Allan C. Wilson

EVOLUTION OF THE CYTOCHROME B GENE OF MAMMALS

SummaryWith the polymerase chain reaction (PCR) and versatile primers that amplify the whole cytochromeb gene (∼ 1140 bp), we obtained 17 complete gene sequences representing three orders of hoofed mammals (ungulates) and dolphins (cetaceans). The fossil record of some ungulate lineages allowed estimation of the evolutionary rates for various components of the cytochromeb DNA and amino acid sequences. The relative rates of substitution at first, second, and third positions within codons are in the ratio 10 to 1 to at least 33. For deep divergences (>5 million years) it appears that both replacements and silent transversions in this mitochondrial gene can be used for phylogenetic inference. Phylogenetic findings include the association of (1) cetaceans, artiodactyls, and perissodactyls to the exclusion of elephants and humans, (2) pronghorn and fallow deer to the exclusion of bovids (i. e., cow, sheep, and goat), (3) sheep and goat to the exclusion of other pecorans (i. e., cow, giraffe, deer, and pronghorn), and (4) advanced ruminants to the exclusion of the chevrotain and other artiodactyls. Comparisons of these cytochromeb sequences support current structure-function models for this membrane-spanning protein. That part of the outer surface which includes the Qo redox center is more constrained than the remainder of the molecule, namely, the transmembrane segments and the surface that protrudes into the mitochondrial matrix. Many of the amino acid replacements within the transmembrane segments are exchanges between hydrophobic residues (especially leucine, isoleucine, and valine). Replacement changes at first and second positions of codons approximate a negative binomial distribution, similar to other protein-coding sequences. At four-fold degenerate positions of codons, the nucleotide substitutions approximate a Poisson distribution, implying that the underlying mutational spectrum is random with respect to position.

Mitochondrial DNA sequences of primates: Tempo and mode of evolution

SummaryWe cloned and sequenced a segment of mitochondrial DNA from human, chimpanzee, gorilla, orangutan, and gibbon. This segment is 896 bp in length, contains the genes for three transfer RNAs and parts of two proteins, and is homologous in all 5 primates. The 5 sequences differ from one another by base substitutions at 283 positions and by a deletion of one base pair. The sequence differences range from 9 to 19% among species, in agreement with estimates from cleavage map comparisons, thus confirming that the rate of mtDNA evolution in primates is 5 to 10 times higher than in nuclear DNA. The most striking new finding to emerge from these comparisons is that transitions greatly outnumber transversions. Ninety-two percent of the differences among the most closely related species (human, chimpanzee, and gorilla) are transitions. For pairs of species with longer divergence times, the observed percentage of transitions falls until, in the case of comparisons between primates and non-primates, it reaches a value of 45. The time dependence is probably due to obliteration of the record of transitions by multiple substitutions at the same nucleotide site. This finding illustrates the importance of choosing closely related species for analysis of the evolutionary process. The remarkable bias toward transitions in mtDNA evolution necessitates the revision of equations that correct for multiple substitutions at the same site. With revised equations, we calculated the incidence of silent and replacement substitutions in the two protein-coding genes. The silent substitution rate is 4 to 6 times higher than the replacement rate, indicating strong functional constraints at replacement sites. Moreover, the silent rate for these two genes is about 10% per million years, a value 10 times higher than the silent rate for the nuclear genes studied so far. In addition, the mean substitution rate in the three mitochondrial tRNA genes is at least 100 times higher than in nuclear tRNA genes. Finally, genealogical analysis of the sequence differences supports the view that the human lineage branched off only slightly before the gorilla and chimpanzee lineages diverged and strengthens the hypothesis that humans are more related to gorillas and chimpanzees than is the orangutan.

Evolution in bacteria: evidence for a universal substitution rate in cellular genomes.

