Platyhelminth mitochondrial DNA: Evidence for early evolutionary origin of a tRNAserAGN that contains a dihydrouridine arm replacement loop,and of serine-specifying AGA and AGG codons

Summary The nucleotide sequence of a segment of the mitochondrial DNA (mtDNA) molecule of the liver flukeFasciola hepatica (phylum Platyhelminthes, class Trematoda) has been determined, within which have been identified the genes for tRNA^ala, tRNA^asp, respiratory chain NADH dehydrogenase subunit I (ND1), tRNA^asn, tRNA^pro, tRNA^ile, tRNA^lys, ND3, tRNA^serAGN, tRNA^trp, and cytochromec oxidase subunit I (COI). The 11 genes are arranged in the order given and are all transcribed from the same strand of the molecule. The overall order of theF. hepatica mitochondrial genes differs from what is found in other metazoan mtDNAs. All of the sequenced tRNA genes except the one for tRNA^serAGN can be folded into a secondary structure with four arms resembling most other metazoan mitochondrial tRNAs, rather than the tRNAs that contain a TψC arm replacement loop, found in nematode mtDNAs. TheF. hepatica mitochondrial tRNA^serAGN gene contains a dihydrouridine arm replacement loop, as is the case in all other metazoan mtDNAs examined to date. AGA and AGG are found in theF. hepatica mitochondrial protein genes and both codons appear to specify serine. These findings concerningF. hepatica mtDNA indicate that both a dihydrouridine arm replacement loop-containing tRNA^serAGN gene and the use of AGA and AGG codons to specify serine must first have occurred very early in, or before, the evolution of metazoa.     

The Mitochondrial Genomes of Two Nematodes, Caenorhabditis Elegans and Ascaris Suum

The nucleotide sequences of the mitochondrial DNA (mtDNA) molecules of two nematodes, Caenorhabditis elegans [13,794 nucleotide pairs (ntp)], and Ascaris suum (14,284 ntp) are presented and compared. Each molecule contains the genes for two ribosomal RNAs (s-rRNA and l-rRNA), 22 transfer RNAs (tRNAs) and 12 proteins, all of which are transcribed in the same direction. The protein genes are the same as 12 of the 13 protein genes found in other metazoan mtDNAs: Cyt b, cytochrome b; COI-III, cytochrome c oxidase subunits I-III; ATPase6, Fo ATPase subunit 6; ND1-6 and 4L, NADH dehydrogenase subunits 1-6 and 4L: a gene for ATPase subunit 8, common to other metazoan mtDNAs, has not been identified in nematode mtDNAs. The C. elegans and A. suum mtDNA molecules both include an apparently noncoding sequence that contains runs of AT dinucleotides, and direct and inverted repeats (the AT region: 466 and 886 ntp, respectively). A second, apparently noncoding sequence in the C. elegans and A. suum mtDNA molecules (109 and 117 ntp, respectively) includes a single, hairpin-forming structure. There are only 38 and 89 other intergenic nucleotides in the C. elegans and A. suum mtDNAs, and no introns. Gene arrangements are identical in the C. elegans and A. suum mtDNA molecules except that the AT regions have different relative locations. However, the arrangement of genes in the two nematode mtDNAs differs extensively from gene arrangements in all other sequenced metazoan mtDNAs. Unusual features regarding nematode mitochondrial tRNA genes and mitochondrial protein gene initiation codons, previously described by us, are reviewed. In the C. elegans and A. suum mt-genetic codes, AGA and AGG specify serine, TGA specifies tryptophan and ATA specifies methionine. From considerations of amino acid and nucleotide sequence similarities it appears likely that the C. elegans and A. suum ancestral lines diverged close to the time of divergence of the cow and human ancestral lines, about 80 million years ago.     

