Comparison of codon usage and tRNAs in mitochondrial genomes of Candida species

To gain insight into the nature of the mitochondrial genomes (mtDNA) of different Candida species, the synonymous codon usage bias of mitochondrial protein coding genes and the tRNAs in C. albicans, C. parapsilosis, C. stellata, C. glabrata and the closely related yeast Saccharomyces cerevisiae were analyzed. Common features of the mtDNA in Candida species are a strong A+T pressure on protein coding genes, and insufficient mitochondrial tRNA species are encoded to perform protein synthesis. The wobble site of the anticodon is always U for the NNR (NNA and NNG) codon families, which are dominated by A-ending codons, and always G for the NNY (NNC and NNU) codon families, which is dominated by U-ending codons, and always U for the NNN (NNA, NNU, NNC and NNG) codon families, which are dominated by A-ending codons and U-ending codons. Patterns of synonymous codon usage of Candida species can be classified into three groups: (1) optimal codon-anticodon usage, Glu, Lys, Leu (translated by anti-codon UAA), Gln, Arg (translated by anti-codon UCU) and Trp are containing NNR codons. NNA, whose corresponding tRNA is encoded in the mtDNA, is used preferentially. (2) Non-optimal codon-anticodon usage, Cys, Asp, Phe, His, Asn, Ser (translated by anti-codon GCU) and Tyr are containing NNY codons. The NNU codon, whose corresponding tRNA is not encoded in the mtDNA, is used preferentially. (3) Combined codon-anticodon usage, Ala, Gly, Leu (translated by anti-codon UAG), Pro, Ser (translated by anti-codon UGA), Thr and Val are containing NNN codons. NNA (tRNA encoded in the mtDNA) and NNU (tRNA not encoded in the mtDNA) are used preferentially. In conclusion, we propose that in Candida species, codons containing A or U at third position are used preferentially, regardless of whether corresponding tRNAs are encoded in the mtDNA. These results might be useful in understanding the common features of the mtDNA in Candida species and patterns of synonymous codon usage.     

Molecular phylogeny of the subfamily Schizothoracinae (Teleostei: Cypriniformes: Cyprinidae) inferred from complete mitochondrial genomes

The schizothoracine fishes, members of the Teleost order Cypriniformes, are one of the most diverse group of cyprinids in the Qinghai–Tibetan Plateau and surrounding regions. However, taxonomy and phylogeny of these species remain unclear. In this study, we determined the complete mitochondrial genome of Schizopygopsis malacanthus. We also used the newly obtained sequence, together with 31 published schizothoracine mitochondrial genomes that represent eight schizothoracine genera and six outgroup taxa to reconstruct the phylogenetic relationships of the subfamily Schizothoracinae by different partitioned maximum likelihood and partitioned Bayesian inference at nucleotide and amino acid levels. The schizothoracine fishes sampled form a strongly supported monophyletic group that is the sister taxon to Barbus barbus. A sister group relationship between the primitive schizothoracine group and the specialized schizothoracine group + the highly specialized schizothoracine group was supported. Moreover, members of the specialized schizothoracine group and the genera Schizothorax, Schizopygopsis, and Gymnocypris were found to be paraphyletic.     

Phylogenetic information from three mitochondrial genomes of Terebelliformia (Annelida) worms and duplication of the methionine tRNA

Mitochondrial genomes have been useful for inferring animal phylogeny across a wide range of clades, however they are still poorly sampled in some animal taxa, limiting our knowledge of mtDNA evolution. For example, despite being one of the most diverse animal phyla, only 5 complete annelid mitochrondial genomes have been published. To address this paucity of information, we obtained complete mitochondrial genomic sequences from Pista cristata (Terebellidae) and Terebellides stroemi (Trichobranchidae) as well as one nearly complete mitochondrial genome from Eclysippe vanelli (Ampharetidae). These taxa are within Terebelliformia (Annelida), which include spaghetti worms, icecream cone worms and their relatives. In contrast to the 37 genes found in most bilaterian metazoans, we recover 38 genes in the mitochondrial genomes of T. stroemi and P. cristata due to the presence of a second methionine tRNA (trnM). Interestingly, the two trnMs are located next to each other and are possibly a synapomorphy of these two taxa. The E. vanelli partial mitochondrial genome lacks this additional trnM at the same position, but it may be present in the region not sampled. Compared to other annelids, gene orders of these three mitochondrial genomes are generally conserved except for the atp6-mSSU region. Phylogenetic analyses reveal that mtDNA data strongly supports a Trichobranchidae/Terebellidae clade.     