This paper constructs a temporal scale for bacterial evolution by tying ecological events that took place at known times in the geological past to specific branch points in the genealogical tree relating the 16S ribosomal RNAs of eubacteria, mitochondria, and chloroplasts. One thus obtains a relationship between time and bacterial RNA divergence which can be used to estimate times of divergence between other branches in the bacterial tree. According to this approach, Salmonella typhimurium and Escherichia coli diverged between 120 and 160 million years (Myr) ago, a date which fits with evidence that the chief habitats occupied now by these two enteric species became available that long ago. The median extent of divergence between S. typhimurium and E. coli at synonymous sites for 21 kilobases of protein-coding DNA is 100%. This implies a silent substitution rate of 0.7-0.8%/Myr--a rate remarkably similar to that observed in the nuclear genes of mammals, invertebrates, and flowering plants. Similarities in the substitution rates of eucaryotes and procaryotes are not limited to silent substitutions in protein-coding regions. The average substitution rate for 16S rRNA in eubacteria is about 1%/50 Myr, similar to the average rate for 18S rRNA in vertebrates and flowering plants. Likewise, we estimate a mean rate of roughly 1%/25 Myr for 5S rRNA in both eubacteria and eucaryotes. For a few protein-coding genes of these enteric bacteria, the extent of silent substitution since the divergence of S. typhimurium and E. coli is much lower than 100%, owing to extreme bias in the usage of synonymous codons. Furthermore, in these bacteria, rates of amino acid replacement were about 20 times lower, on average, than the silent rate. By contrast, for the mammalian genes studied to date, the average replacement rate is only four to five times lower than the rate of silent substitution.

Immunological Time Scale for Hominid Evolution

Several workers have observed that there is an extremely close immunological resemblance between the serum albumins of apes and man. Our studies with the quantitative micro-complement fixation method confirm this observation. To explain the closeness of the resemblance, previous workers suggested that there has been a slowing down of albumin evolution since the time of divergence of apes and man. Recent evidence, however, indicates that the albumin molecule has evolved at a steady rate. Hence, we suggest that apes and man have a more recent common ancestry than is usually supposed. Our calculations lead to the suggestion that, if man and Old World monkeys last shared a common ancestor 30 million years ago, then man and African apes shared a common ancestor 5 million years ago, that is, in the Pliocene era.

Calibration of mitochondrial DNA evolution in geese

SummaryMitochondrial DNA was purified from five American species of geese representing the generaAnser andBranta, which have fossil records. The results of electrophoretic comparisons of about 75 fragments per individual produced by 14 restriction enzymes imply that the mean extent of sequence divergence between species ofAnser andBranta is about 9%. Fossil evidence suggests that these two groups of geese had a common ancestor 4–5 million years ago. Thus, the mean rate of sequence divergence in goose mitochondiral DNA is not far from 2% per million years, the value in mammals.

Molecular Evolution in Drosophila and the Higher Diptera II. A Time Scale for Fly Evolution

In this paper, we examine first the steadiness of the rate of evolutionary change in a larval hemolymph protein, LHP, in numerousDrosophila species. We estimated amino acid sequence divergence from immunological distances measured with the quantitative microcomplement fixation technique. Using tests not depending on knowledge of absolute times of divergence, we estimated the variance of the rate of evolutionary change to be at least 4 times as large as that for a process resembling radioactive decay. Thus, the rate of evolution of this protein is as uniform as that of vertebrate proteins. Our analysis indicates no acceleration of protein evolution in the lineages leading to Hawaiian drosophilines. Second, we give an explicit description of a procedure for calculating the absolute value of the mean rate of evolutionary change in this protein. This procedure is suggested for general use in calculating absolute rates of molecular evolution. The mean rate of evolution of LHP is about 1.2 immunological distance units per million years, which probably coreesponds to a unit evolutionary period of 4 million years; LHP thus evolves at a rate comparable to that of mammalian hemoglobins. Finally, we utilize the calibrated rate of LHP evolution to derive a time scale of evolution in the Drosophilidae and higher Diptera.SummaryIn this paper, we examine first the steadiness of the rate of evolutionary change in a larval hemolymph protein, LHP, in numerousDrosophila species. We estimated amino acid sequence divergence from immunological distances measured with the quantitative microcomplement fixation technique. Using tests not depending on knowledge of absolute times of divergence, we estimated the variance of the rate of evolutionary change to be at least 4 times as large as that for a process resembling radioactive decay. Thus, the rate of evolution of this protein is as uniform as that of vertebrate proteins. Our analysis indicates no acceleration of protein evolution in the lineages leading to Hawaiian drosophilines. Second, we give an explicit description of a procedure for calculating the absolute value of the mean rate of evolutionary change in this protein. This procedure is suggested for general use in calculating absolute rates of molecular evolution. The mean rate of evolution of LHP is about 1.2 immunological distance units per million years, which probably coreesponds to a unit evolutionary period of 4 million years; LHP thus evolves at a rate comparable to that of mammalian hemoglobins. Finally, we utilize the calibrated rate of LHP evolution to derive a time scale of evolution in the Drosophilidae and higher Diptera.

