Significance of variation

McDonald and Kreitman 1991

Mutations are the raw material of evolution

• Source of new alleles• Source of new genes• Produce heritable variation that is transmitted across

generations

1. Small-scale mutations2. Macromutations

• Polyploidy• Doubling of whole genomic DNA

Genome and gene duplications create evolutionary novelty

Sanetra et al.Frontiers in Zoology 2005 2:15

• Vertebrate evolution punctuated by three widespread gene or genome duplications

• Associated with:• Increases in

morphological complexity

• Adaptive radiations in body design

• Is genome duplication the explanation?

Secretory calcium binding phosphoprotein (SCPP) gene family

Tunicate Ray-finned fish Lobe-finned fish

In mammals, formation of

tooth, bone and milk depends

upon SCPP

Genome duplications = evolutionary novelty • If this is true, what pattern would you expect to see

on a phylogenetic tree after genome duplication event in terms of species diversity?

• Burst of diversification• Not supported

Extant lineages

Extinct lineages

Number of families per

clade

Donoghue and Purnell 2005

Clades

Evolution of jawed fish

• If this is true, what pattern would you expect to see on a phylogenetic tree after genome duplication event?

• Burst of diversification• Not supported

Extant lineages

Extinct lineages

Fossil evidence

Donoghue and Purnell 2005

Evolution of ray-finned fishes

Genome duplications = evolutionary novelty

Crow and Wagner 2005. Mol. Bio. Evol. 23:887-892

Genome duplications provide robustness

Extant lineages

Extinct lineages

• Focus on the high rate of extinction before duplication

• Provides robustness against extinction

Genome duplication associated with diversification of the angiosperms

De Bodt et al. 2005

• Appear suddenly in the fossil record

Darwin referred to the rapid rise and early diversification of the angiosperms

as an “abominable mystery”

• Tried to identify a single causal factor

• Described his efforts “wretchedly poor”

Letter to J.D. Hooker dated July 22 1897

Genome duplication associated with diversification of the angiosperms

De Bodt et al. 2005

• Appear suddenly in the fossil record

• Polyploidy created developmental and regulatory gene families found in angiosperms

Are genome-wide and single-gene duplications equally valuable from an evolutionary perspective?• Genes involved in signal transduction

transcriptional regulation and are preferentially retained following polyploidy

Maere et al. 2005. PNAS.102, 5454–5459Blanc & Wolfe. 2004.Plant Cell 16, 1679–1691Seoighe & Gehring. 2004. Trends Genet. 20, 461–464

Are genome-wide and single-gene duplications equally valuable from an evolutionary perspective?• Genes involved in signal transduction

transcriptional regulation and are preferentially retained following polyploidy

• Developmental genes also retained at higher frequency

• Fewer of these genes survive single gene duplications

• Transcription factors and genes involved in signal transduction show high dosage effects

• Protein components must be present in stoichiometric qualities

Meiotic Nonreduction

Fertilization

Polyploidy in plants is an ancient and ongoing process

X

Tetraploid

Diploid

• 70-80% of plants have polyploidy origins

• Speciation via polyploidy has been observed in modern times

2x

4x

6x

8x

10x

Chrysanthemum

Many species posses a ploidy series

2x

4x

6x

8x

10x

Do higher ploidy levels possess greater potential for evolutionary change?

More gene products

Greater genetic

diversity

Opportunity for

duplicated genes to

diverge in function

More gene

interactions

Artificial selection on

timing of flowering

Will polyploids evolve faster?

G0

G1

G2

G3

G4

Days to flower

control

Days to flower

control

Tetraploid

Chromosome count = 36

Diploid

Chromosome count = 18

Figure 1. Countable chromosome images from the ploidy levels included in this artificial selection experiment

Tetraploid

Chromosome count = 36

Diploid

Chromosome count = 18

Figure 1. Countable chromosome images from the ploidy levels included in this artificial selection experiment

Do higher ploidy levels possess greater potential for evolutionary change?

