in budding yeast suggested a role in FA traffic through membranes. Overexpression of CmFAX1 in C. merolae cells resulted in a 0.4-fold decrease in intracellular free FAs and a 2.4-fold increase in intracellular TAGs under normal growth conditions, suggesting that export of the free FAs produced in the chloroplast was enhanced and they were consumed for TAG synthesis in the cytosol. The accumulated TAGs in the CmFAX1 overexpression strain included short chain FAs, 14:0 and 14:1, which were not observed in the TAGs of the control strain and are suitable for biojet/biodiesel production because of their low cloud points. Overproduction of CmFAX1 resulted in higher cell yields after exponential growth phase culture. This was probably because of decreased photo-synthetic pigments (chlorophyll a) and increased light permeability into the highly dense cell culture, which are beneficial characteristics for increased biomass production. These results indicate that CmFAX1 could be a useful target protein for improvement of algal biofuel production.