Requirement and functional redundancy of two large ribonucleotide reductase subunit genes for cell cycle, chloroplast biogenesis in tomato

Ribonucleotide reductase (RNR), functioning in the de novo synthesis of dNTPs, is crucial for DNA replication and cell cycle progression. However, the knowledge about the RNR in plants is still limited. In this study, we isolated ylc1 ( y oung l eaf c hlorosis 1 ) mutant, which exhibited many development defects such as dwarf stature, chlorotic young leaf, and smaller fruits. Map-based cloning, complementation, and knocking-out experiments confirmed that YLC1 encodes a large subunit of RNR (SlRNRL1), an enzyme involved in the de novo biosynthesis of dNTPs. Physiological and transcriptomic analyses indicate that SlRNRL1 plays a crucial role in the regulation of cell cycle, chloroplast biogenesis, and photosynthesis in tomato. In addition, we knocked out SlRNRL2 (a SlRNRL1 homolog) using CRISPR-Cas9 technology in the tomato genome, and found that SlRNRL2, possessing a redundant function with SlRNRL1, played a weak role in the formation of RNR complex due to its low expression intensity. Genetic analysis reveals that SlRNRL1 and SlRNRL2 are essential for tomato growth and development as the double mutant slrnrl1slrnrl2 is lethal. This also implies that the de novo synthesis of dNTPs is required for seed development in tomato. Overall, our results provide a new insight for understanding the SlRNRL1 and SlRNRL2 functions and the mechanism of de novo biosynthesis of dNTPs in plants.