Comparison of base composition and codon usage in insect mitochondrial genomes

Insects, the most biodiverse taxonomic group, have high AT content in their mitochondrial genomes. Although codon usage tends to be AT-rich, base composition and codon usage of mitochondrial genomes may vary among taxa. Thus, we compare base composition and codon usage patterns of 49 insect mitochondrial genomes. For protein coding genes, AT content is as high as 80% in the Hymenoptera and Lepidoptera and as low as 72% in the Orthopotera. The AT content is high at positions 1 and 3, but A content is low at position 2. A close correlation occurs between codon usage and tRNA abundance in nuclear genomes. Optimal codons can pair well with the antr codons of the most abundant tRNAs. One tRNA gene translates a synonymous codon family in vertebrate mitochondrial genomes and these tRNA anticodons can pair with optimal codons. However, optimal codons cannot pair with anticodons in mtDNA ofCochiiomyia hominivorax (Dipteral: CaLliphoridae). Ten optimal codons cannot pair with tRNA anticodons in all 49 insect mitochondrial genomes; non-optimal codon-anticodon usage is common and codon usage is not influenced by tRNA abundance.     

Complete sequences of mitochondrial genomes from the Baltic mussel Mytilus trossulus

Marine mussels Mytilus possess two mitochondrial (mt) genomes, which undergo doubly uniparental inheritance (DUI). Female (F) and male (M) genomes are usually highly diverged at the sequence level. Both genomes contain the same set of metazoan genes (for 12 proteins, 2 rRNAs and 23 tRNAs), both lack the atp8 gene and have two tRNAs for methionine. However, recently recombination between those variants has been reported. Both original F and M mt genomes of M. trossulus were replaced by M. edulis mtDNA in the Baltic populations. Highly diverged M genome occurs rarely in the Baltic mussels. Full sequences of the M genome identified in males (sperm) and F genome in females (eggs) were obtained. Both genomes were diverged by 24% in nucleotide sequence, but had similar nucleotide composition and codon usage bias. Constant domain (CD) of the control region (CR), the tRNA and rRNA genes were the most conserved. The most diverged was the variable domain 1 (VD1) of the control region. The F genome was longer than M by 147 bp. and the main difference was localised in the VD1 region. No recombination was observed in whole mtDNA of both studied variants. Nuclear mitochondrial pseudogenes (numts) have not been found by hybridisation with probes complementary to several fragments of the Baltic M. trossulus mtDNA.     

Complete DNA sequence of the mitochondrial genome ofCepaea nemoralis (Gastropoda: Pulmonata)

The nucleotide sequence of a mitochondrial genome of the pulmonate gastropod molluscCepaea nemoralis has been determined. Contained within the 14,100 basepairs (bp) are the two ribosomal RNA genes and 13 protein coding genes typical of metazoan mitochondrial genomes. TheCepaea mtDNA does contain a gene for ATPase subunit 8, like the clausiliid pulmonate,Albinaria, and the chiton,Katharina, but unlike the bivalve mollusc,Mytilus. The mitochondrial genetic code ofCepaea is proposed to be the same as that ofMytilus, Katharina, andDrosophila. Only 14 putative tRNA genes are presented, although there is sufficient unassigned sequence to encode the remainder of the expected total of 22 tRNA genes. These 14 tRNA genes are a mixture of standard cloverleaf structures and nonstandard structures containing TV replacement loops as seen in nematode and mosquito mitochondrial genomes. If the eight unidentified tRNA genes are indeed present, very little unassigned sequence would remain to serve as a control region. Genes are transcribed from both strands of the molecule. Base composition is the least biased for any reported animal mitochondrial genome and is also very little skewed between strands using measures independent of base composition. TheCepaea mitochondrial gene order is quite unlike that of any other reported metazoan mtDNA, with the exception of the recently reported partial sequences ofAlbinaria. No gene bound-aries are shared among all the reported molluscan taxa, demonstrating a complete lack of conservation of mitochondrial gene order across the phylum Mollusca.     

