A likelihood ratio test for evolutionary rate shifts and functional divergence among proteins.

Changes in protein function can lead to changes in the selection acting on specific residues. This can often be detected as evolutionary rate changes at the sites in question. A maximum-likelihood method for detecting evolutionary rate shifts at specific protein positions is presented. The method determines significance values of the rate differences to give a sound statistical foundation for the conclusions drawn from the analyses. A statistical test for detecting slowly evolving sites is also described. The methods are applied to a set of Myc proteins for the identification of both conserved sites and those with changing evolutionary rates. Those positions with conserved and changing rates are related to the structures and functions of their proteins. The results are compared with an earlier Bayesian method, thereby highlighting the advantages of the new likelihood ratio tests.

Molecular and morphological supertrees for eutherian (placental) mammals.

A large body of diverse comparative data now exists for a major phylogenetic synthesis of the higher-level relationships among eutherian (placental) mammals. We present such a phylogenetic synthesis using the composite trees or supertrees from the combined and separate analyses of their published molecular and morphological source phylogenies. Our combined and separate supertrees largely support the same suprafamilial taxa and orders, but different interordinal clades. These similarities and differences reinforce the continuing contributions of morphological studies, while highlighting the growing influence of molecular information on the field. As current summaries of past research, our supertrees emphasize opportunities for future work, while providing a step toward the eventual integration of the data and characters themselves.

Function-structure analysis of proteins using covarion-based evolutionary approaches: Elongation factors.

The divergent evolution of protein sequences from genomic databases can be analyzed by the use of different mathematical models. The most common treat all sites in a protein sequence as equally variable. More sophisticated models acknowledge the fact that purifying selection generally tolerates variable amounts of amino acid replacement at different positions in a protein sequence. In their "stationary" versions, such models assume that the replacement rate at individual positions remains constant throughout evolutionary history. "Nonstationary" covarion versions, however, allow the replacement rate at a position to vary in different branches of the evolutionary tree. Recently, statistical methods have been developed that highlight this type of variation in replacement rates. Here, we show how positions that have variable rates of divergence in different regions of a tree ("covarion behavior"), coupled with analyses of experimental three-dimensional structures, can provide experimentally testable hypotheses that relate individual amino acid residues to specific functional differences in those branches. We illustrate this in the elongation factor family of proteins as a paradigm for applications of this type of analysis in functional genomics generally.

Evolution of CpG islands within the myc gene family.

CpG islands are discrete regions of DNA with significantly greater frequencies of CpG doublets than bulk genomic DNA. They are most frequently associated with the 5'-ends of housekeeping genes and are involved in the regulation of their expression. In this study, the structure and evolution of CpG islands within genes of the myc family were evaluated with the protein-coding sequences of animals and their transducing viruses. These evaluations relied on a gene tree for the entire myc family to test the origins of CpG islands within their two protein-coding exons. Overall, CG-very rich and CG-rich islands are associated with exon 2 of the different myc genes of warm-blooded vertebrates and with exon 3 of the N-myc and s-myc sequences of mammals, but not birds. These overall distributions of well-developed islands can be related to the major transitions of the CG-rich genomes of warm-blooded vertebrates from the CG-poor ones of other animals. In turn, the greater variability of well-developed islands within exon 3 of the N-myc gene and among the different retrogenes of the myc family can be attributed to their reduced functional constraints, as evidenced by their limited and very restricted patterns of expression, respectively.

c-myc gene sequences and the phylogeny of bats and other eutherian mammals.

The complete protein-coding sequences of the c-myc proto-oncogene were determined for five species of four new orders of eutherian (placental) mammals. These newly obtained sequences were aligned to each other and to other available orthologs for the phylogenetic estimation of eutherian interordinal relationships. Several measures of sequence difference and base composition were first calculated to assess the major evolutionary properties of the three codon positions and two protein-coding exons of the gene. On the basis of these calculations, different parsimony, distance, and maximum likelihood approaches were adopted, with the most sophisticated involving the separate, then combined, likelihood analyses of the third codon positions of exon 2 versus all other sites. These phylogenetic approaches provided clear support for the grouping of Chiroptera (bats) with Artiodactyla (ruminants, camels, and pigs) and Carnivora (cats, dogs, and their allies), an interordinal arrangement that receives strong corroboration from other lines of evidence including complete mitochondrial DNA sequences. In contrast, these analyses failed to provide strong to reasonable support for any other interordinal group. This study concludes with specific recommendations about sampling and other strategies for maximizing the phylogenetic contributions of the c-myc gene to the continued resolution of the eutherian ordinal tree.

Molecular systematics: Perfect SINEs of evolutionary history?