Mitochondrial Resolution of a Deep Branch in the Genealogical Tree for Perching Birds

Animal mitochondrial DNA (mtDNA) is known to contain information about the genealogical relations among closely related species and is shown here to yield information about distant relations as well. Our results also draw attention to the need for caution in using third positions of codons for tree construction. This is evident from comparative studies of the cytochrome b gene in 13 species representing major groups within the order of perching birds (Passeriformes). Sequences of a 924 base-pair segment of this gene were obtained from each of these species via the polymerase chain reaction and a novel set of versatile primers. With a woodpecker sequence as an outgroup, trees that separate songbirds from other perching birds and resolve the ancient branch leading to songbirds were obtained utilizing the conservative first and second positions of codons. Analysis of positions within codons suggests that, for deep branches, the skewed base composition at the fast-changing third positions can result in phylogenetic disinformation, which conflicts with the information retained in the first and second positions. The mitochondrial tree shows broad concordance with that based on hybridization of nuclear DNA; however, parsimony and maximum likelihood methods suggest a close kinship between thrushes and Australian babblers, in agreement with the traditional morphological classification.

Ancient origin of lactalbumin from lysozyme: Analysis of DNA and amino acid sequences

SummaryParsimony trees relating DNA sequences coding for lysozymesc and α-lactalbumins suggest that the gene duplication that allowed lactalbumin to evolve from lysozyme preceded the divergence of mammals and birds. Comparisons of the amino acid sequences of additional lysozymes and lactalbumins are consistent with this view. When all base positions are considered, the probability that the duplication leading to the lactalbumin gene occurred after the start to mammalian evolution is estimated to be 0.05–0.10. Elimination of the phylogenetic noise generated by fast evolution and compositional bias at third positions of codons reduced this probability to 0.002–0.03. Thus the gene duplication may have long preceded the acquisition of lactalbumin function.

Molecular time scale for evolution

Abstract Molecular evolutionary clocks have ticked at much the same rate per year in many eubacterial genes as in the nuclear genes of animals and plants. This implication emerges from comparative studies of ribosomal RNA and protein-coding genes. The existence of nearly universal molecular clocks in cellular genomes provides a challenge to explain them as well as an opportunity to use them for putting knowledge of biological diversity on a temporal framework.

Wormy mice in a hybrid zone.

As one approach to analysing the genetic barriers between species, we studied the numbers and types of parasitic worms in two species of house mice (Mus musculus and M. domesticus) and in their natural hybrids. Where the ranges of these two species meet in southern Germany, there is a zone of hybridization less than 20 kilometres across1, in which about 98% of the mice have backcross genotypes. Fourteen of the 46 mice tested from within the zone have over 500 pinworms per gut, a number far exceeding the mean of 40 per gut for other mice inside and outside the zone. Other nematodes have a similar, non-random distribution. The number of mice bearing 9 or more tapeworms per gut is also excessive in the hybrid zone. These extraordinarily wormy mice may be unusually susceptible to parasitism; the different species may have different genes for resistance, and recombinant backcross animals may lose both2. Our findings support the view that the hybrid populations may have reduced fitness and thereby act as a genetic sink, interfering with the flow of genes between the two species1,3. The possibility that environmental or ecological peculiarities in the zone of hybridization make the mice more liable to infection is not supported.