Ploidy Level

Tetraploid Diploid

% o

f Sel

ectio

n lin

es th

at a

re

sign

fican

tly d

iffer

ent t

han

the

cont

rols

0

20

40

60

80

100

Chi-square = 9.9; P = 0.002

Diploid

Chromosome count = 18

experiment

Tetraploid

Chromosome count = 36

Chromosome count = 18

Do ploidy levels differ in their geographic distributions?

2x4x>4x

2x4x>4x

Cytotypes

Diploid Triploid Tetraploid Higher

Gen

ome

size

(pg

DN

A)

3.0

3.5

4.0

4.5

5.0

5.5

6.0

6.5

F3,768 = 2591 P < 0.0001

Cytotypes

Diploid Triploid Tetraploid Higher

Gen

ome

size

(pg

DN

A)

3.0

3.5

4.0

4.5

5.0

5.5

6.0

6.5

F3,768 = 2591 P < 0.0001

Figure 1. Genome size for Solidago altissima plants with different cytotypes which are putatively diploid, triploid, tetraploid, and a mix of pentaploids and hexaploids. Calclations based on an internal standard with a known genome size (chicken red blood cells)

Solidago altissimaLate goldenrod

Futuyma 1998

Do alleles differ in their geographic distribution?

• Cline in alcohol dehydrogenase locus of Drosophila melanogaster

• Repeated on three continents

Are different alleles being favored over time?

Anderson et al. 2005. The latitudinal cline in the In(3R)Payne inversion polymorphism has shifted in the last 20 years in Australian Drosophila

melanogaster populations. Molecular Ecology 14: 851–858

Selectionist mission

Link biochemical differences to fitness in nature

Few well-known examples where natural selection is clearly involved in the maintenance of enzyme polymorphism

http://anthro.palomar.edu/synthetic/images/map_of_sickle_cell_frequencies.gif

Significance of variation

• Chimps and humans differ in 1% of our genes

• ~3,400,000 nucleotides• ~60,000 amino acid differences

• What proportion of these differences have been fixed because they were beneficial and allowed us to adapt to our environments?

• How many of these differences were simply fixed by random genetic drift?

To what extent does natural selection operate at the molecular level?

Some history…

Significance of variation

• Study of variation at the molecular level began with proteins (allozymes).

Hubby and Lewontin. 1966. A molecular approach to the study of genic heterozygosity in natural populations. The number of alleles at differ loci in Drosophila pseudoobscura. Genetis 54:577-94.Harris 1966. Enzyme polymorphism in man. Proc. Roy. Soc. B. 164:298-310.

• Discovered astonishing level of polymorphism• Challenged our fundamental understanding of how

adaptive evolution occurs• Most variation must be neutral – Kimura’s neutral theory

of evolution (1970)

“The maintenance of abundant polymorphism and heterozygosity in populations demands, however, an explanation… The easiest way to cut the Gordian knot is, of course, to assume that a great majority of the polymorphisms observed involve gene variants that are selectively neutral, that is, have no appreciable effects on the fitness of their carriers” Dobzhansky 1970

The beginning of neutralist-selectionist debate

• Kimura (1968)• Most mutations are deleterious and are rapidly

eliminated• A very small number of mutations are favorable and

are rapidly fixed• Most of the variation that we observe within species

is selectively neutral and is governed by the interplay of mutation and drift

• Most differences between species are simply due to the random fixation of mutations

The central tenants of the neutral theory

• Estimates of overall heterozygosityNei 1983. Genetic polymorphism and role of mutation in evolution. In The evolution of genes and proteins p. 165-190

• Distribution of single locus heterozygosityNei et al. 1976 Testing the neutral mutation hypothesis by distribution of single locus heterozygosity. Nature 262:491-493

• Variance in heterozygosityGojobori 1982 Means and variances of heterozygosity and protein function. In Molecular Evoluton, Protein Polymorphism and the Neutral Theory pp 137-150

• Number of alleles per locusChakaraborty et al. 1980. Statistical studies on protein polymorphism in natural populations. III. Distribution of allele frequencies and the number of alleles per locus. Genetics 94:1039-1063

• The correlation of single-locus heterozygosity between related species

Braverman et al. 1995. The hitchhiking effect on the site frequency spectrum of DNA polymorphism Genetics 140:783-795

Neutral theory seemed to work!