Short interspersed repetitive elements - SINEs - are being championed as near-perfect phylogenetic characters; they have recently been used with notable success to resolve some phylogenetic conundrums, but they do have certain limitations that restrict their use as 'perfect' characters for molecular systematics.

Phylogenetic assessment of molecular and morphological data for eutherian mammals.

The interordinal relationships of eutherian (placental) mammals were evaluated by a phylogenetic analysis of four published data sets (three sequences and one morphological). The nature and degree of support and conflict for particular groups were assessed by separate bootstrap and homogeneity tests, which were followed by combined analyses of the sequence and morphological data. Between orders, strong support (i.e., > or = 95% bootstrap scores) was found for a paraphyletic Artiodactyla (relative to Cetacea) and a monophyletic Cetartiodactyla (Artiodactyla and Cetacea) and Paenungulata (Hyracoidea, Proboscidea, and Sirenia). In turn, some reasonable to strong evidence (> or = 85%) was obtained for Hyracoidea with Sirenia, Dermoptera with Scandentia, Glires (Lagomorpha with Rodentia), and Afrotheria (Amblysomus, Macroscelidea, Paenungulata, and Tubulidentata). Otherwise, no other interordinal clades were supported at these reasonable to strong levels. This overall lack of resolution for eutherian interordinal clusters agrees with other studies that suggest further progress will continue to be slow and difficult. Further resolution will require the integration of more recently published data, the continued sampling of taxa and characters, and the use of more powerful methods of data analysis.

Constraints on protein evolution and the age of the eubacteria/eukaryote split.

NASA: This article discusses research done by Doolittle et al. to determine the evolutionary distance between eubacteria and eukaryotes. Several equations for estimating evolutionary distances and divergence times are considered. Molecular clock calculations and their effects on these estimates are discussed. The authors conclude that their research supports the range of dates for the cenancestor of modern eubacteria, archaebacteria, and eukaryotes estimated by Doolittle et al.^ieng

A congruence study of molecular and morphological data for eutherian mammals.

The four orders of eutherian mammals which are traditionally placed in the superorder Archonta [Chiroptera (microbats and megabats), Dermoptera (flying lemurs), Primates (primates), and Scandentia (tree shrews)] are among the best-studied taxa of their infraclass from both the molecular and morphological perspectives. Nevertheless, the ordinal relationships of archontans remain unresolved. While morphological studies favor their monophyly, molecular investigations do not. To evaluate these opposing conclusions, parsimony analyses were conducted with three separate sets of DNA sequences from both the nuclear and mitochondrial genomes and one file of morphological data for archontans and other eutherian mammals. Statistical tests of character support and ordinal branching pattern differences documented that the three sets of DNA sequences and their results were homogeneous and congruent, thereby allowing for the combination of these data into one large matrix for further phylogenetic analysis. In contrast, these same tests revealed that the combined sequence and morphological files and their topologies were in strong conflict. Archontan monophyly was supported by the morphological evidence, but this arrangement was strongly rejected by the combined DNA sequences that favored instead a grouping of Dermoptera, Primates, and Scandentia with Lagomorpha (rabbits) and Rodentia (rodents). Resolution of these significant differences will require further evaluations about the homologies and evolutionary properties of the molecular and morphological characters and about the appropriateness of the chosen phylogenetic methods, as well as the incorporation of new comparative data from both sources.

Phylogeography and population structure of the Atlantic and Mediterranean green turtle Chelonia mydas: a mitochondrial DNA control region sequence assessment.

Mitochondrial (mt) DNA sequences were analysed to resolve the phylogeography and population genetic structure of Atlantic and Mediterranean populations of green turtles (Chelonia mydas). Analysis of sequence variation over 487 base pairs of the control (D-loop) region identified 18 haplotypes among 147 individuals from nine nesting populations. Pairwise comparisons of haplotype frequencies distinguished most nesting colonies, indicating significant genetic differentiation among rookeries and a strong propensity for natal homing behaviour by nesting females. Comparison of control region sequence data to earlier restriction fragment length polymorphism (RFLP) data for the same individuals demonstrates approximately a sixfold higher substitution rate in the 5' end of the control region. The sequence data provide higher resolution both in terms of the number of mtDNA genotype variants and the phylogeographic relationships detected within the Atlantic region, and reveal a gene genealogy that distinguishes two groups of haplotypes corresponding to (i) the western Caribbean and Mediterranean, and (ii) eastern Caribbean, South Atlantic and West Africa. The data suggest that phylogeographic patterns in the Atlantic Ocean may be interpreted in terms of female nest site fidelity and episodic dispersal events. The distribution of mtDNA haplotypes within the region is thus explained by the geological and climatic alternations (glacial and interglacial) over the last million years.

Support for interordinal eutherian relationships with an emphasis on primates and their archontan relatives.

