Genomic and transcriptomic analysis of plant PPR genes and organellar RNA editing

Abstract

The precise and perplexing coordination of plant organellar gene expression through nucleus-encoded trans-acting factors is puzzling from the genomic and evolutionary perspectives. Pentatricopeptide repeat (PPR) proteins are such players that are targeted exclusively to plant chloroplasts and mitochondria to participate nearly all of the post-transcriptional processes during RNA maturation. As a typical helix repeat modular protein, PPR constitutes one of the largest gene family in land plants, with more than 400 members estimated in most seed plants or larger than 2,000 copies in some genomes in bryophytes and ferns. The accurate recognition and regulation of target organellar RNA molecules by PPR proteins follow a modular manner with high specificity (one-motif-one-base), which has been described as a ―PPR code‖ to predict PPR-RNA interactions. However, this code is still poor in prediction capability due to the low-quality of the architectural annotation of the RNA-binding motifs and structural array arrangement. Thus, the accurate definition of PPR motif and the consistent annotation of PPR protein structure across species is a prerequisite for many downstream analyses, like the illumination of the molecular functions for diverse PPRs and the comparative analysis of PPR gene family evolution. On the other hand, the PLS-type PPRs are particularly interesting as key players in RNA editing activity in plant chloroplast and mitochondria, though their co-evolutionary patterns are still elusive. In this study, I performed a systematic study on PPR gene family in a wide range of plant genomes and transcriptomes, and further explored the co-evolution between PPR and RNA editing events in several model species. Firstly, structural modeling and contact prediction approach was used to redefine and confirm ten different variants of PPR motifs in plant proteins. Based on this, a robust computational pipeline for PPR gene annotation was developed through the combination of protein-based annotation results and genome-based predictions working on directly from the six-frame translation of genome assembly. I further applied my computational pipeline to analyze 109 published genomes, resulting in an unprecedented large PPR datasets, with high accuracy and consistency. A user-friendly PPR database searching and on-line annotation portal was built in this study: http://www.plantppr.com. This consistently annotated dataset is a valuable resource for comparative genomics studies and for large-scale prediction of PPR function. Secondly, I expanded the analysis by investigation of the 1KP data (1000 plant transcriptomes) that covers a broader species across evolution, revealing tens of thousands of new PPR proteins that were never identified before. Interestingly, a spectacular expansion of PPR gene family in land plants was con-firmed, especially during the colonization of land and evolutionary transition from a gametophyte- dominant to a sporophyte-dominant lifestyle. Thirdly, I explored the evolutionary characteristics of PPR gene family and the correlated RNA editing events in the newly sequenced ferns (Azolla filiculoides and Salvinia cucullata), by whole genome and transcriptome sequencing, RNA-seq alignment and variation calling, as well as comparative analysis of these two ferns with other closely-related plants. In summary, by comparative and computational genomic and transcriptomic studies on a broad wide of species including the newly sequenced model ferns, I redefined and reannotated PPR motifs and proteins followed by an extensive comparative analysis for PPR gene family and RNA editing events in plant organellar transcripts. These unprecedented studies revealed a remarkable journey of the evolution of PPR gene family and its co-evolution with the entire RNA editing machinery.