Three divergent mitochondrial genomes from California populations of the copepod Tigriopus californicus

Previous work on the harpacticoid copepod Tigriopus californicus has focused on the extensive population differentiation in three mtDNA protein coding genes (COXI, COXII, Cytb). In order to get a more complete understanding of mtDNA evolution in this species, we sequenced three complete mitochondrial genomes (one from each of three California populations) and compared them to two published mtDNA genomes from an Asian congener, Tigriopus japonicus. Several features of the mtDNA genome appear to be conserved within the genus: 1) the unique order of the protein coding genes, rRNA genes and most of the tRNA genes, 2) the genome is compact, varying between 14.3 and 14.6 kb, and 3) all genes are encoded on the same strand of the mtDNA. Within T. californicus, extremely high levels of nucleotide divergence (>20%) are observed across much of the mitochondrial genome. Inferred amino acid sequences of the proteins encoded in the mtDNAs also show high levels of divergence; at the extreme, the three ND3 variants in T. californicus showed >25% amino acid substitutions, compared with <3% amino acid divergence at the previously studied COXI locus. Unusual secondary structures make functional assignments of some tRNAs difficult. The only apparent tRNA(trp) in these genomes completely overlaps the 5' end of the 16S rRNA in all three T. californicus mtDNAs. Although not previously noted, this feature is also conserved in T. japonicus mtDNAs; whether this sequence is processed into a functional tRNA has not been determined. The putative control region contains a duplicated segment of different length (from 88 to 155 bp) in each of the T. californicus sequences. In each case, the duplicated segments are not tandem repeats; despite their different lengths, the distance between the start of the first and the start of the second repeat is conserved (520 bp). The functional significance, if any, of this repeat structure remains unknown.     

Discovery of a large number of previously unrecognized mitochondrial pseudogenes in fish genomes

Nuclear inserted copies of mitochondrial origin (numts) vary widely among eukaryotes, with human and plant genomes harboring the largest repertoires. Numts were previously thought to be absent from fish species, but the recent release of three fish nuclear genome sequences provides the resource to obtain a more comprehensive insight into the extent of mtDNA transfer in fishes. From the sequence analyses of the genomes of Fugu rubripes, Tetraodon nigroviridis, and Danio rerio, we have identified 2, 5, and 10 recent numt integrations, respectively, which integrated into those genomes less than 0.6 million years (Myr) ago. Such results contradict the hypothesis of absence or rarity of numts in fishes, as (i) the ratio of numts to the total size of the nuclear genome in T. nigroviridis was superior to the ratio observed in several higher vertebrate species (e.g., chicken, mouse, and rat), and only surpassed by humans, and (ii) the mtDNA coverage transferred to the nuclear genome of D. rerio is exceeded only by human and mouse, within the whole range of eukaryotic genomes surveyed for numts. Additionally, 335, 336, and 471 old numts (>12.5 Myr) were detected in F. rubripes, T. nigroviridis, and D. rerio, respectively. Surprisingly, old numts are inserted preferentially into known or predicted genes, as inferred for recent numts in human. However, because in fish genomes such integrations are old, they are likely to represent evolutionary successes and they may be considered a potential important evolutionary mechanism for the enhancement of genomic coding regions.     