Current knowledge about mammalian interordinal relationships is growing rapidly; thus this contribution is an attempt to summarize the past 5 years of this literature. We have focused on the recent controversies in mammalian phylogenetics including hypotheses concerning the monophyly of Archonta, the diphyly of Chiroptera, and the polyphyly of Rodentia. All of these issues have been proposed recently, challenging these phylogenetic hypotheses. We have attempted to include all of the comprehensive analyses of eutherian mammal systematics with an emphasis on morphological and molecular data sets where discrete characters are listed so they could be compiled and used in support of interordinal relationships. Particular attention is given to determining which of the living eutherian orders is the closest relative to primates. In reviewing relationships among the mammals, we have focused on collating all of the available evidence so that one could know where each of the specific data sets is in support of the various relationships.

Testing the covarion hypothesis of molecular evolution.

The covarion hypothesis of molecular evolution states that the fixation of mutations may alter the probability that any given position will fix the next change. Tests of this hypothesis using the divergence of real sequences are compromised because models of rate variation among sites (e.g., the gamma version of the one-parameter equation) predict sequence divergence values similar to those for the covarion process. This study therefore focuses on the extent to which the varied and unvaried codons of two well-diverged taxa are the same, because fewer are expected by the covarion hypothesis than by the gamma model. The data for these tests are the protein sequences of Cu, Zn superoxide dismutase (SOD) for mammals and plants. Simulation analyses show that the covarion hypothesis makes better predictions about the frequencies of varied and unhit positions in common between these two taxa than does the gamma version of the one-parameter model. Furthermore, the analysis of SOD tertiary structure demonstrates that mammal and plant variabilities are distributed differently on the protein. These results support the conclusions that the variable and invariable codons of mammal and plant SODs are different and that the covarion model explains the evolution of this protein better than the gamma version of the one-parameter process. Unlike other models, the covarion hypothesis accounts for rate fluctuations among positions over time, which is an important parameter of molecular evolution.

Molecular evolution and population genetics of Greater Caribbean green turtles (Chelonia mydas) as inferred from mitochondrial DNA control region sequences.

The molecular evolution and population genetics of migratory green turtles (Chelonia mydas) in the Greater Caribbean were examined with mitochondrial DNA (mtDNA) control region I sequences. A total of 488 base positions (bp) per individual were aligned for 44 individuals from four nesting populations in Florida, Costa Rica, Aves Island (Venezuela), and Surinam. Twelve sequence polymorphisms were detected, representing ten transitions, one transversion, and one 10-bp repeat. Sequence analyses of within- and between-population diversity revealed a deep divergence between western and eastern Caribbean nesting colonies and an inverse relationship between reproductive female population size and mtDNA diversity. In small populations, genetic admixture was important to maintaining high diversity, whereas larger populations appear to have experienced historical bottlenecks or resulted from founder effects. Mitochondrial DNA sequences of the control region offer an order of magnitude greater resolution than restriction site data for addressing questions about mtDNA variation, both within and between populations of green turtles.

DNA systematics and evolution of the artiodactyl family Bovidae.

Nine additional sequences from representatives of different tribes of the family Bovidae were combined with six published artiodactyl sequences to provide orthologous mtDNA for investigation of bovid phylogeny and evolution. Each species was represented by a homologous 2.7-kilobase-pair stretch of mtDNA for the complete 12S and 16S rRNA genes and three adjacent tRNA genes. These data, when compared to other results, provided evidence for a monophyletic Bovidae and for two clades within the family: one including the tribes Boselaphini, Bovini, and Tragelaphini and another for an Antilopini/Neotragini grouping. All other intrafamilial relationships were only weakly supported. These sequence comparisons suggest that most bovid tribes originated early in the Miocene with all extant lineages present by approximately 16-17 million years ago. Thus, bovid tribes provide an example of rapid cladogenesis, following the origin of families in the infraorder Pecora.

Phylogeny and evolution of antlered deer determined from mitochondrial DNA sequences.

Mitochondrial DNA sequences of both ribosomal RNA genes and three adjacent transfer RNA genes were obtained for the three extant subfamilies of antlered deer (Cervinae, Muntiacinae, and Odocoileinae) as well as for their antlerless sister group Hydropotinae (family Cervidae). Phylogenetic analysis of these sequences (each nearly 2.7 kilobase pairs in length) supports a cervine/muntiacine clade to the exclusion of odocoileines. These results are statistically significant, stable, and congruent with some independent data. Our mitochondrial DNA sequences, when coupled with other information, indicate that the earliest fossil antlered deer are not closely related to living muntiacines or any other contemporary subfamily. From this information, we hypothesize an Old World, Late Miocene origin of Odocoileinae.