A Novel Mitochondrial Genome Organization for the Blue Mussel, Mytilus Edulis

The sequence of 13.9 kilobases (kb) of the 17.1-kb mitochondrial genome of Mytilus edulis has been determined, and the arrangement of all genes has been deduced. Mytilus mitochondrial DNA (mtDNA) contains 37 genes, all of which are transcribed from the same DNA strand. The gene content of Mytilus is typically metazoan in that it includes genes for large and small ribosomal RNAs, for a complete set of transfer RNAs and for 12 proteins. The protein genes encode the cytochrome b apoenzyme, cytochrome c oxidase (CO) subunits I-III, NADH dehydrogenase (ND) subunits 1-6 and 4L, and ATP synthetase (ATPase) subunit 6. No gene for ATPase subunit 8 could be found. The reading frames for the ND1, COI, and COIII genes contain long extensions relative to those genes in other metazoan mtDNAs. There are 23 tRNA genes, one more than previously found in any metazoan mtDNA. The additional tRNA appears to specify methionine, making Mytilus mtDNA unique in having two tRNA(Met) genes. Five lengthy unassigned intergenic sequences are present, four of which vary in length from 79 to 119 nucleotides and the largest of which is 1.2 kb. The base compositions of these are unremarkable and do not differ significantly from that of the remainder of the mtDNA. The arrangement of genes in Mytilus mtDNA is remarkably unlike that found in any other known metazoan mtDNA.     

Complete DNA Sequence of the Mitochondrial Genome of the Black Chiton, Katharina Tunicata

The DNA sequence of the 15,532-base pair (bp) mitochondrial DNA (mtDNA) of the chiton Katharina tunicata has been determined. The 37 genes typical of metazoan mtDNA are present: 13 for protein subunits involved in oxidative phosphorylation, 2 for rRNAs and 22 for tRNAs. The gene arrangement resembles those of arthropods much more than that of another mollusc, the bivalve Mytilus edulis. Most genes abut directly or overlap, and abbreviated stop codons are inferred for four genes. Four junctions between adjacent pairs of protein genes lack intervening tRNA genes; however, at each of these junctions there is a sequence immediately adjacent to the start codon of the downstream gene that is capable of forming a stem-and-loop structure. Analysis of the tRNA gene sequences suggests that the D arm is unpaired in tRNA(ser(AGN)), which is typical of metazoan mtDNAs, and also in tRNA(ser(UCN)), a condition found previously only in nematode mtDNAs. There are two additional sequences in Katharina mtDNA that can be folded into structures resembling tRNAs; whether these are functional genes is unknown. All possible codons except the stop codons TAA and TAG are used in the protein-encoding genes, and Katharina mtDNA appears to use the same variation of the mitochondrial genetic code that is used in Drosophila and Mytilus. Translation initiates at the codons ATG, ATA and GTG. A + T richness appears to have affected codon usage patterns and, perhaps, the amino acid composition of the encoded proteins. A 142-bp non-coding region between tRNA(glu) and CO3 contains a 72-bp tract of alternating A and T.     

黄毛纺蚋线粒体基因组全序列测定与分析

利用PCR步移法对黄毛纺蚋的线粒体基因组全序列进行了测定和分析。黄毛纺蚋线粒体基因组全长15904 bp(Gen Bank序列号KP793690),包括13个蛋白编码基因、22个tRNA基因、2个rRNA基因以及长度为939 bp的非编码区。A、T、C、G碱基含量分别为39.1%、35.8%、10.4%、14.7%。9个蛋白编码基因和14个tRNA基因在J链编码,其余4个蛋白编码基因和8个tRNA基因在N链编码,基因排列顺序与其它已知双翅目昆虫相同。13个蛋白编码基因中除COI以TTG作为起始密码外,其余蛋白质基因均以ATN作为起始密码子,终止密码子多数为典型的TAA、TAG,只有COI和ND4L以单独的T作为终止密码子。在所测得的22个tRNA基因中,除tRNASer(AGN)缺少DHU臂外,其余tRNA均能形成典型的三叶草结构。     

Mytilus mitochondrial DNA contains a functional gene for a tRNASer(UCN) with a dihydrouridine arm-replacement loop and a pseudo-tRNASer(UCN) gene.

