The MRS2/MGT gene family in Arabidopsis thaliana belongs to the superfamily of CorA-MRS2-ALR-type membrane proteins. Proteins of this type are characterized by a GMN tripeptide motif (Gly-Met-Asn) at the end of the first of two C-terminal transmembrane domains and have been characterized as magnesium transporters. Using the recently established mag-fura-2 system allowing direct measurement of Mg 2+ uptake into mitochondria of Saccharomyces cerevisiae, we find that all members of the Arabidopsis family complement the corresponding yeast mrs2 mutant. Highly different patterns of tissue-specific expression were observed for the MRS2/MGT family members in planta. Six of them are expressed in root tissues, indicating a possible involvement in plant magnesium supply and distribution after uptake from the soil substrate. Homozygous T-DNA insertion knockout lines were obtained for four members of the MRS2/MGT gene family. A strong, magnesium-dependent phenotype of growth retardation was found for mrs2-7 when Mg 2+ concentrations were lowered to 50 mM in hydroponic cultures. Ectopic overexpression of MRS2-7 from the cauliflower mosaic virus 35S promoter results in complementation and increased biomass accumulation. Green fluorescent protein reporter gene fusions indicate a location of MRS2-7 in the endomembrane system. Hence, contrary to what is frequently found in analyses of plant gene families, a single gene family member knockout results in a strong, environmentally dependent phenotype.

The RNA-binding pentatricopeptide repeat (PPR) family comprises hundreds to thousands of genes in most plants, but only a few dozen in algae, indicating massive gene expansions during land plant evolution. The nature and timing of these expansions has not been well defined due to the sparse sequence data available from early-diverging land plant lineages. In this study, we exploit the comprehensive OneKP datasets of over 1000 transcriptomes from diverse plants and algae toward establishing a clear picture of the evolution of this massive gene family, focusing on the proteins typically associated with RNA editing, which show the most spectacular variation in numbers and domain composition across the plant kingdom. We characterize over 2 250 000 PPR motifs in over 400 000 proteins. In lycophytes, polypod ferns, and hornworts, nearly 10% of expressed protein-coding genes encode putative PPR editing factors, whereas they are absent from algae and complex-thalloid liverworts. We show that rather than a single expansion, most land plant lineages with high numbers of editing factors have continued to generate novel sequence diversity. We identify sequence variations that imply functional differences between PPR proteins in seed plants versus non-seed plants and variations we propose to be linked to seed-plant-specific editing co-factors. Finally, using the sequence variations across the datasets, we develop a structural model of the catalytic DYW domain associated with C-to-U editing and identify a clade of unique DYW variants that are strong candidates as U-to-C RNA-editing factors, given their phylogenetic distribution and sequence characteristics.

BackgroundGene expression in plant chloroplasts and mitochondria is affected by RNA editing. Numerous C-to-U conversions, accompanied by reverse U-to-C exchanges in some plant clades, alter the genetic information encoded in the organelle genomes. Predicting and analyzing RNA editing, which ranges from only few sites in some species to thousands in other taxa, is bioinformatically demanding.ResultsHere, we present major enhancements and extensions of PREPACT, a WWW-based service for analysing, predicting and cataloguing plant-type RNA editing. New features in PREPACT’s core include direct GenBank accession query input and options to restrict searches to candidate U-to-C editing or to sites where editing has been documented previously in the references. The reference database has been extended by 20 new organelle editomes. PREPACT 3.0 features new modules “EdiFacts” and “TargetScan”. EdiFacts integrates information on pentatricopeptide repeat (PPR) proteins characterized as site-specific RNA editing factors. PREPACT’s editome references connect into EdiFacts, linking editing events to specific co-factors where known. TargetScan allows position-weighted querying for sequence motifs in the organelle references, optionally restricted to coding regions or sequences around editing sites, or in queries uploaded by the user. TargetScan is mainly intended to evaluate and further refine the proposed PPR-RNA recognition code but may be handy for other tasks as well. We present an analysis for the immediate sequence environment of more than 15,000 documented editing sites finding strong and different bias in the editome data sets.ConclusionsWe exemplarily present the novel features of PREPACT 3.0 aimed to enhance the analyses of plant-type RNA editing, including its new modules EdiFacts integrating information on characterized editing factors and TargetScan aimed to analyse RNA editing site recognition specificities.Electronic supplementary materialThe online version of this article (10.1186/s12859-018-2244-9) contains supplementary material, which is available to authorized users.

Summary Hornworts are crucial to understand the phylogeny of early land plants. The emergence of ‘reverse’ U‐to‐C RNA editing accompanying the widespread C‐to‐U RNA editing in plant chloroplasts and mitochondria may be a molecular synapomorphy of a hornwort–tracheophyte clade. C‐to‐U RNA editing is well understood after identification of many editing factors in models like Arabidopsis thaliana and Physcomitrella patens, but there is no plant model yet to investigate U‐to‐C RNA editing. The hornwort Anthoceros agrestis is now emerging as such a model system. We report on the assembly and analyses of the A. agrestis chloroplast and mitochondrial genomes, their transcriptomes and editomes, and a large nuclear gene family encoding pentatricopeptide repeat (PPR) proteins likely acting as RNA editing factors. Both organelles in A. agrestis feature high amounts of RNA editing, with altogether > 1100 sites of C‐to‐U and 1300 sites of U‐to‐C editing. The nuclear genome reveals > 1400 genes for PPR proteins with variable carboxyterminal DYW domains. We observe significant variants of the ‘classic’ DYW domain, in the meantime confirmed as the cytidine deaminase for C‐to‐U editing, and discuss the first attractive candidates for reverse editing factors given their excellent matches to U‐to‐C editing targets according to the PPR‐RNA binding code.