The sugar transporter STP1-driven trophic conversion of Chlamydomonas reinhardtii.

backgroundChlamydomonas reinhardtii is a versatile model microalga with strong potential for industrial applications in light of its rapid growth, ability to synthesize valuable metabolites, and GRAS status. However, large-scale phototrophic cultivation faces challenges such as self-shading, uneven illumination, as well as the prerequisite of lighting equipment, all of which limit productivity and economic feasibility. Glucose-based heterotrophic cultivation is a scalable alternative, yet C. reinhardtii lacks endogenous glucose transporters, which inherently precludes its glucose uptake.

resultsTo overcome this limitation and expand its trophic flexibility, we engineered C. reinhardtii by introducing the STP1 glucose transporter originating from Arabidopsis thaliana, which confers the ability to uptake glucose. Along with the expression of STP1 in C. reinhardtii, two previously studied glucose transporters were also expressed individually for comparison: GLUT1 from Homo sapiens and HUP1 from Parachlorella kessleri because both are known to enable glucose uptake in C. reinhardtii. Resultingly, all transporter-expressing mutants acquired the ability to uptake glucose. Among tested, the STP1-expressing strain (3S7) exhibited the highest growth under glucose-supplemented heterotrophic conditions despite the marginal difference. The cell number of the 3S7 strain was 3.56 times higher than that of the wild-type (WT) strain, while the cell numbers of the GLUT1- and HUP1-expressing strains were 2.84 and 2.79 times higher, respectively. Dry cell mass analysis confirmed the glucose-based heterotrophic growth of the STP1-expressing strain, showing a 2.03-fold increase compared to the WT, whose weight is the highest among the transformants. Assays using the fluorescent glucose analog 2-NBDG and the non-metabolizable glucose analog 2-DG validated glucose transport into the 3S7 strain and others. The reduced residual glucose in the culture supernatant also supported the 3S7 strain’s primary ability to uptake glucose compared to the other strains, while exhibiting the biomass yield on glucose of 18.9 ± 8.6%.

conclusionsThese results demonstrate that STP1 can be exploited as a promising glucose importer, conferring glucose uptake ability to C. reinhardtii. This study establishes a foundation for increasing its trophic flexibility by broadening its usable organic carbon source, opening up a new opportunity to use the model alga as a cell factory platform for scalable industrial bioprocesses.