Duplicate genes• Faster rates of nonsynonymous replacement than of

synonymous replacement• High selective value of protein divergence

• Hemoglobins• Visual pigments• Adrenergic receptors in humans• Antigen recognition sites in humans and mouse• Immunoglobulins• Growth hormone genes in humans and bovines

A few clear cases showed accelerated protein evolution

Duplicate genes• Faster rates of nonsynonymous replacement than of

synonymous replacement• High selective value of protein divergence

• Hemoglobins• Visual pigments• Adrenergic receptors in humans• Antigen recognition sites in humans and mouse• Immunoglobulins• Growth hormone genes in humans and bovines

A few clear cases showed accelerated protein evolution Can you make this

into a general test?

What can we learn by comparing the rate of synonymous and nonsynonymous

replacements?

• New light recently shed on debate because of:• Increase in DNA sequence data• New methods of analysis

• McDonald-Kreitman test (MK)• Neutral theory predicts the amount of

variation there should be within and between species

• We can use sequence data to calculate the amount of variation within a species (polymorphism) to the amount of variation between species (divergence):1. Synonymous (no change in amino

acid sequence or regulatory sequences)

2. Nonsynonymous (change in amino acid sequence or regulatory sequence)

Does selection act at the molecular level?

PolymorphismPs and Pn

DivergenceDs and Dn

If neutral - expect more syn or nonsyn?

If neutral - expect more syn or nonsyn?

• McDonald-Kreitman test (MK)• Nonsynonymous mutations that are

adaptive () contribute to divergence (Dn) but not so much to polymorphism (Pn)

• Rapidly fixed by selection so they are not segregating within species

• Synonymous mutations will accumulate at the neutral rate (Ds and Ps)

• If most molecular evolution is neutral then:

• If under selection?

The verbal argument…

PolymorphismPs and Pn

DivergenceDs and Dn

Dn Pn

Ds Ps

=>

Ps = 4NeLsk

Pn = 4NeLnfk

Ds = 2Lst

Dn = 2Lntf 1 -

Ne = effective population size = nucleotide mutation rateLs = number of synonymous sitesk = constant

number sequences sampledsampling strategydemography

Ln = number of nonsynonymous sitesf = proportion of amino acid mutations that are neutral t = time since divergence of two species = proportion of nonsynonymous mutations that are adaptive

Time it takes a new mutation to become fixed if that is its destiny from Kimura’s equations

Dn = Pn

Ds Ps

The mathmatical argument…

= 1 – DsPn

DnPs

Dn = Pn

Ds Ps

Divergence Polymorphism

Non-synonymous Synonymous

3648 4397365 1741

0.49 0.25

= 1 – DsPn

DnPs

= 1 – (7365)(439) (3648)(1741) = 0.49

MK test for adaptive evolution

115 genes in two species of Drosophila

• From this and other studies, adaptive value evolution in Drosophila protein-coding sequences converging at ~50%

• Can extrapolate to the whole genome• If = 0.45, then Drosophila would have one substitution

every 45 years or 450 generations• 22,000 substitutions per million years

Inferring the strength of adaptive evolution

Eyre-Walker 2006

Survey of rate of adaptive evolution at molecular level

• No.• The proportion of nonsynonymous mutations that are

adaptive is particularly low in comparisons of humans and other organisms

• Ranges 0-35%• Bias in 35% estimate of Fay et. al 2001

• Used genes associated disease and other critical function

• Also little evidence of adaptive evolution in Arabidopsis.• High levels of adaptive evolution in viruses and bacteria

Is the pattern uniform across species?

• D. melanogaster and D. simulans differ by 110,000 adaptive amino acid differences

• Species are almost identical morphologically• Physiology• Ecology• Arms race between hosts and parasites

• “It might be that we just have no idea how complex the environment really is and how it is constantly changing in ways that challenge organisms to adapt.” (Eyre-Walker 2006)

What do all these substitutions do?

Evolution is the interplay between two tangled banks

Natural turbulence Genetic turbulence