ACCUMULATION OF PHOTOSYSTEM ONE2 plays an essential role in chloroplast intron splicing during chloroplast biogenesis

ACCUMULATION OF PHOTOSYSTEM ONE2 (APO2) is an essential protein for embryo development, yet its functional role remains largely unexplored. In this study, we generated inducible APO2 RNA interference (RNAi) lines to investigate the function of APO2. Upon induction, APO2-RNAi plants exhibited a yellow phenotype in newly emerging leaves, accompanied by defective chloroplast development. We demonstrated that APO2 localizes to chloroplasts and that APO2 is highly expressed in green leaves. Phylogenetic analysis of the APO protein family across land plants revealed that APO2 proteins cluster into a distinct clade separate from other APO family members. The splicing of several chloroplast introns was affected in APO2-RNAi-induced yellow leaves, with splicing efficiencies of the ycf3.1 and ndhA introns dramatically lower than those in green leaves. We determined that APO2 interacts with two splicing factors, chloroplast RNA splicing 2-associated Factor 1 (CAF1)/CAF2 and DEAD-box RNA helicase 3 (RH3), which are involved in group II intron splicing. Our results also revealed splicing deficiencies in both ycf3.1 and ndhA introns in RH3 cosuppression lines. Notably, both APO2 and RH3 directly bound the 5' region of ycf3.1. These results establish that APO2 is indispensable for chloroplast biogenesis, primarily due to its essential role in the splicing of chloroplast introns.