Structure and evolution of the avian mitochondrial control region

The structural and evolutionary characteristics of the mitochondrial control region were studied by using control region sequences of 68 avian species. The distribution of the variable nucleotide positions within the control region was found to be genus specific and not dependant on the level of divergence, as suggested before. Saturation was shown to occur at the level of divergence of 10% in pairwise comparisons of the control region sequences, as has also been reported for the third codon positions in ND2 and cytochrome b genes of mtDNA. The ratio of control region vs cytochrome b divergence in pairwise comparisons of the sequences was shown to vary from 0.13 to 21.65, indicating that the control region is not always the most variable region of the mtDNA, but also that there are differences in the rate of divergence among the lineages. Only two of the conserved sequence blocks localized earlier for other species, D box and CSB-1, were found to show a considerable amount of sequence conservation across the avian and mammalian sequences. Additionally, a novel avian-specific sequence block was found.     

Sequence and properties of the human KB cell and mouse L cell D-loop regions of mitochondrial DNA.

The sequences of the displacement-loop (D-loop) regions of mitochondrial DNA (mtDNA) from mouse L cells and human KB cells have been determined and provide physical maps to aid in the identification of sequences involved in the regulation of replication and expression of mammalian mtDNA. Both D-loop regions are bounded by the genes for tRNAPhe and tRNAPro. This region contains the most highly divergent sequences in mtDNA with the exceptions of three small conserved sequence blocks near the 5' ends of D-loop strands, a 225 nucleotide conserved sequence block in the center of the D-loop strand template region, and a short sequence associated with the 3' ends of D-loop strands. A sequence similar to that associated with the 3' termini of D-loop strands overlaps one of the conserved sequence blocks near the 5' ends of D-loop strands. The large, central conserved sequence probably does not code for a protein since no open reading frames are discretely conserved. Numerous symmetric sequences and potential secondary structures exist in these sequences, but none appear to be clearly conserved between species.     

The main regulatory region of mammalian mitochondrial DNA: Structure-function model and evolutionary pattern

Summary The evolution of the main regulatory region (D-loop) of the mammalian mitochondrial genome was analyzed by comparing the sequences of eight mammalian species: human, common chimpanzee, pygmy chimpanzee, dolphin, cow, rat, mouse, and rabbit. The best alignment of the sequences was obtained by optimization of the sequence similarities common to all these species.The two peripheral left and right D-loop domains, which contain the main regulatory elements so far discovered, evolved rapidly in a species-specific manner generating heterogeneity in both length and base composition. They are prone to the insertion and deletion of elements and to the generation of short repeats by replication slippage. However, the preservation of some sequence blocks and similar cloverleaf-like structures in these regions, indicates a basic similarity in the regulatory mechanisms of the mitochondrial genome in all mammalian species.We found, particularly in the right domain, significant similarities to the telomeric sequences of the mitochondrial (mt) and nuclear DNA ofTetrahymena thermophila. These sequences may be interpreted as relics of telomeres present in ancestral linear forms of mtDNA or may simply represent efficient templates of RNA primase-like enzymes.Due to their peculiar evolution, the two peripheral domains cannot be used to estimate in a quantitative way the genetic distances between mammalian species. On the other hand the central domain, highly conserved during evolution, behaves as a good molecular clock.Reliable estimates of the times of divergence between closely and distantly related species were obtained from the central domain using a Markov model and assuming nonhomogeneous evolution of nucleotide sites.     

Efficient site-specific cleavage by RNase MRP requires interaction with two evolutionarily conserved mitochondrial RNA sequences.

RNase MRP is a site-specific endonuclease that processes primer mitochondrial RNA from the leading-strand origin of mitochondrial DNA replication. Using deletional analysis and saturation mutagenesis, we have determined the substrate requirements for cleavage by mouse mitochondrial RNase MRP. Two regions of sequence homology among vertebrate mitochondrial RNA primers, conserved sequence blocks II and III, were found to be critical for both efficient and accurate cleavage; a third region of sequence homology, conserved sequence block I, was dispensable. Analysis of insertion and deletion mutations within conserved sequence block II demonstrated that the specificity of RNase MRP accommodates the natural sequence heterogeneity of conserved sequence block II in vivo. Heterologous assays with human RNase MRP and mutated mouse mitochondrial RNA substrates indicated that sequences essential for substrate recognition are conserved between mammalian species.     

