Prime editing of an evolutionarily informed PEPC residue enhances photosynthesis and grain nutrition in C 3 rice

Abstract C 4 photosynthesis evolved through adaptive changes in key metabolic enzymes that enhance carbon fixation efficiency, yet transferring such traits into C 3 crops has met limited success. Using precision prime editing, we introduced an evolutionarily conserved single amino acid substitution (Arg→Gly) into phosphoenolpyruvate carboxylase (PEPC) in rice. Edited plants exhibited enhanced PEPC activity in the presence of the feedback inhibitor malate, increased chlorophyll content, elevated photosynthetic rates under ambient CO 2 , and increased seed size and weight. Remarkably, seeds from the edited lines showed simultaneous enrichment of zinc (Zn), iron (Fe), and protein, hereafter referred to as ZiP rice. This work demonstrates that a single, evolutionarily guided residue change can reprogram core carbon metabolism in a C 3 crop, enabling concurrent improvements in photosynthesis and grain nutritional quality.