A 2500-nucleotide pair (ntp) sequence of F-type mitochondrial (mt) DNA of the Pacific Rim mussel Mytilus californianus (class Bivalvia, phylum Mollusca) that contains two complete (ND2 and ND3) and two partial (COI and COIII) protein genes and nine tRNA genes is presented. Seven of the encoded tRNAs (Ala, Arg, His, Met(AUA), Pro, Ser(UCN), and Trp) have the potential to fold into the orthodox four-armed tRNA secondary structure, while two [tRNASer(AGN) and a second tRNASer(UCN)] will fold only into tRNAs with a dihydrouridine (DHU) arm-replacement loop. Comparison of these mt-tRNA gene sequences with previously published, corresponding M. edulis F-type mtDNA indicates that similarity between the four-armed tRNASer(UCN) genes is only 63.8% compared with an average of 92.1% (range 86.2-98. 5%) for the remaining eight tRNA genes. Northern blot analysis indicated that mature tRNAs encoded by the DHU arm-replacement loop-containing tRNASer(UCN), tRNASer(AGN), tRNAMet(AUA), tRNATrp, and tRNAPro genes occur in M. californianus mitochondria, strengthening the view that all of these genes are functional. However, Northern blot and 5' RACE (rapid amplification of cDNA ends) analyses indicated that the four-armed tRNASer(UCN) gene is transcribed into a stable RNA that includes the downstream COI sequence and is not processed into a mature tRNA. On the basis of these observations the M. californianus and M. edulis four-armed tRNASer(UCN) sequences are interpreted as pseudo-tRNASer(UCN) genes.     

Mitochondrial genomes of two demosponges provide insights into an early stage of animal evolution

Mitochondrial DNA (mtDNA) of multicellular animals (Metazoa) is typically a small ( approximately 16 kbp), circular-mapping molecule that encodes 37 tightly packed genes. The structures of mtDNA-encoded transfer RNAs (tRNAs) and ribosomal RNAs (rRNAs) are usually highly unorthodox, and proteins are translated with multiple deviations from the standard genetic code. In contrast, mtDNA of the choanoflagellate Monosiga brevicollis, the closest unicellular relative of animals, is four times larger, contains 1.5 times as many genes, and lacks mentioned peculiarities of animal mtDNA. To investigate the evolutionary transition that led to the specific organization of metazoan mtDNA, we determined complete mitochondrial sequences from the demosponges Geodia neptuni and Tethya actinia, two representatives of the most basal animal phylum, the Porifera. We found that poriferan mtDNAs resemble those of other animals in their compact organization, lack of introns, and a well-conserved animal-like gene order. Yet, they contain several extra genes, encode bacterial-like rRNAs and tRNAs, and use a minimally derived genetic code. Our findings suggest that the evolution of the typical metazoan mtDNA has been a multistep process in which the compact genome organization and the reduced gene content were established prior to the reduction of tRNA and rRNA structures and the introduction of multiple changes of the translation code.     

Mitochondrial DNA of the sea anemone,Metridium senile (Cnidaria): Prokaryote-like genes for tRNAf-Met and small-subunit ribosomal RNA,and standard genetic code specificities for AGR and ATA codons

The nucleotide sequence of a segment of the mitochondrial DNA (mtDNA) molecule of the sea anemone Metridium senile (phylum Cnidaria, class Anthozoa, order Actiniaria) has been determined, within which have been identified the genes for respiratory chain NADH dehydrogenase subunit 2 (ND2), the small-subunit rRNA (s-rRNA), cytochrome c oxidase subunit II(COII), ND4, ND6, cytochrome b (Cyt b), tRNA^f-Met, and the large-subunit rRNA (1-rRNA). The eight genes are arranged in the order given and are all transcribed from the same strand of the molecule. The overall order of the M. senile mt-genes differs from that of other metazoan mtDNAs. In M. senile mt-protein genes, AGA and AGG codons appear to have the standard genetic code specification of arginine, rather than serine as found for other invertebrate mt-genetic codes. Also, ATA has the standard genetic code specification of isoleucine. TGA occurs in three M. senile mt-protein genes and may specify tryptophan as in other metazoan, protozoan, and some fungal mt-genetic codes. The M. senile mt-rRNA^f-Met gene has primary and secondary structure features closely resembling those of the Escherichia coli initiator tRNA, including standard dihydrouridine and TC loop sequences and a mismatch pair at the top of the aminoacyl stem. Determinations of the 5 and 3 end nucleotides of the M. senile mt-srRNAs indicated that these molecules have a homogenous size of 1,081 ntp, larger than any other known metazoan mt-s-rRNAs. Consistent with its larger size, the M. senile mt-s-rRNA can be folded into a secondary structure that more closely resembles that of the E. coli 16S rRNA than can any other metazoan mt-s-rRNA. These findings concerning M. senile mtDNA indicate that most of the unusual features regarding metazoan mt-genetic codes, rRNAs, and probably tRNAs developed after divergence of the Cnidarian line from the ancestral line common to other metazoa.Correspondence to: D.R. Wolstenholme     

