The Pentatricopeptide Repeat Protein PGR3 Is Required for the Translation of petL and ndhG by Binding Their 5' UTRs

Abstract

PGR3 is a P-class pentatricopeptide repeat (PPR) protein required for the stabilization of petL operon RNA and the translation of the petL gene in plastids. Irrespective of its important roles in plastids, key questions have remained unanswered, including how PGR3 protein promotes translation and which plastid mRNA PGR3 activates the translation. Here, we show that PGR3 facilitates the translation from ndhG, in addition to petL, through binding to their 5' untranslated regions (UTRs). Ribosome profiling and RNA sequencing in pgr3 mutants revealed that translation from petL and ndhG was specifically suppressed. Harnessing small RNA fragments protected by PPR proteins in vivo, we probed the PGR3 recruitment to the 5' UTRs of petL and ndhG. The putative PGR3-bound RNA segments per se repress the translation possibly with a strong secondary structure and thereby block ribosomes' access. However, the PGR3 binding antagonizes the effects and facilitates the protein synthesis from petL and ndhG in vitro. The prediction of the 3-dimensional structure of PGR3 suggests that the 26th PPR motif plays important roles in target RNA binding. Our data show the specificity of a plastidic RNA-binding protein and provide a mechanistic insight into translational control.

Keywords: Arabidopsis; Chloroplast; Gene expression; In vitro translation; P-type PPR protein; Translational regulation.