16. Molecular Phylogenetics

Learning outcomes When you have read Chapter 16, you should be able to: Recount how taxonomy led to phylogeny and discuss the reasons why molecular markers are important in phylogenetics Describe the key features of a phylogenetic tree and distinguish between inferred trees, true trees, gene trees and species trees Explain how phylogenetic trees are reconstructed, including a description of DNA sequence alignment, the methods used to convert alignment data into a phylogenetic tree, and how the accuracy of a tree is assessed Discuss, with examples, the applications and limitations of molecular clocks Give examples of the use of phylogenetic trees in studies of human evolution and the evolution of the human and simian immunodeficiency viruses Describe how molecular phylogenetics is being used to study the origins of modern humans, and the migrations of modern humans into Europe and the New World

16.1. The Origins of Molecular Phylogenetics 16.2. The Reconstruction of DNA-based Phylogenetic Trees 16.3. The Applications of Molecular Phylogenetics

16.1. The Origins of Molecular Phylogenetics

Figure 16.1. The tree of life. An ancestral species is at the bottom of the ‘trunk' of the tree. As time passes, new species evolve from earlier ones so the tree repeatedly branches until we reach the present time, when there are many species descended from the ancestor.

Figure 16. 2. DNA yields more phylogenetic information than protein Figure 16.2. DNA yields more phylogenetic information than protein. The two DNA sequences differ at three positions but the amino acid sequences differ at only one position. These positions are indicated by green dots. Two of the nucleotide substitutions are therefore synonymous and one is non-synonymous (see Figure 14.11 ).

Box 16.1. Phenetics and cladistics

16.2. The Reconstruction of DNA-based Phylogenetic Trees

Figure 16. 3. Phylogenetic trees Figure 16.3. Phylogenetic trees. (A) An unrooted tree with four external nodes. (B) The five rooted trees that can be drawn from the unrooted tree shown in part A. The positions of the roots are indicated by the numbers on the outline of the unrooted tree.

Figure 16. 4. The use of an outgroup to root a phylogenetic tree Figure 16.4. The use of an outgroup to root a phylogenetic tree. The tree of human, chimpanzee, gorilla and orangutan genes is rooted with a baboon gene because we know from the fossil record that baboons split away from the primate lineage before the time of the common ancestor of the other four species. For more information on phylogenetic analysis of humans and other primates see Section 16.3.1.

Figure 16.5. The difference between a gene tree and a species tree

Figure 16.6. Mutation might precede speciation, giving an incorrect time for a speciation event if a molecular clock is used. See the text for details. Based on Li (1997).

igure 16.7. A gene tree can have a different branching order from a species tree. In this example, the gene has undergone two mutations in the ancestral species, the first mutation giving rise to the ‘blue' allele and the second to the ‘green' allele. Random genetic drift in association with the two subsequent speciations results in the red allele lineage appearing in species A, the green allele lineage in species B and the blue allele lineage in species C. Molecular phylogenetics based on the gene sequences will reveal that the red-blue split occurred before the blue-green split, giving the gene tree shown on the right. However, the actual species tree is different, as shown on the left. Based on Li (1997)

Figure 16. 8. Sequence alignment Figure 16.8. Sequence alignment. (A) Two sequences that have not diverged to any great extent can be aligned easily by eye. (B) A more complicated alignment in which it is not possible to determine the correct position for an indel. If errors in indel placement are made in a multiple alignment then the tree reconstructed by phylogenetic analysis is unlikely to be correct. In this diagram, the red asterisks indicate nucleotides that are the same in both sequences.

Figure 16. 9. The dot matrix technique for sequence alignment Figure 16.9. The dot matrix technique for sequence alignment. The correct alignment stands out because it forms a diagonal of continuous dots, broken at point mutations and shifting to a different diagonal at indels.