Rates of nuclear and cytoplasmic mitochondrial DNA sequence divergence in mammals

Differential rates of nucleotide substitution among different gene segments and between distinct evolutionary lineages is well documented among mitochondrial genes and is likely a consequence of locus-specific selective constraints that delimit mutational divergence over evolutionary time. We compared sequence variation of 18 homologous loci (15 coding genes and 3 parts of the control region) among 10 mammalian mitochondrial DNA genomes which allowed us to describe different mitochondrial evolutionary patterns and to produce an estimation of the relative order of gene divergence. The relative rates of divergence of mitochondrial DNA genes in the family Felidae were estimated by comparing their divergence from homologous counterpart genes included in nuclear mitochondrial DNA (Numt, pronounced "new might"), a genomic fossil that represents an ancient transfer of 7.9 kb of mitochondrial DNA to the nuclear genome of an ancestral species of the domestic cat (Felis catus). Phylogenetic analyses of mitochondrial (mtDNA) sequences with multiple outgroup species were conducted to date the ancestral node common to the Numt and the cytoplasmic (Cymt) mtDNA genes and to calibrate the rate of sequence divergence of mitochondrial genes relative to nuclear homologous counterparts. By setting the fastest substitution rate as strictly mutational, an empirical "selective retardation index" is computed to quantify the sum of all constraints, selective and otherwise, that limit sequence divergence of mitochondrial gene sequences over time.      

Human Mitochondrial DNA Variation and Evolution: Analysis of Nucleotide Sequences from Seven Individuals

We have analyzed nucleotide sequence variation in an approximately 900-base pair region of the human mitochondrial DNA molecule encompassing the heavy strand origin of replication and the D-loop. Our analysis has focused on nucleotide sequences available from seven humans. Average nucleotide diversity among the sequences is 1.7%, several-fold higher than estimates from restriction endonuclease site variation in mtDNA from these individuals and previously reported for other humans. This disparity is consistent with the rapidly evolving nature of this noncoding region. However, several instances of convergent or parallel gain and loss of restriction sites due to multiple substitutions were observed. In addition, other results suggest that restriction site (as well as pairwise sequence) comparisons may underestimate the total number of substitutions that have occurred since the divergence of two mtDNA sequences from a common ancestral sequence, even at low levels of divergence. This emphasizes the importance of recognizing the large standard errors associated with estimates of sequence variability, particularly when constructing phylogenies among closely related sequences. Analysis of the observed number and direction of substitutions revealed several significant biases, most notably a strand dependence of substitution type and a 32-fold bias favoring transitions over transversions. The results also revealed a significantly nonrandom distribution of nucleotide substitutions and sequence length variation. Significantly more multiple substitutions were observed than expected for these closely related sequences under the assumption of uniform rates of substitution. The bias for transitions has resulted in predominantly convergent or parallel changes among the observed multiple substitutions. There is no convincing evidence that recombination has contributed to the mtDNA sequence diversity we have observed.     

Nucleotide sequence of a Xenopus laevis mitochondrial DNA fragment containing the D-loop, flanking tRNA genes and the apocytochrome b gene

Extensive corrections of the nucleotide sequence of the Xenopus laevis mitochondrial (mt) displacement (D) loop and surrounding genes [Wong et al., Nucl. Acids Res. 11 (1983) 4977-4995] are reported, including addition of two stretches of nucleotides and 60 scattered modifications. The additional sequences presented here correspond to the apocytochrome b gene, the tRNAGlu gene and part of URF6. This allows us to propose a conformational model for the X. laevis apocytochrome b protein and also permits comparisons with mammalian mtDNA. The D-loop sequence is poorly conserved except for sequences involved in the regulation of the mt genome (conserved sequence blocks and the DNA polymerase stop sequences). On the other hand, all genes show marked conservation both of their nucleotide sequence and their respective location on the mt genome. Organization of the genetic information described for mammalian mtDNA also holds for the X. laevis mtDNA. This result strongly suggests that all animal vertebrate mtDNAs have followed the same evolutionary pathway.     

Origin and evolution of homologous repeated sequences in the mitochondrial DNA control region of shrews

The complete mitochondrial DNA (mtDNA) control region was amplified and directly sequenced in two species of shrew, Crocidura russula and Sorex araneus (Insectivora, Mammalia). The general organization is similar to that found in other mammals: a central conserved region surrounded by two more variable domains. However, we have found in shrews the simultaneous presence of arrays of tandem repeats in potential locations where repeats tend to occur separately in other mammalian species. These locations correspond to regions which are associated with a possible interruption of the replication processes, either at the end of the three-stranded D-loop structure or toward the end of the heavy-strand replication. In the left domain the repeated sequences (R1 repeats) are 78 bp long, whereas in the right domain the repeats are 12 bp long in C. russula and 14 bp long in S. araneus (R2 repeats). Variation in the copy number of these repeated sequences results in mtDNA control region length differences. Southern blot analysis indicates that level of heteroplasmy (more than one mtDNA form within an individual) differs between species. A comparative study of the R2 repeats in 12 additional species representing three shrew subfamilies provides useful indications for the understanding of the origin and the evolution of these homologous tandemly repeated sequences. An asymmetry in the distribution of variants within the arrays, as well as the constant occurrence of shorter repeated sequences flanking only one side of the R2 arrays, could be related to asymmetry in the replication of each strand of the mtDNA molecule. The pattern of sequence and length variation within and between species, together with the capability of the arrays to form stable secondary structures, suggests that the dominant mechanism involved in the evolution of these arrays in unidirectional replication slippage.