Cytochrome B

THE rate of mitochondrial DNA (mtDNA) evolution has been carefully calibrated only in primates 1 . Similarity between the primate calibration and rates estimated for other vertebrates 2–4 has led to widespread assumption of a constant molecular clock in vertebrates even though ...

Because of functional constraints, substitution rates vary among the positions of a protein but are usually assumed to be constant at a given site during evolution. The distribution of the rates across the sequence positions generally fits a Gamma distribution. Models of sequence evolution were accordingly designed and led to improved phylogenetic reconstruction. However, it has been convincingly demonstrated that the evolutionary rate of a given position is not always constant throughout time. We called such within-site rate variations heterotachy (for "different speed" in Greek). Yet, heterotachy was found among homologous sequences of distantly related organisms, often with different functions. In such cases, the functional constraints are likely different, which would explain the different distribution of variable sites. To evaluate the importance of heterotachy, we focused on amino acid sequences of mitochondrial cytochrome b, for which the function is likely the same...

Mitochondrial DNA sequences are often used to construct molecular phylogenetic trees among closely related animals. In order to examine the usefulness of mtDNA sequences for deep-branch phylogenetics, genes in previously reported mtDNA sequences were analyzed among several animals that diverged 20–600 million years ago. Unambiguous alignment was achieved for stem-forming regions of mitochondrial tRNA genes by virtue of their conservative secondary structures. Sequences derived from stem parts of the mitochondrial tRNA genes appeared to accumulate much variation linearly for a long period of time: nearly 100 Myr for transition differences and more than 350 Myr for transversion differences. This characteristic could be attributed, in part, to the structural variability of mitochondrial tRNAs, which have fewer restrictions on their tertiary structure than do nonmitochondrial tRNAs. The tRNA sequence data served to reconstruct a well-established phylogeny of the animals with 100% bootstrap probabilities by both maximum parsimony and neighbor joining methods. By contrast, mitochondrial protein genes coding for cytochrome b and cytochrome oxidase subunit I did not reconstruct the established phylogeny or did so only weakly, although a variety of fractions of the protein gene sequences were subjected to tree-building. This discouraging phylogenetic performance of mitochondrial protein genes, especially with respect to branches originating over 300 Myr ago, was not simply due to high randomness in the data. It may have been due to the relative susceptibility of the protein genes to natural selection as compared with the stem parts of mitochondrial tRNA genes. On the basis of these results, it is proposed that mitochondrial tRNA genes may be useful in resolving deep branches in animal phylogenies with divergences that occurred some hundreds of Myr ago. For this purpose, we designed a set of primers with which mtDNA fragments encompassing clustered tRNA genes were successfully amplified from various vertebrates by the polymerase chain reaction.

Phylogenetic relationships among South American sigmodontine rodents were examined based on the complete sequence for the mitochondrial cytochrome b gene [1140 base pairs (bp)] for 66 species and between 759 and 1140 bp for an additional 19 species. Thirty-eight South American genera were represented, coming from eight of nine tribes. Outgroups included the North American murid rodents Peromyscus, Reithrodontomys, Scotinomys, and Neotoma, the Old World murine rodents Mus and Rattus, and the geomyoid genera Thomomys, Geomys, Dipodomys, and Perognathus as the most distant outgroup. The South American sigmodontines were supported as a monophyletic lineage. Within this radiation several clear-cut suprageneric groupings were identified. Many of the currently recognized tribal groupings of genera were found fairly consistently, although not always with high levels of bootstrap support. The various tribes could not be linked hierarchically with any confidence. In addition, several genera stand out as unique entities, without any apparent close relatives. The overall pattern suggests a rapid radiation of the sigmodontines in South America, followed by differentiation at the tribal and generic levels.

A strictly anaerobic bacterium dechlorinating tetrachloroethene (perchloroethylene, PCE) via trichloroethene (TCE) to cis-1, 2-dichloroethene (DCE) was isolated from activated sludge with pyruvate plus PCE as energy substrates. The organism, called Dehalospirillum ...

Two previous X-ray structures of antimycin bound to vertebrate bc 1 complex gave conflicting results. A new structure reported here of the bovine mitochondrial bc 1 complex at 2.28 Å resolution with antimycin bound, allows us for the first time to reliably describe the binding of ...

To investigate the potential role of somatic mitochondrial DNA (mtDNA) mutations in tumorigenesis, the occurrence of mutations in mtDNA of ovarian carcinomas was studied. We sequenced the D-loop region of mtDNA of 15 primary ovarian carcinomas and their matched normal controls. Somatic mtDNA mutations were detected in 20% (3 of 15) tumor samples carrying single or multiple changes. Complete sequence analysis of the mtDNA genomes of another 10 pairs of primary ovarian carcinomas and control tissues revealed somatic mtDNA mutations in 60% (6 of 10) of tumor samples. Most of these mutations were homoplasmic, and most were T-->C or G-->A transitions, but one represented a differential length within a run of identical C residues. A region of mtDNA sequence including the 16S and 12S rRNA genes, the D-loop and the cytochrome b gene, may represent the zone of preferred mtDNA mutation in ovarian cancer. The high incidence of mtDNA mutations found in ovarian carcinomas and other human c...

