Molecular evolution

1. Molecular evolution Part I: The evolution of macromolecules. Part II: The reconstruction of the evolutionary history of genes and organisms. Biol336-12

2. www.ncbi.nlm.nih.gov Molecular Evolution AGAMOUS; transcription factor [ Arabidopsis thaliana ] What information can DNA sequences give us? Evaluating the role of drift/demography vs. selection on trait divergence. Identify function. Looking at genes whose evolutionary history was shared. Biol336-12

3. Molecular Evolution 3D Protein Structure of Human proinsulin 1952: Frederick Sanger and coworkers determine the complete amino acid sequence of insulin. MALWMRLLPLLALLALWGPDPAAAFVNQHLCG Biol336-12

4. Molecular Evolution How and why have molecular sequences evolved to be the way they are? Biol336-12

5. Molecular Evolution Learning Objectives: • Variability within a population • Subsitution rates • Neutral Theory • Detecting selection at the DNA level. Biol336-12

6. Molecular Evolution REVIEW: NUCLEOTIDE SUBSITUTIONS Lys Ala Leu Val Leu Leu AgAt145 AAG GCA CTG GTC CTG TTG AgAt134 AAA GCA CTG GTC CTC TTG SepAt145 AGG GCA CTG GTC CTG GTG SepAt134 AGG GCA CTG GTC CTG GTG CalAt145 AAG - - - CTG TTC CTG TTG CalAt134 AAG - - - CTG TTC CTG TTG Biol336-12 SPECIES GENE

7. Molecular Evolution What happens after a mutation arises in the DNA sequence at a locus? Polymorphism: mutant allele is one of -several present in population. Substitution: the mutant allele fixes in the population. (New mutations at other nucleotides may occur later.) Biol336-12

8. Molecular Evolution 0 aaat aaat aaat aaat aaat aaat aaat 10 aaat aaat aaat aaat acat aaat aaat 20 aaat aaat acat aaat acat acat acat 30 acat acat acat acat acat acat acat 40 acat acat actt acat acat acat acat Generation 10-29 new mutation polymorphism Generation 30 mutation fixed substitution Generation 40 new mutation polymorphism L3 L2 L4 L5 L6 L7 L1 Time (generations) Biol336-12

9. Molecular Evolution Imagine that five sequences are obtained from each of two species, and that the sequences are related to each other as shown here. Biol336-12

10. Molecular Evolution Any mutation that happens on a red branch will appear as a polymorphism within species 1. Any mutation that happens on a blue branch will appear as a polymorphism within species 2. Any mutation that happens on the green branch will appear as a fixed difference between the species within species between species Biol336-12

11. Molecular Evolution What happens after a mutation arises in the DNA sequence at a locus? Polymorphism: mutant allele is one of -several present in population. Substitution: the mutant allele fixes in the population. (New mutations at other nucleotides may occur later.) Biol336-12

12. Molecular Evolution Substitution rate: the rate at which mutant alleles rise to fix within a lineage By comparing DNA sequences from different organisms, we can estimate the rate at which mutations appear and fix, causing basepair substitutions. Biol336-12

13. Molecular Evolution How many selectively neutral mutants reach fixation per unit time? Neutral mutations occur at a rate, μ per locus per generation. In a diploid population at a particular locus, there are 2N alleles. The number of mutants arising every generation at a given locus in a diploid population of size N is The probability of fixation of selectively neutral allele? Thus, the substitution rate for neutral alleles is 2N*μ 1/2N (1/2N)(2N*μ) = μ Biol336-12

14. (2Nμ)(2s) = 4Nμs Fixation probability for a beneficial allele Molecular Evolution What is the substitution rate for neutral alleles? μ What is the substitution rate for beneficial alleles (s>0)? What is the substitution rate for deleterious alleles? Close to zero. Biol336-12

15. Molecular Evolution Consider a numerical example: A new mutant arises in a population of 1000 individuals. If it is neutral the probability it will fix is If it confers a selective advantage of s=0.01, then the probability it will fix is, If it has a selective disadvantage of s=-0.001? 1/2N=1/(2*1000) 2*s=0.02 (2%) 0.004% Biol336-12