Nucleotide sequence of nine protein-coding genes and 22 tRNAs in the mitochondrial DNA of the sea starPisaster ochraceus

Summary We have cloned and sequenced over 9 kb of the mitochondrial genome from the sea starPisaster ochraceus. Within a continuous 8.0-kb fragment are located the genes for NADH dehydrogenase subunits 1, 2, 3, and 4L (ND1, ND2, ND3, and ND4L), cytochrome oxidase subunits I, II, and III (COI, COII, and COIII), and adenosine triphosphatase subunits 6 and 8 (ATPase 6 and ATPase 8). This large fragment also contains a cluster of 13 tRNA genes between ND1 and COI as well as the genes for isoleucine tRNA between ND1 and ND2, arginine tRNA between COI and ND4L, lysine tRNA between COII and ATPase 8, and the serine (UCN) tRNA between COIII and ND3. The genes for the other five tRNAs lie outside this fragment. The gene for phenylalanine tRNA is located between cytochrome b and the 12S ribosomal genes. The genes for tRNA^glu and tRNA^thr are 3 to the 12S ribosomal gene. The tRNAs for histidine and serine (AGN) are adjacent to each other and lie between ND4 and ND5. These data confirm the novel gene order in mitochondrial DNA (mtDNA) of sea stars and delineate additional distinctions between the sea star and other mtDNA molecules.     

Mitochondrial sequence evolution in spiders: intraspecific variation in tRNAs lacking the TPsiC Arm

Analyses of mitochondrial DNA sequences from three species of Habronattus jumping spiders (Chelicerata: Arachnida: Araneae) reveal unusual inferred tRNA secondary structures and gene arrangements, providing new information on tRNA evolution within chelicerate arthropods. Sequences from the protein-coding genes NADH dehydrogenase subunit 1 (ND1), cytochrome oxidase subunit I (COI), and subunit II (COII) were obtained, along with tRNA, tRNA, and large-subunit ribosomal RNA (16S) sequences; these revealed several peculiar features. First, inferred secondary structures of tRNA and, likely, tRNA, lack the TPsiC arm and the variable arm and therefore do not form standard cloverleaf structures. In place of these arms is a 5-6-nt T arm-variable loop (TV) replacement loop such as that originally described from nematode mitochondrial tRNAs. Intraspecific variation occurs in the acceptor stem sequences in both tRNAs. Second, while the proposed secondary structure of the 3' end of 16S is similar to that reported for insects, the sequence at the 5' end is extremely divergent, and the entire gene is truncated about 300 nt with respect to Drosophila yakuba. Third, initiation codons appear to consist of ATY (ATT and ATC) and TTG for ND1 and COII, respectively. Finally, Habronattus shares the same ND1-tRNA-16S gene arrangement as insects and crustaceans, thus illustrating variation in a tRNA gene arrangement previously proposed as a character distinguishing chelicerates from insects and crustaceans.     

The mitochondrial genome of the stramenopile alga Chrysodidymus synuroideus. Complete sequence, gene content and genome organization

This is the first report of a complete mitochondrial genome sequence from a photosynthetic member of the stramenopiles, the chrysophyte alga Chrysodidymus synuroideus. The circular-mapping mitochondrial DNA (mtDNA) of 34 119 bp contains 58 densely packed genes (all without introns) and five unique open reading frames (ORFs). Protein genes code for components of respiratory chain complexes, ATP synthase and the mitoribosome, as well as one product of unknown function, encoded in many other protist mtDNAs (YMF16). In addition to small and large subunit ribosomal RNAs, 23 tRNAs are mtDNA-encoded, permitting translation of all codons present in protein-coding genes except ACN (Thr) and CGN (Arg). The missing tRNAs are assumed to be imported from the cytosol. Comparison of the C.synuroideus mtDNA with that of other stramenopiles allowed us to draw conclusions about mitochondrial genome organization, expression and evolution. First, we provide evidence that mitochondrial ORFs code for highly derived, unrecognizable versions of ribosomal or respiratory genes otherwise ‘missing’ in a particular mtDNA. Secondly, the observed constraints in mitochondrial genome rearrangements suggest operon-based, co-ordinated expression of genes functioning in common biological processes. Finally, stramenopile mtDNAs reveal an unexpectedly low variability in genome size and gene complement, testifying to substantial differences in the tempo of mtDNA evolution between major eukaryotic lineages.     

