Chlamydomonas Mutants Null for Chloroplast Triose Phosphate Transporter3 are Metabolically Compromised and Light Sensitive

Abstract

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.