OrganellarGenomeDRAW (OGDRAW) version 1.3.1: expanded toolkit for the graphical visualization of organellar genomes.

Organellar (plastid and mitochondrial) genomes play an important role in resolving phylogenetic relationships, and next-generation sequencing technologies have led to a burst in their availability. The ongoing massive sequencing efforts require software tools for routine assembly and annotation of organellar genomes as well as their display as physical maps. OrganellarGenomeDRAW (OGDRAW) has become the standard tool to draw graphical maps of plastid and mitochondrial genomes. Here, we present a new version of OGDRAW equipped with a new front end. Besides several new features, OGDRAW now has access to a local copy of the organelle genome database of the NCBI RefSeq project. Together with batch processing of (multi-)GenBank files, this enables the user to easily visualize large sets of organellar genomes spanning entire taxonomic clades. The new OGDRAW server can be accessed at https://chlorobox.mpimp-golm.mpg.de/OGDraw.html.

GB2sequin - A file converter preparing custom GenBank files for database submission.

The typical wet lab user often annotates smaller sequences in the GenBank format, but resulting files are not accepted for database submission by NCBI. This makes submission of such annotations a cumbersome task. Here we present "GB2sequin" an easy-to-use web application that converts custom annotations in the GenBank format into the NCBI direct submission format Sequin. Additionally, the program generates a "five-column, tab-delimited feature table" and a FASTA file. Those are required for submission through BankIt or the update of an existing GenBank entry. We specifically developed "GB2sequin" for the regular wet lab researcher with strong focus on user-friendliness and flexibility. The application is equipped with an intuitive graphical interface and a comprehensive documentation. It can be employed to prepare any GenBank file for database submission and is freely available online at https://chlorobox.mpimp-golm.mpg.de/GenBank2Sequin.html.

GeSeq - versatile and accurate annotation of organelle genomes.

We have developed the web application GeSeq (https://chlorobox.mpimp-golm.mpg.de/geseq.html) for the rapid and accurate annotation of organellar genome sequences, in particular chloroplast genomes. In contrast to existing tools, GeSeq combines batch processing with a fully customizable reference sequence selection of organellar genome records from NCBI and/or references uploaded by the user. For the annotation of chloroplast genomes, the application additionally provides an integrated database of manually curated reference sequences. GeSeq identifies genes or other feature-encoding regions by BLAT-based homology searches and additionally, by profile HMM searches for protein and rRNA coding genes and two de novo predictors for tRNA genes. These unique features enable the user to conveniently compare the annotations of different state-of-the-art methods, thus supporting high-quality annotations. The main output of GeSeq is a GenBank file that usually requires only little curation and is instantly visualized by OGDRAW. GeSeq also offers a variety of optional additional outputs that facilitate downstream analyzes, for example comparative genomic or phylogenetic studies.

Chloroplast ribonucleoprotein CP31A is required for editing and stability of specific chloroplast mRNAs.

Chloroplast ribonucleoproteins (cpRNPs) are nuclear-encoded, highly abundant, and light-regulated RNA binding proteins. They have been shown to be involved in chloroplast RNA processing and stabilization in vitro and are phylogenetically related to the well-described heterogeneous nuclear ribonucleoproteins (hnRNPs). cpRNPs have been found associated with mRNAs present in chloroplasts and have been regarded as nonspecific stabilizers of chloroplast transcripts. Here, we demonstrate that null mutants of the cpRNP family member CP31A exhibit highly specific and diverse defects in chloroplast RNA metabolism. First, analysis of cp31a and cp31a/cp31b double mutants uncovers that these 2 paralogous genes participate nonredundantly in a combinatorial fashion in processing a subset of chloroplast editing sites in vivo. Second, a genome-wide analysis of chloroplast transcript accumulation in cp31a mutants detected a virtually complete loss of the chloroplast ndhF mRNA and lesser reductions for specific other mRNAs. Fluorescence analyses show that the activity of the NADH dehydrogenase complex, which also includes the NdhF subunit, is defective in cp31a mutants. This indicates that cpRNPs are important in vivo for calibrating the expression levels of specific chloroplast mRNAs and impact chloroplast physiology. Taken together, the specificity and combinatorial aspects of cpRNP functions uncovered suggest that these chloroplast proteins are functional equivalents of nucleocytosolic hnRNPs.

The evolution of chloroplast RNA editing.

RNA editing alters the nucleotide sequence of an RNA molecule so that it deviates from the sequence of its DNA template. Different RNA-editing systems are found in the major eukaryotic lineages, and these systems are thought to have evolved independently. In this study, we provide a detailed analysis of data on C-to-U editing sites in land plant chloroplasts and propose a model for the evolution of RNA editing in land plants. First, our data suggest that the limited RNA-editing system of seed plants and the much more extensive systems found in hornworts and ferns are of monophyletic origin. Further, although some eukaryotic editing systems appear to have evolved to regulate gene expression, or at least are now involved in gene regulation, there is no evidence that RNA editing plays a role in gene regulation in land plant chloroplasts. Instead, our results suggest that land plant chloroplast C-to-U RNA editing originated as a mechanism to generate variation at the RNA level, which could complement variation at the DNA level. Under this model, many of the original sites, particularly in seed plants, have been subsequently lost due to mutation at the DNA level, and the function of extant sites is merely to conserve certain codons. This is the first comprehensive model for the evolution of the chloroplast RNA-editing system of land plants and may also be applicable to the evolution of RNA editing in plant mitochondria.