The complete mitochondrial genome of the damsel bug Alloeorhynchus bakeri (Hemiptera: Nabidae)

The complete sequence of the mitochondrial DNA (mtDNA) of the damsel bug, Alloeorhynchus bakeri, has been completed and annotated in this study. It represents the first sequenced mitochondrial genome of heteropteran family Nabidae. The circular genome is 15, 851 bp in length with an A+T content of 73.5%, contains the typical 37 genes that are arranged in the same order as that of the putative ancestor of hexapods. Nucleotide composition and codon usage are similar to other known heteropteran mitochondrial genomes. All protein-coding genes (PCGs) use standard initiation codons (methionine and isoleucine), except COI, which started with TTG. Canonical TAA and TAG termination codons are found in eight protein-coding genes, the remaining five (COI, COII, COIII, ND5, ND1) have incomplete termination codons (T or TA). PCGs of two strands present opposite CG skew which is also reflected by the nucleotide composition and codon usage. All tRNAs have the typical clover-leaf structure, except the dihydrouridine (DHU) arm of tRNA(Ser (AGN))which forms a simple loop as known in many other metazoa. Secondary structure models of the ribosomal RNA genes of A. bakeri are presented, similar to those proposed for other insect orders. There are six domains and 45 helices and three domains and 27 helices in the secondary structures of rrnL and rrnS, respectively. The major non-coding region (also called control region) between the small ribosomal subunit and the tRNA(Ile )gene includes two special regions. The first region includes four 133 bp tandem repeat units plus a partial copy of the repeat (28 bp of the beginning), and the second region at the end of control region contains 4 potential stem-loop structures. Finally, PCGs sequences were used to perform a phylogenetic study. Both maximum likelihood and Bayesian inference analyses highly support Nabidae as the sister group to Anthocoridae and Miridae.     

Avoidance of antisense, antiterminator tRNA anticodons in vertebrate mitochondria

Protein synthesis (translation) stops at stop codons, codons not complemented by tRNA anticodons. tRNAs matching stops, antitermination (Ter) tRNAs, prevent translational termination, producing dysfunctional proteins. Genomes avoid tRNAs with anticodons whose complement (the anticodon of the ‘antisense’ tRNA) matches stops. This suggests that antisense tRNAs, which also form cloverleaves, are occasionally expressed. Mitochondrial antisense tRNA expression is plausible, because both DNA strands are transcribed as single RNAs, and tRNA structures signal RNA maturation. Results describe potential antisense Ter tRNAs in mammalian mitochondrial genomes detected by tRNAscan-SE, and evidence for adaptations preventing translational antitermination: genomes possessing Ter tRNAs use less corresponding stop codons; antisense Ter tRNAs form weaker cloverleaves than homologuous non-Ter antisense tRNAs; and genomic stop codon usages decrease with stabilities of codon-anticodon interactions and of Ter tRNA cloverleaves. This suggests that antisense tRNAs frequently function in translation. Results suggest that opposite strand coding is exceptional in modern genes, yet might be frequent for mitochondrial tRNAs. This adds antisense tRNA templating to other mitochondrial tRNA functions: sense tRNA templating, formation and regulation of secondary (light strand DNA) replication origins. Antitermination probably affects mitochondrial degenerative diseases and ageing: pathogenic mutations are twice as frequent in tRNAs with antisense Ter anticodons than in other tRNAs, and species lacking mitochondrial antisense Ter tRNAs have longer mean maximal lifespans than those possessing antisense Ter tRNAs.     

