Engineering cytosolic carbonic anhydrase to establish C4 photosynthesis in rice
In mesophyll cells, carbonic anhydrase is mainly located in the chloroplast, however it is in the cytosol in plants with a C4 carbon concentrating mechanism. There is interest in relocating carbonic anhydrase to the cytosol of C3 plants as a first step in the introduction of a carbon concentrating mechanism. Here, Jethva et al. used CRISPR/Cas9 to generate rice (Oryza sativa) plants with cytosolic-localized carbonic anhydrase. They generated a mutant with a frame shift that disrupts the chloroplast transit peptide, however, to ensure the rest of the protein is produced correctly, the reading frame was restored at the end of the chloroplast transit peptide sequence. Carbonic anhydrase activity assays revealed that the mutants have 17-30% of the carbonic anhydrase activity compared to wild type plants, but crucially all of this is in the cytosol, with no chloroplastic activity detected. Despite the reduction in enzymatic activity, the plants have no abnormal growth or yield phenotype. Hence, it is possible to relocate carbonic anhydrase to the cytosol of mesophyll cells in rice, which could be a key step for the introduction of a complete carbon concentration mechanism. (Summary by Rose McNelly @Rose_McN) bioRxiv https://doi.org/10.1101/2024.05.21.595093