Figure 16. 10. A simple distance matrix Figure 16.10. A simple distance matrix. The matrix shows the evolutionary distance between each pair of sequences in the alignment. In this example the evolutionary distance is expressed as the number of nucleotide differences per nucleotide site for each sequence pair. For example, sequences 1 and 2 are 20 nucleotides in length and have four differences, corresponding to an evolutionary difference of 4/20 = 0.2. Note that this analysis assumes that there are no multiple substitutions (also called multiple hits). Multiple substitution occurs when a single site undergoes two or more changes (e.g. the ancestral sequence … ATGT … gives rise to two modern sequences: … AGGT … and … ACGT …). There is only one nucleotide difference between the two modern sequences, but there have been two nucleotide substitutions. If this multiple hit is not recognized then the evolutionary distance between the two modern sequences will be significantly underestimated. To avoid this problem, distance matrices for phylogenetic analysis are usually constructed using mathematical methods that include statistical devices for estimating the amount of multiple substitution that has occurred.

Figure 16.11. Manipulations carried out when using the neighbor-joining method for tree reconstruction. See the text for details.

Figure 16.12. Constructing a new multiple alignment in order to bootstrap a phylogenetic tree. The new alignment is built up by taking columns at random from the real alignment. Note that the same column can be sampled more than once

Figure 16. 13. Calculating a human molecular clock Figure 16.13. Calculating a human molecular clock. The number of substitutions is determined for a pair of homologous genes from human and orangutan: call this number ‘x'. The number of substitutions per lineage is therefore x/2, and the number per million years is x/2 × 13.

16.3. The Applications of Molecular Phylogenetics

Figure 16.14. Different interpretations of the evolutionary relationships between humans, chimpanzees and gorillas. See the text for details. Abbreviation: Myr, million years

Figure 16.15. The phylogenetic tree reconstructed from HIV and SIV genome sequences. The AIDS epidemic is due to the HIV-1M type of immunodeficiency virus. ZR59 is positioned near the root of the star-like pattern formed by genomes of this type. Based on Wain-Hobson (1998).

Figure 16.16. Two competing hypotheses for the origins of modern humans. (A) The multiregional hypothesis states that Homo erectus left Africa over 1 million years ago and then evolved into modern humans in different parts of the Old World. (B) The Out of Africa hypothesis states that the populations of Homo erectus in the Old World were displaced by new populations of modern humans that followed them out of Africa.

Figure 16.17. Phylogenetic tree reconstructed from mitochondrial RFLP data obtained from 147 modern humans. The ancestral mitochondrial DNA is inferred to have existed in Africa because of the split in the tree between the seven modern African mitochondrial genomes placed below the ancestral sequence and all the other genomes above it. Because this lower branch is purely African it is deduced that the ancestor was also African. The scale bars at the bottom indicate sequence divergence from which, using the mitochondrial molecular clock, it is possible to assign dates to the branch points in the tree. The clock suggests that the ancestral sequence existed between 140 000 and 290 000 years ago. Reprinted with permission from Cann et al. (1987)Nature, 325, 31–36. Copyright 1987 Macmillan Magazines Limited.

Figure 16.18. The spread of agriculture from the Middle East to Europe. The dark-green area is the ‘Fertile Crescent', the area of the Middle East where many of today's crops - wheat, barley, etc. - grow wild and where these plants are thought to have first been taken into cultivation.

Figure 16. 19. A genetic gradation across modern Europe Figure 16.19. A genetic gradation across modern Europe. See the text for details.

Figure 16. 20. The eleven major European mitochondrial haplotypes Figure 16.20. The eleven major European mitochondrial haplotypes. The calculated time of origin for each haplotype is shown, the closed and open parts of each bar indicating different degrees of confidence. The percentages refer to the proportions of the modern European population with each haplotype. All the haplotypes except J and T1 had entered Europe before the origin of agriculture 9000–10 000 years ago. Redrawn from Richards et al. (2000)

Figure 16.21. The route by which humans first entered the New World.

Research Briefing 16.1. Neandertal DNA

The End