High altitude adaptation of the schizothoracine fishes (Cyprinidae) revealed by the mitochondrial genome analyses

The schizothoracine fishes, also known as “mountain carps” are widely distributed in the Qinghai-Tibetan Plateau and its peripheral regions. Although they provide a prime example of high altitude adaptation, the phylogenetic relationships and the divergence times among these carp lineages are still controversial. Moreover, the genetic basis for high altitude adaptation is also poorly understood. In this study, we determined the mitochondrial genomes from two species of the schizothoracine fishes, representing a “morphologically primitive” clade and “morphologically specialized” clade, respectively. The phylogenetic tree and the divergence times were estimated within the evolutionary framework of the entire order Cypriniformes. Our results indicate a polyphylyetic relationship of the schizothoracine fishes and suggest two independent migration events into the Qinghai-Tibetan Plateau: one by the “morphologically primitive” clade in the Late Miocene and another by the “morphologically specialized” clade in the Eocene. Rapid speciation events of each clade from the Late Miocene to the Pliocene correspond to the timing of the geologic acceleration of the Qinghai-Tibetan Plateau. Interestingly, we found evidence for positive selection acting on the protein coding genes in the mitochondrial genomes of the “morphologically specialized” clade, implying a possible genetic basis for high altitude adaptation in this derived lineage of cypriniform fishes.     

Gene rearrangements in snake mitochondrial genomes: highly concerted evolution of control-region-like sequences duplicated and inserted into a tRNA gene cluster

Mitochondrial DNA (mtDNA) regions corresponding to two major tRNA gene clusters were amplified and sequenced for the Japanese pit viper, himehabu. In one of these clusters, which in most vertebrates characterized to date contains three tightly connected genes for tRNA(Ile), and tRNA(Gln), and tRNA(Met), a sequence of approximately 1.3 kb was found to be inserted between the genes for tRNA(Ile) and tRNA(Gln). The insert consists of a control-region-like sequence possessing some conserved sequence blocks, and short flanking sequences which may be folded into tRNA(Pro), tRNA(Phe), and tRNA(Leu) genes. Several other snakes belonging to different families were also found to possess a control-region-like sequence and tRNA(Leu) gene between the tRNA(Ile)and tRNA(Gln) genes. We also sequenced a region surrounded by genes for cytochrome b and 12S rRNA, where the control region and genes for tRNA(Pro) and tRNA(Phe) are normally located in the mtDNAs of most vertebrates. In this region of three examined snakes, a control-region- like sequence exists that is almost completely identical to the one found between the tRNA(Ile) and tRNA(Gln) genes. The mtDNAs of these snakes thus possess two nearly identical control-region-like sequences which are otherwise divergent to a large extent between the species. These results suggest that the duplicate state of the control-region- like sequences has long persisted in snake mtDNAs, possibly since the original insertion of the control-region-like sequence and tRNA(Leu) gene into the tRNA gene cluster, which occurred in the early stage of the divergence of snakes. It is also suggested that the duplicated control-region-like sequences at two distant locations of mtDNA have evolved concertedly by a mechanism such as frequent gene conversion. The secondary structures of the determined tRNA genes point to the operation of simplification pressure on the T psi C arm of snake mitochondrial tRNAs.      

Sequence and gene organization of the chicken mitochondrial genome. A novel gene order in higher vertebrates

The 16,775 base-pair mitochondrial genome of the white Leghorn chicken has been cloned and sequenced. The avian genome encodes the same set of genes (13 proteins, 2 rRNAs and 22 tRNAs) as do other vertebrate mitochondrial DNAs and is organized in a very similar economical fashion. There are very few intergenic nucleotides and several instances of overlaps between protein or tRNA genes. The protein genes are highly similar to their mammalian and amphibian counterparts and are translated according to the same variant genetic code. Despite these highly conserved features, the chicken mitochondrial genome displays two distinctive characteristics. First, it exhibits a novel gene order, the contiguous tRNA(Glu) and ND6 genes are located immediately adjacent to the displacement loop region of the molecule, just ahead of the contiguous tRNA(Pro), tRNA(Thr) and cytochrome b genes, which border the displacement loop region in other vertebrate mitochondrial genomes. This unusual gene order is conserved among the galliform birds. Second, a light-strand replication origin, equivalent to the conserved sequence found between the tRNA(Cys) and tRNA(Asn) genes in all vertebrate mitochondrial genomes sequenced thus far, is absent in the chicken genome. These observations indicate that galliform mitochondrial genomes departed from their mammalian and amphibian counterparts during the course of evolution of vertebrate species. These unexpected characteristics represent useful markers for investigating phylogenetic relationships at a higher taxonomic level.     

