An Improved Assembly Strategy Helps Parsing the Cryptic Mitochondrial Genome Evolution in Plants

Although plant mitochondrial genomes (mitogenomes) are small, they exhibit considerable complexity not seen in other eukaryotic mitogenomes. Assembly and analysis of plant mitogenomes is hampered by their large variations in structure and size, and the mitogenome remains the last genome to be deciphered in many plant species. As a result, very few plant mitogenomes have been assembled and little is known regarding their evolution. In this study, a strategy was devised for assembly of mitogenomes from existing short reads from whole-genome sequencing projects. The strategy combined current tools and manual steps to resolve the two main challenges to mitogenome assembly: repeat and plastid insertion sequences. High-quality complete mitogenomes were assembled for 23 species from five families of the Fagales. Mitogenomes varied 2.4 times in size. The largest, Carpinus cordata, did not contain large amounts of unique sequences, but instead contained a high proportion of sequences homologous to other Fagales. Further analysis of the Fagales mitogenomes revealed highly mosaic characteristics, with horizonal transfer (HGT)-like sequences identified from almost all seed plant taxa. Independent and unequal transfers of third-party DNA may partially account for the HGT-like fragments and unbalanced size expansions observed in Fagales mitogenomes. Supporting this, a mitochondrial plasmid of nuclear origin was found in Carpinus, and this may represent an intermediate stage prior to incorporation into the mitogenome. The approaches used in this study are widely applicable and provide new insights into the mechanisms of mitogenome evolution in plants.

Complete vertebrate mitogenomes reveal widespread repeats and gene duplications

Background Modern sequencing technologies should make the assembly of the relatively small mitochondrial genomes an easy undertaking. However, few tools exist that address mitochondrial assembly directly. Results As part of the Vertebrate Genomes Project (VGP) we develop mitoVGP, a fully automated pipeline for similarity-based identification of mitochondrial reads and de novo assembly of mitochondrial genomes that incorporates both long (> 10 kbp, PacBio or Nanopore) and short (100–300 bp, Illumina) reads. Our pipeline leads to successful complete mitogenome assemblies of 100 vertebrate species of the VGP. We observe that tissue type and library size selection have considerable impact on mitogenome sequencing and assembly. Comparing our assemblies to purportedly complete reference mitogenomes based on short-read sequencing, we identify errors, missing sequences, and incomplete genes in those references, particularly in repetitive regions. Our assemblies also identify novel gene region duplications. The presence of repeats and duplications in over half of the species herein assembled indicates that their occurrence is a principle of mitochondrial structure rather than an exception, shedding new light on mitochondrial genome evolution and organization. Conclusions Our results indicate that even in the “simple” case of vertebrate mitogenomes the completeness of many currently available reference sequences can be further improved, and caution should be exercised before claiming the complete assembly of a mitogenome, particularly from short reads alone.

A functional bacteria-derived restriction modification system in the mitochondrion of a heterotrophic protist

The overarching trend in mitochondrial genome evolution is functional streamlining coupled with gene loss; therefore, gene acquisition by mitochondria is considered to be exceedingly rare. Selfish elements in the form of self-splicing introns occur in many organellar genomes, but the wider diversity of selfish elements, and how they persist in the DNA of organelles, has not been explored. In the mitochondrial genome of a marine heterotrophic katablepharid protist, we identify a functional type II restriction modification (RM) system originating from a horizontal gene transfer (HGT) event involving bacteria related to flavobacteria. This RM system consists of an HpaII-like endonuclease and a cognate cytosine methyltransferase (CM). We demonstrate that these proteins are functional by heterologous expression in both bacterial and eukaryotic cells. These results suggest that a mitochondrial-encoded RM system can function as a toxin–antitoxin selfish element and that such elements could be co-opted by eukaryotic genomes to drive biased organellar inheritance.

An updated assembly strategy helps parsing the cryptic mitochondrial genome evolution in plants

Although plant mitogenomes are small in size, their variations are no less than any other complex genomes. They are under rapid structure and size changes. These characters make the assembly a great challenge. This caused two intertwined problems, a slow growth of known mitogenomes and a poor knowledge of their evolution. In many species, mitogenome becomes the last genome that undeciphered. To have a better understanding of these two questions, we developed a strategy using short sequencing reads and combining current tools and manual steps to get high quality mitogenomes. This strategy allowed us to assembled 23 complete mitogenomes from 5 families in Fagales. Our large-scale comparative genomic analyses indicated the composition of mitogenomes is very mosaic that “horizontal transfers” can be from almost all taxa in seed plants. The largest mitogenome contains more homologous DNA with other Fagales, rather than unique sequences. Besides of real HGTs, sometimes mitovirus, nuclear insertions and other third-part DNA could also produce HGT-like sequences, accounting partially for the unusual evolutionary trajectories, including the cryptic size expansion in Carpinus. Mitochondrial plasmid was also found. Its lower GC content indicates that it may be only an interphase of a foreign DNA before accepting by the main chromosome. Our methods and results provide new insights into the assembly and mechanisms of mitogenome evolution.

