Analysis of RNA Recognition and Binding Characteristics of OsCPPR1 Protein in Rice

Abstract

Pentatricopeptide repeat (PPR) proteins represent one of the largest protein families in plants and typically localize to organelles like mitochondria and chloroplasts. By contrast, CYTOPLASM- LOCALIZED PPR1 (OsCPPR1) is a cytoplasm-localized PPR protein that can degrade OsGOLDEN- LIKE1 (OsGLK1) mRNA in the tapetum of rice anther. However, the mechanism, by which OsCPPR1 recognizes and binds to OsGLK1 transcripts, remains unknown. Through protein structure prediction and macromolecular docking experiments, we observed that distinct PPR motif structures of OsCPPR1 exhibited varying binding efficiencies to OsGLK1 RNA. Moreover, RNA-electrophoretic mobility shift assay experiment demonstrated that the recombinant OsCPPR1 can directly recognize and bind to OsGLK1 mRNA in vitro. This further confirmed that the mutations in the conserved amino acids in each PPR motif resulted in loss of activity, while truncation of OsCPPR1 decreased its binding efficiency. These findings collectively suggest that it may require some co-factors to assist in cleavage, a facet that warrants further exploration in subsequent studies.