Complete mitochondrial DNA sequences of two haplotypes of the smallmouth bass, <Emphasis Type="Italic">Micropterus dolomieu</Emphasis>, collected from nonindigenous populations in Japan

We determined the complete mitochondrial genome sequences of two haplotypes of the smallmouth bass, Micropterus dolomieu; individuals used in the analysis were collected from nonindigenous populations in Japan. Both genomes comprised 16,488 bp, with genome contents and gene orders being identical to those of other teleost fishes. A previous study revealed that the Japanese smallmouth bass had only two haplotypes, and the present study revealed that the complete mitochondrial DNA (mtDNA) sequences of the haplotypes differed in only one nucleotide difference. The low genetic diversity in the mtDNA of the smallmouth bass individuals in our study and the results of the mtDNA sequence comparison between the Japanese and the North American individuals suggested that the fish had been transplanted from a fish farm with a low-diversity stock.     

Phylogenetic studies of three sinipercid fishes (Perciformes: Sinipercidae) based on complete mitochondrial DNA sequences

The sinipercids are a group of 12 species of freshwater percoid fish endemic to East Asia and their phylogenetic placements have perplexed generations of taxonomists. We cloned and sequenced the complete mitochondrial DNA (mtDNA) of three sinipercid fishes (Siniperca chuatsi, S. kneri, and S. scherzeri) to characterize and compare their mitochondrial genomes. The mitochondrial genomes of S. chuatsi, S. kneri, and S. scherzeri were 16,496, 17,002, and 16,585?bp in length, respectively. The organization of the three mitochondrial genomes is similar to those reported from other fish mitochondrial genomes, which contains 37 genes (13 protein-coding genes, 2 ribosomal RNAs, and 22 transfer RNAs) and a major non-coding control region. Among the 13 protein-coding genes of all the three sinipercid fishes, three reading-frame overlaps were found on the same strand. There is an 81-bp tandem repeat cluster at the end of CSB-3 in the S. scherzeri control region. The complete mitochondrial genomes of the three sinipercids should be useful for the evolutionary studies of sinipercids and other vertebrate species.     

A low rate of replacement substitutions in two major Ovis aries mitochondrial genomes

Two mitochondrial DNA molecules which represent major Ovis aries mtDNA haplogroups were cloned and comparatively sequenced to assess the degree of intraspecific variation. A total of 9623 bp that correspond to 58% of both mitochondrial genomes were determined. The control region, the Cyt b , ND2, ND3, ND4L, COIII and 12 tRNA genes, including the origin of L-strand replication, were completely characterized. Partial sequence information was obtained from the 12S and 16S rRNA and an additional six protein coding and six tRNA genes. The control regions of the two mtDNAs showed a nucleotide divergence of 4·34% while coding regions differed by 0·44%. The number of sheep coding region substitutions was similar to values observed in intraspecific comparisons of mitochondrial DNAs that represent remote points in genealogical trees of mice and humans. However, replacement substitutions were only observed at ∼30% of the rate in mice and ∼20% of the rate in humans. Nucleotide substitutions with a potential for phenotypic effects were found in the 12S and 16S rRNA and in the ND1 and COIII genes.     

