OsWHY1 Interacts with OsTRX z and is Essential for Early Chloroplast Development in Rice.

WHIRLY (WHY) family proteins, a small family of single-stranded DNA (ssDNA) binding proteins, are widely found in plants and have multiple functions to regulate plant growth and development. However, WHY in rice has received less attention. In this study, we continued our previous study on OsTRX z that is important for chloroplast development. OsTRX z was discovered to interact with OsWHY1, which was confirmed using yeast two-hybrid, pull-down, and BiFC assays. Subsequently, the oswhy1 mutants were obtained by CRISPR/Cas9, which exhibited an albino phenotype and died after the three-leaf stage. Consistent with this albino phenotype, low amounts of Chl a, Chl b, and Car were detected in the oswhy1-1 mutant. Moreover, the oswhy1-1 mutant had chloroplasts with disrupted architecture and no stacked grana and thylakoid membranes. Subcellular localization showed that the OsWHY1-GFP fusion protein was targeted to the chloroplast. What's more, OsWHY1 was found to be preferentially expressed in young leaves and was involved in chloroplast RNA editing and splicing. Mutation of OsWHY1 significantly affected the expression of chloroplast and ribosome development-related and chlorophyll synthesis-related genes. In conclusion, OsWHY1 contributes to early chloroplast development and normal seedling survival in rice. These results will further elucidate the molecular mechanism of chloroplast development and expand our understanding of WHY1 functions.