Correction to: Potential causes and consequences of rapid mitochondrial genome evolution in thermoacidophilic Galdieria (Rhodophyta)

An amendment to this paper has been published and can be accessed via the original article.

Potential causes and consequences of rapid mitochondrial genome evolution in thermoacidophilic Galdieria (Rhodophyta)

Background The Cyanidiophyceae is an early-diverged red algal class that thrives in extreme conditions around acidic hot springs. Although this lineage has been highlighted as a model for understanding the biology of extremophilic eukaryotes, little is known about the molecular evolution of their mitochondrial genomes (mitogenomes). Results To fill this knowledge gap, we sequenced five mitogenomes from representative clades of Cyanidiophyceae and identified two major groups, here referred to as Galdieria-type (G-type) and Cyanidium-type (C-type). G-type mitogenomes exhibit the following three features: (i) reduction in genome size and gene inventory, (ii) evolution of unique protein properties including charge, hydropathy, stability, amino acid composition, and protein size, and (iii) distinctive GC-content and skewness of nucleotides. Based on GC-skew-associated characteristics, we postulate that unidirectional DNA replication may have resulted in the rapid evolution of G-type mitogenomes. Conclusions The high divergence of G-type mitogenomes was likely driven by natural selection in the multiple extreme environments that Galdieria species inhabit combined with their highly flexible heterotrophic metabolism. We speculate that the interplay between mitogenome divergence and adaptation may help explain the dominance of Galdieria species in diverse extreme habitats.

Potential causes and consequences of rapid mitochondrial genome evolution in thermoacidophilic Galdieria (Rhodophyta)

BackgroundThe Cyanidiophyceae is an early-diverged red algal class that thrives in extreme conditions around acidic hot springs. Although this lineage has been highlighted as a model for understanding the biology of extremophilic eukaryotes, little is known about the molecular evolution of their mitochondrial genomes (mitogenomes). ResultsTo fill this knowledge gap, we sequenced five mitogenomes from representative clades of Cyanidiophyceae and identified two major groups, here referred to as Galdieria-type (G-type) and Cyanidium-type (C-type). G-type mitogenomes exhibit the following three features: (i) reduction in genome size and gene inventory, (ii) evolution of unique protein properties including charge, hydropathy, stability, amino acid composition, and protein size, and (iii) distinctive GC-content and skewness of nucleotides. Based on GC-skew-associated characteristics, we postulate that unidirectional DNA replication may have resulted in the rapid evolution of G-type mitogenomes. ConclusionsThis high divergence was likely driven by natural selection in the multiple extreme environments that Galdieria species inhabit combined with their highly flexible heterotrophic metabolism. We speculate that the interplay between mitogenome divergence and adaptation may help explain the dominance of Galdieria species in diverse extreme habitats.

Microhomologies Are Associated with Tandem Duplications and Structural Variation in Plant Mitochondrial Genomes

Short tandem repeats (STRs) contribute to structural variation in plant mitochondrial genomes, but the mechanisms underlying their formation and expansion are unclear. In this study, we detected high polymorphism in the nad7-1 region of the Pinus tabuliformis mitogenome caused by the rapid accumulation of STRs and rearrangements over a few million years ago. The STRs in nad7-1 have a 7-bp microhomology (TAG7) flanking the repeat array. We then scanned the mitogenomes of 136 seed plants to understand the role of microhomology in the formation of STR and mitogenome evolution. A total of 13,170 STRs were identified, and almost half of them were associated with microhomologies. A substantial amount (1,197) of microhomologies was long enough to mediate structural variation, and the length of microhomology is positively correlated with the length of tandem repeat unit. These results suggest that microhomology may be involved in the formation of tandem repeat via microhomology-mediated pathway, and the formation of longer duplicates required greater length of microhomology. We examined the abundance of these 1,197 microhomologies, and found 75% of them were enriched in the plant mitogenomes. Further analyses of the 400 prevalent microhomologies revealed that 175 of them showed differential enrichment between angiosperms and gymnosperms and 186 differed between angiosperms and conifers, indicating lineage-specific usage and expansion of microhomologies. Our study sheds light on the sources of structural variation in plant mitochondrial genomes and highlights the importance of microhomology in mitochondrial genome evolution.