No abstract

Modulation of photoassimilate export from the chloroplast is essential for controlling the distribution of fixed carbon in the cell and maintaining optimum photosynthetic rates. In this study we identified chloroplast TRIOSE PHOSPHATE/PHOSPHATE TRANSLOCATOR2 (CreTPT2) and CreTPT3 in the green alga Chlamydomonas (Chlamydomonas reinhardtii), which exhibit similar substrate specificities but whose encoding genes are differentially expressed over the diurnal cycle. We focused mostly on CreTPT3 because of its high level of expression and the severe phenotype exhibited by tpt3 relative to tpt2 mutants. Null mutants for CreTPT3 had a pleiotropic phenotype that affected growth, photosynthetic activities, metabolite profiles, carbon partitioning, and organelle-specific accumulation of H2O2. These analyses demonstrated that CreTPT3 is a dominant conduit on the chloroplast envelope for the transport of photoassimilates. In addition, CreTPT3 can serve as a safety valve that moves excess reductant out of the chloroplast and appears to be essential for preventing cells from experiencing oxidative stress and accumulating reactive oxygen species, even under low/moderate light intensities. Finally, our studies indicate subfunctionalization of the CreTPT transporters and suggest that there are differences in managing the export of photoassimilates from the chloroplasts of Chlamydomonas and vascular plants.

Photosynthetic eukaryotic organisms contain several chloroplast-associated metabolite transporters that enable energetic/metabolic exchange between the chloroplast and other cellular compartments. In this study, we used the model photosynthetic alga Chlamydomonas reinhardtii to investigate a highly expressed chloroplast triose phosphate transporter. The triose phosphate/phosphate translocator 3 (CreTPT3), located on the Chlamydomonas chloroplast envelope, was found to be highly expressed under both non-stressed/stressed conditions (RNA level) and was characterized for substrate specificity in vitro using a yeast liposome uptake system. The CreTPT3 transporter showed high DHAP and 3-PGA transport activities, but little activity with PEP. Null mutants for CreTPT3, generated by CRISPR-Cas9 editing of the CreTPT3 gene, resulted in a pleiotropic phenotype impacting photosynthetic activity, metabolite pools, carbon partitioning, and storage, the redox status of the chloroplast, and the accumulation of reactive oxygen species. The results presented demonstrate that CreTPT3 is a major conduit on the chloroplast envelope for the intracellular distribution of fixed carbon and reductant generated by photosynthetic electron transport. Its function is critical for optimizing the use of resources supporting cell fitness, especially as light intensities increase, the rate of photosynthetic CO2 fixation is elevated and the chloroplast environment becomes highly reducing.