16. Molecular Evolution If the population size is very large then the probability of fixation for an advantageous mutation converges to 2*s Given s=0.01, N=1000, P(fixation)= 0.02 or 2%, Given s=0.01, N=100, P(fixation)=0.02037 Biol336-12

17. Molecular Evolution What about slightly deleterious mutations? s= -0.001, N=1000 P(fixation)=0.000406 s=-0.001, N=100, P(fixation)= 0.0049 s=-0.001, N=10, P(fixation) = 0.0499 Biol336-12

18. Molecular Evolution Are most substitutions (fixed changes) due to drift or natural selection? vs. Agree that: Most mutations are deleterious and are removed.Some mutations are favourable and are fixed. At Dispute: Are most replacement mutations that fix beneficial or neutral? Is observed polymorphism due to selection or drift? Biol336-12

19. Molecular Evolution Silent (or synonymous) mutations, where the amino acid remains unchanged, are more likely to be neutral. Replacement (or non-synonymous) mutations causing an amino acid change are more likely to experience selection. • Form and strength depends on gene and its function Biol336-12

20. Molecular Evolution Biol336-12

21. Molecular Evolution Histones seem to have an unusually low replacement substitution rate. This suggests that mutations causing basepair changes in histones are deleterious WHY? Biol336-12

22. Molecular Evolution Histones are DNA binding proteins around which DNA is coiled to form chromatin. Many positions within the protein interact with the DNA or other histones. Biol336-12

23. Molecular Evolution Most amino acid changes in histone proteins may have negative or even lethal consequences. Histone proteins have strong functional constraints. Biol336-12

24. Molecular Evolution Biol336-12

25. Molecular Evolution Active sites (antigen binding sites of immunoglobins often have higher substitution rates than silent sites Biol336-12

26. Molecular Evolution It could be that selection favours mutations in these regions, thereby increasing the diversity among antibodies produced by the body and improving the immune response Biol336-12

27. Molecular Evolution How and why have molecular sequences evolved to be the way they are? Biol336-12

28. Molecular Evolution To infer that selection has acted within a genome, one must reject the null hypothesis that no selection has acted. Null hypothesis: describes pattern of sequence evolution under the forces of mutation and drift. Remember from neutral theory: The rate at which one nucleotide is replaced by another nucleotide throughout a population (substitution) equals the rate of mutation (μ) at that site. Biol336-12

29. Molecular Evolution How do we detect selection at DNA sequences? Comparing intra-species polymorphism to inter-species differences (McDonald-Kreitman test). Linked/neighbouring neutral markers. Examine genes for Dn/Ds ratios. Biol336-12

30. Molecular Evolution: The McDonald Kreitman Test Kreitman and Hudson (1991) sequenced a 4750 basepair region near the alcohol dehydrogenase (ADH) gene from 11 individuals of D. melanogaster and found higher than expected levels of polymorphism Biol336-12

31. Molecular Evolution: The McDonald Kreitman Test There is only one amino acid polymorphism (AdhF/AdhS) within this region which occurs at site 1490. Biol336-12

32. Molecular Evolution: The McDonald Kreitman Test Selection may be maintaining this polymorphism at or near this site. Biol336-12

33. Molecular Evolution:The McDonald Kreitman Test ADH is an enzyme that breaks down ethanol. Flies carrying the ADHF allele survive better when their food is spiked with ethanol than do flies carrying the ADHS allele (Cavener and Clegg 1981) Nonetheless, the factor that maintains ADHF/ADHS polymorphism remains unknown. Alchohol dehydrogenase Biol336-12

34. Molecular Evolution:The McDonald-Kreitman Test How and why have molecular sequences evolved to be the way they are? How do we explain the patterns of variation observed in ADH DNA sequences? Biol336-12

35. Molecular Evolution: McDonald Kreitman Test Imagine that five sequences are obtained from each of two species, and that the sequences are related to each other as shown here. Biol336-12

