A single nucleotide mutation in BrECB2 impaired RNA editing efficiency and early chloroplast biosynthesis in pak choi

Chloroplasts are important organelles for photosynthesis, which is essential for increasing the yields of pak choi. In this study, we evaluated a delayed chloroplast development mutant ‘M136’ identified during self-crossing of the pak choi inbred line ‘136’. The young true leaves of ‘M136’ were yellow and gradually turned green with maturation. Chloroplast development, pigment contents, and photosynthetic parameters were impaired and gradually recovered with growth in ‘M136’, and chlorophyll fluorescence parameters were also impaired in ‘M136’. Genetic analysis and bulk segregant analysis (BSA)-seq revealed that the mutant phenotype was controlled by a single recessive gene, identified as BraA06g011520.3.5C (BrECB2), which encodes a DYW-type pentatricopeptide repeat (PPR) protein. In ‘M136’, a T-to-C single nucleotide polymorphism (SNP) in the 4th PPR motif of BrECB2 caused a threonine-to-isoleucine amino acid substitution. BrECB2 was mainly expressed in young leaves. The chloroplast RNA editing efficiency of ‘M136’ was affected and significantly recovered after the leaves turned green, and the editing efficiency was partially restored in complementation lines. The plastid-encoded RNA polymerase activity was not affected in ‘M136’. Functional complementation analyses revealed that the transient overexpression of BrECB2 partially rescued the mutant phenotype and the RNA editing efficiency in ‘M136’. In summary, this study indicates that BrECB2 is involved in early chloroplast development and RNA editing, providing a theoretical basis for understanding the regulatory network involved in chloroplast development in pak choi.