Noninvasive methods using genetic markers have been suggested as ways to overcome difficulties associated with documenting the presence of elusive species. We present and assess a novel, reliable and effective molecular genetic technique for the unequivocal genetic identification of faeces from the endangered Iberian lynx (Lynx pardinus). From mitochondrial DNA (mtDNA) cytochrome b and D-loop region sequences, we designed four species-specific primers (for products 130-161 bp long) that were considered to be likely to amplify degraded DNA. We compared two DNA extraction methods, various DNA amplification conditions and the robustness and specificity of the primer pairs with 87 lynx samples from 5 potentially different lynx populations and with 328 samples of other carnivore species. The utility of the identification technique was tested with faeces of different ages, with faeces from controlled field experiments, and with faeces collected from locales with possible lynx populations from throughout the state of Andalusia, Spain (8052 km2). Faecal mtDNA extraction was more efficient using PBS wash of the faeces instead of a faeces homogenate. Our assay increased from 92.6 to 99% efficiency with a second amplification and a reduction in template concentration to overcome polymerase chain reaction (PCR) inhibition. Our assay never produced false positives, and correctly identified all lynx faeces. Of 252 faeces samples of unknown species collected throughout Andalusia, 26.6% (from three different areas) were classified as Iberian lynx, 1.4% showed evidence of PCR inhibition and 1.2% were of uncertain origin. This method has proven to be a reliable technique that can be incorporated into large-scale surveys of Iberian lynx populations and exemplifies an approach that can easily be extended to other species.

To obtain information relative to the phylogenesis and microevolutionary rate of fish mitochondrial DNA, the nucleotide sequence of cytochrome b gene in seven fish species belonging to the order of Perciformes was determined. Sequence analysis showed that fish mitochondrial DNA has a nucleotide compositional bias similar to that of sharks but lower compared to mammals and birds. Quantitative evolutionary analysis, carried out by using a markovian stochastic model, clarifies some phylogenetic relationships within the Perciformes order, particularly in the Scombridae family, and between Perciformes, Gadiformes, Cypriniformes, and Acipenseriformes. The molecular clock of mitochondrial DNA was calibrated with the nucleotide substitution rate of cytochrome b gene in five shark species having divergence times inferred from paleontological estimates. The results of such analysis showed that Acipenseriformes diverged from Perciformes by about 200 MY, that the Perciformes common ancestor dates back to 150 MY, and that fish mitochondrial DNA has a nucleotide substitution rate three to five times lower than that of mammals.

 Population genetic theory predicts that marine animal species with planktonic larvae will have less genetic structure than those with direct development. We compared the genetic structure of four species of littorinid snails – two with planktonic egg capsules that hatch as planktonic larvae and two with benthic egg masses that hatch as crawl-away juveniles. We used DNA sequencing and single stranded conformational polymorphism (SSCP) to assess sequence variation in a 480 bp fragment of the mitochondrial cytochrome b gene and then used an analysis of molecular variance (AMOVA) to estimate Φst for populations from the northeastern Pacific coast. One of the two direct-developing species, Littorina subrotundata, had a moderate amount of population structure (Φst=0.209) as expected but the other direct-developing species, L. sitkana, was nearly fixed for a single haplotype that made it impossible to precisely estimate Φst. One of the two planktonic-developing species, L. scutulata, did not show any significant population structure (Φst=0.004). In contrast to our expectations, the other planktonic-developing species, L. plena, showed some weak but statistically significant population structure (Φst=0.052). We discuss how differences in population genetic structure between species with the same type of development may reflect differences in their historical demography.

The entire mitochondrial DNA control region (mtDNA D-loop) was sequenced in the seven extant species of Alectoris partridges. The D-loop length is very conserved (1155 ± 2 nucleotides), and substitution rates are lower than for the mitochondrial cytochrome b gene of the same species, on average. Comparative analyses suggest that these D-loops can be divided into three domains, corresponding to the highly variable peripheral domains I and III and to the central conserved domain II of vertebrates (Baker and Marshall 1997). Nevertheless, the first 161 nucleotides of domain I of the Alectoris, immediately flanking the tRNAGlu, evolve at an unusually low rate and show motifs similar to the mammalian extended termination-associated sequences [ETAS1 and ETAS2 (Sbisà et al. 1997)], which can form stable secondary structures. The second part of domain I contains a hypervariable region with two divergent copies of a tandemly repeated sequence described previously in other species of anseriforms and galliforms (Quinn and Wilson 1993; Fumihito et al. 1995). Some of the conserved sequence blocks of mammals can be mapped in the central domain of Alectoris. Domain III is highly variable and has sequences similar to mammalian CSB1. The bidirectional transcription promoter HSP/LSP box of the chicken is partially conserved among the Alectoris. This structural organization can be found in the anseriform and galliform species studied so far, suggesting that strong functional constraints might have controlled the evolution of the D-loop since the origin of Galloanserae. Their conserved organization and slow molecular evolution make D-loops of galliforms appropriate for phylogenetic studies, although homoplasy can be be generated at a few hypervariable sites and at some sites which probably have mutated by strand slippage during DNA replication. Phylogenetic analyses of D-loops of Alectoris are concordant with previously published cytochrome b and allozyme phylogenies (Randi 1996). Alectoris is monophyletic and includes three major clades: (1) basal barbara and melanocephala; (2) intermediate rufa and graeca; and (3) recent philbyi, magna, and chukar. Comparative description of the organization and substitution patterns of the mitochondrial control region can aid in mapping hypervariable sites and avoid some sources of homoplasy in data sets which are to be used in phylogenetic analyses.