36. Molecular Evolution: McDonald Kreitman Test Any mutation that happens on a red branch will appear as a polymorphism within species 1. Any mutation that happens on a blue branch will appear as a polymorphism within species 2. Any mutation that happens on the green branch will appear as a fixed difference between the species within species between species Biol336-12

37. Molecular Evolution: McDonald Kreitman Test Some abbreviations: Within species Ps=numbers of synonymous polymorphisms Pn=numbers of non-synonymous polymorphisms Between species Ds=numbers of synonymous substitutions Dn=numbers of non-synonymous substitutions Biol336-12

38. Molecular Evolution: McDonald Kreitman Test If mutations occur randomly over time and if the chance that a mutation does or does not cause an amino acid change remains constant, then the ratio of replacement to silent changes should be the same along any of these branches Between species Biol336-12

39. Molecular Evolution: McDonald Kreitman Test If mutations are neutral any of these mutations has an equal chance of persisting. So the ratio of replacement to silent polymorphisms within a species (Pn/Ps) should be the same as the ratio of replacement to silent differences fixed between species (Dn/Ds) Pn/Ps Dn/Ds Biol336-12

40. Molecular Evolution The McDonald-Kreitman Test: Ho: If all changes are neutral, the ratio of replacement to silent changes at polymorphic sites (within species) should equal the ratio among fixed differences (between species). H1: If replacement mutations are advantageous, they fix rapidly, causing ahigherreplacement to silent ratio between speciesand alowerreplacement to silent ratiowithin species. Biol336-12

41. Molecular Evolution The McDonald-Kreitman Test: H2: If replacement mutations are deleterious, they rarely fix. Thus there will be a lower ratio of replacement to silent changes between species and a higher replacement to silent ratio within species. H3: If replacement mutations are subject to heterozygote advantage or frequency dependent selection, they rarely fix, causing a lower replacement to silent ratio between species and a higher replacement to silent ratio within species. Biol336-12

42. Molecular Evolution Null: all changes are neutral : drift H1: changes are advantageous, positive selection H2: changes are deleterious, purifying selection H3: replacement changes never fix because of heterozygote advantage. Biol336-12

43. Molecular Evolution: McDonald Kreitman Test Btwn species: Ratio of replacement to silent = 7/17 =0.41 Wn species: Ratio of replacement to silent = 2/42 =0.05 FIXED>POLYMORPHISM Biol336-12

44. Molecular Evolution: McDonald Kreitman Test Using a X2 test, the null hypothesis that selection is absent is statistically rejected for ADH. The excess of replacement differences between species suggests that mutations have been postively favoured. Biol336-12

45. Molecular Evolution: McDonald Kreitman Test Assumes: All synonymous mutations are neutral (codon bias). All non-synonymous mutations are either strongly deleterious, neutral or strongly advantageous. Levels of polymorphism are governed by the neutral mutation rate. Within a species, advantageous mutations contribute little to polymorphism but can contribute to divergence between species. A problem with this test is that: A failure to reject the null hypothesis could be because both purifying and directional selection have taken place. Not all synonymous changes are in fact neutral. In some organisms, some codons are preferentially used. Biol336-12

46. Molecular Evolution How else might you detect selection in the genome, in particular the presence of selective sweeps? Biol336-12

47. Molecular Evolution: Neighbouring marker sites If a beneficial mutation appears and sweeps through a population, what will happen to the level of polymorphism present at neighbouring DNA sites? Biol336-12

48. Molecular Evolution: Neighbouring marker sites If a beneficial mutation appears and sweeps through a population, what will happen to the level of polymorphism present at neighbouring DNA sites? Genetic hitchhiking will decrease variation. Biol336-12

49. Molecular Evolution: Neighbouring marker sites In the case of Plasmodium falciparum, diversity at neighbouring marker loci decreased. Biol336-12

50. Molecular Evolution: Neighbouring marker sites Biol336-12 Wootton et al.(2002) Nature