The mitochondrial genome of the brachiopod Laqueus rubellus

The complete nucleotide sequence of the 14,017-bp mitochondrial (mt) genome of the articulate brachiopod Laqueus rubellus is presented. Being one of the smallest of known mt genomes, it has an extremely compact gene organization. While the same 13 polypeptides, two rRNAs, and 22 tRNAs are encoded as in most other animal mtDNAs, lengthy noncoding regions are absent, with the longest apparent intergenic sequence being 54 bp in length. Gene-end sequence overlaps are prevalent, and several stop codons are abbreviated. The genes are generally shorter, and three of the protein-coding genes are the shortest among known homologues. All of the tRNA genes indicate size reduction in either or both of the putative TPsiC and DHU arms compared with standard tRNAs. Possession of a TV (TPsiC arm-variable loop) replacement loop is inferred for tRNA(R) and tRNA(L-tag). The DHU arm appears to be unpaired not only in tRNA(S-tct) and tRNA(S-tga), but also in tRNA(C), tRNA(I), and tRNA(T), a novel condition. All the genes are encoded in the same DNA strand, which has a base composition rich in thymine and guanine. The genome has an overall gene arrangement drastically different from that of any other organisms so far reported, but contains several short segments, composed of 2-3 genes, which are found in other mt genomes. Combined cooccurrence of such gene assortments indicates that the Laqueus mt genome is similar to the annelid Lumbricus, the mollusc Katharina, and the octocoral Sarcophyton mt genomes, each with statistical significance. Widely accepted schemes of metazoan phylogeny suggest that the similarity with the octocoral could have arisen through a process of convergent evolution, while it appears likely that the similarities with the annelid and the mollusc reflect phylogenetic relationships.     

Trichinella spiralis mtDNA: a nematode mitochondrial genome that encodes a putative ATP8 and normally structured tRNAS and has a gene arrangement relatable to those of coelomate metazoans

The complete mitochondrial DNA (mtDNA) of the nematode Trichinella spiralis has been amplified in four overlapping fragments and 16,656 bp of its sequence has been determined. This sequence contains the 37 genes typical of metazoan mtDNAs, including a putative atp8, which is absent from all other nematode mtDNAs examined. The genes are transcribed from both mtDNA strands and have an arrangement relatable to those of coelomate metazoans, but not to those of secernentean nematodes. All protein genes appear to initiate with ATN codons, typical for metazoans. Neither TTG nor GTT start codons, inferred for several genes of other nematodes, were found. The 22 T. spiralis tRNA genes fall into three categories: (i) those with the potential to form conventional "cloverleaf" secondary structures, (ii) those with TPsiC arm + variable arm replacement loops, and (iii) those with DHU-arm replacement loops. Mt-tRNA(R) has a 5'-UCG-3' anticodon, as in most other metazoans, instead of the very unusual 5'-ACG-3' present in the secernentean nematodes. The sequence also contains a large repeat region that is polymorphic in size at the population and/or individual level.     

The complete nucleotide sequence and gene organization of the mitochondrial genome of the oriental mole cricket, Gryllotalpa orientalis (Orthoptera: Gryllotalpidae)

The complete nucleotide sequences of the mitochondrial genome (mitogenome) of the oriental mole cricket, Gryllotalpa orientalis (Orthoptera: Gryllotalpidae), were determined. The 15,521-bp-long G. orientalis mitogenome contains typical gene complement, base composition, and codon usage found in metazoan mitogenomes. The G. orientalis mitogenome contains the third lowest A+T content (70.5%) among the complete insects mt genome sequences. The initiation codon for the G. orientalis COI gene appears to be ATG, instead of the tetranucleotides, which have been postulated to act as initiation codon for Locusta migratoria and some lepidopteran COI genes. The initiation codon for ND2 appears to be GTG, which is rare, but has been designated as an initiator of Tricholepidion gertschi ND2. All anticodons of G. orientalis tRNAs were identical to Drosophila yakuba and L. migratoria. The tRNA(Ser)(AGN) could not form a stable stem loop structure in the DHU arm as shown in many other insect tRNA(Ser)(AGN). Phylogenetic analysis of nucleotide sequence information from all mt genes supported a monophyletic Diptera, a monophyletic Lepidoptera, a monophyletic Coleoptera, a monophyletic Mecopterida (Diptera+Lepidoptera), and a monophyletic Endopterygota (Diptera+Lepidoptera+Coleoptera), suggesting that the complete insect mitogenome sequence has a resolving power to the diversification events within Endopterygota. However, the relationships of ancient insect orders were unstable, indicating the limited use of mitogenome information at deeper phylogenetic depth.