Towards a comprehensive picture of alloacceptor tRNA remolding in metazoan mitochondrial genomes

Remolding of tRNAs is a well-documented process in mitochondrial genomes that changes the identity of a tRNA. It involves a duplication of a tRNA gene, a mutation that changes the anticodon and the loss of the ancestral tRNA gene. The net effect is a functional tRNA that is more closely related to tRNAs of a different alloacceptor family than to tRNAs with the same anticodon in related species. Beyond being of interest for understanding mitochondrial tRNA function and evolution, tRNA remolding events can lead to artifacts in the annotation of mitogenomes and thus in studies of mitogenomic evolution. Therefore, it is important to identify and catalog these events. Here we describe novel methods to detect tRNA remolding in large-scale data sets and apply them to survey tRNA remolding throughout animal evolution. We identify several novel remolding events in addition to the ones previously mentioned in the literature. A detailed analysis of these remoldings showed that many of them are derived from ancestral events.     

Seventeen new complete mtDNA sequences reveal extensive mitochondrial genome evolution within the Demospongiae

Two major transitions in animal evolution--the origins of multicellularity and bilaterality--correlate with major changes in mitochondrial DNA (mtDNA) organization. Demosponges, the largest class in the phylum Porifera, underwent only the first of these transitions and their mitochondrial genomes display a peculiar combination of ancestral and animal-specific features. To get an insight into the evolution of mitochondrial genomes within the Demospongiae, we determined 17 new mtDNA sequences from this group and analyzing them with five previously published sequences. Our analysis revealed that all demosponge mtDNAs are 16- to 25-kbp circular molecules, containing 13-15 protein genes, 2 rRNA genes, and 2-27 tRNA genes. All but four pairs of sampled genomes had unique gene orders, with the number of shared gene boundaries ranging from 1 to 41. Although most demosponge species displayed low rates of mitochondrial sequence evolution, a significant acceleration in evolutionary rates occurred in the G1 group (orders Dendroceratida, Dictyoceratida, and Verticillitida). Large variation in mtDNA organization was also observed within the G0 group (order Homosclerophorida) including gene rearrangements, loss of tRNA genes, and the presence of two introns in Plakortis angulospiculatus. While introns are rare in modern-day demosponge mtDNA, we inferred that at least one intron was present in cox1 of the common ancestor of all demosponges. Our study uncovered an extensive mitochondrial genomic diversity within the Demospongiae. Although all sampled mitochondrial genomes retained some ancestral features, including a minimally modified genetic code, conserved structures of tRNA genes, and presence of multiple non-coding regions, they vary considerably in their size, gene content, gene order, and the rates of sequence evolution. Some of the changes in demosponge mtDNA, such as the loss of tRNA genes and the appearance of hairpin-containing repetitive elements, occurred in parallel in several lineages and suggest general trends in demosponge mtDNA evolution.     

Recombination sequences in plant mitochondrial genomes: diversity and homologies to known mitochondrial genes.

Several plant mitochondrial genomes contain repeated sequences that are postulated to be sites of homologous intragenomic recombination (1-3). In this report, we have used filter hybridizations to investigate sequence relationships between the cloned mitochondrial DNA (mtDNA) recombination repeats from turnip, spinach and maize and total mtDNA isolated from thirteen species of angiosperms. We find that strong sequence homologies exist between the spinach and turnip recombination repeats and essentially all other mitochondrial genomes tested, whereas a major maize recombination repeat does not hybridize to any other mtDNA. The sequences homologous to the turnip repeat do not appear to function in recombination in any other genome, whereas the spinach repeat hybridizes to reiterated sequences within the mitochondrial genomes of wheat and two species of pokeweed that do appear to be sites of recombination. Thus, although intragenomic recombination is a widespread phenomenon in plant mitochondria, it appears that different sequences either serve as substrates for this function in different species, or else surround a relatively short common recombination site which does not cross-hybridize under our experimental conditions. Identified gene sequences from maize mtDNA were used in heterologous hybridizations to show that the repeated sequences implicated in recombination in turnip and spinach/pokeweed/wheat mitochondria include, or are closely linked to genes for subunit II of cytochrome c oxidase and 26S rRNA, respectively. Together with previous studies indicating that the 18S rRNA gene in wheat mtDNA is contained within a recombination repeat (3), these results imply an unexpectedly frequent association between recombination repeats and plant mitochondrial genes.