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The neutral theory

of molecular evolution

the neutral theory

detecting natural selection

exercises

Objectives

1 - learn about the neutral theory

2 - be able to detect natural selection at the molecular level

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Two theses in Darwins Origin of Species

1 - organisms descend with modification from common ancestors

phylogenetics - pattern

2 - mechanism for this modification is natural selection

molecular basis of adaptation - process

Molecular evolution and Darwin

The field of molecular evolution has been dominated by phylogenetics and molecular

systematics. These endeavors have been extremely successful in supporting and elucidating

the dynamics of point #1 above.

Molecular evolutionists have been relatively less successful (Sharp 1997) at uncoveringevidence detailing the mechanism of descent with modification - the molecular basis of

adaptation. Recent years have seen tremendous strides in this area (Hughes 1999) but it

remains to a great extent uncharted territory.

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Population genetics

interested in genetic variation - understand generation and maintenance

initially (until 1960s) only able to study indirectly - phenotype

paucity of data led to controversy on the extent of genetic variation

Classic school - very little genetic

variation, cost associated withnatural selection Muller

Balance school - lots of genetic variation

maintained by natural selectionDobzhansky

debate settled with advent of molecular approaches (direct) to the

study of genetic variation - electrophoresis, sequencing

tremendous amount of genetic variation exists

thought was that this variation was maintained by natural selectionaccumulation of adaptively advantageous variants

heterozygote advantage or heterosis

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A new explanation neutralist for the

high levels of molecular variationMotoo Kimura (1968) Evolutionary rate at the molecular level. Nature 217: 624

Electrophoresis studies (Lewontin & Hubby) and sequence comparisons (Pauling & Zuckerkandl)

reveal high levels of molecular variation

Kimura reasoned that variation was too high and accumulated to rapidly to be explained by selection

This is the so-called cost of selection argument (see appendix slide # 9)

Conclude that these observed differences are selectively neutral that is they do not confer any

selective advantage or disadvantage to the organisms that bear them (because they do not alter the

function of the protein that they encode)

J.L. King & T.H. Jukes (1969) Non-Darwinian evolution.Science 164: 788

Note that many genetic changes have no effect on organismic fitness they are neutral

Natural selection can not alter changes that it can not perceive Marshall biochemical evidence in support of these assertions

e.g. synonymous substitutions, functionally equivalent cytochrome c variants, rapid evolution of

fibrinopeptides (removed from functional fibrinogen)

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The neutral theory

change too rapid to be explained by natural selection therefore most of the changes observed are selectively neutral -

no effect on fitness

Kimura reasoned that the majority of both polymorphism (allelic frequencies within

populations) and substitution (fixed differences between populations) result from

fixation of selectively neutral variants by random genetic drift

- the main role of natural selection is elimination of deleterious variants (maintenance

of the status quo) - molecular evolution is conservative

- adaptively favorable mutations fixed by natural selection are a small minority of

all nucleotide substitutions

huge debate ensued between selectionists (believe that extensive variationis a product of natural selection) and neutralists (believe that variation is

a product of random fixation of neutral variants)

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Predictions of the neutral theory

neutral theory makes explicit quantitative predictions about levels ofgenetic variation - null hypothesis of molecular evolution

functionally important parts of a molecule will change more slowly than

functionally unimportant parts

Those mutant substitutions that disrupt less the existing structure and

function of a molecule (conservative substitutions) occur more frequently

in evolution than more disruptive ones. Kimura and Ohta 1974

most important for our purposes:

Absolutely essential concept in modern molecular biology: basis of

programs to align sequences and make functional predictions

Challenge to the Darwinian view: if selection is driving force in evolution, rateof evolution should be most rapid where selection operates most - in the functionally

important parts of molecules (opposite to the neutral view)

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synonymous subs - do not change encoded amino acidnonsynoymous subs - do change encoded amino acid

in virtually every gene ever studied synonymous sites change

at a higher rate than nonsynonymous sites

Maximum evolutionary rate &

selection versus neutrality do relative rates of change better fit selectionist or neutralist prediction? overwhelming support for neutralist prediction:

1 synonymous versus nonsynonymous subs rate (Kimura 1977, Jukes 1978)

2 accelerated rate of psuedogene evolution (Li et al 1981)

GAT AAC ATC CAA GGAATAACT GCAATC

GAC AAC ATC CAA GGT ATC ACG GCT ATC

Asp Asn Ile Gln Gly Ile Thr Ala Ile

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Detection of natural selection using synonymous

& non-synonymous substitution ratesTypes of natural selection:

1 purifying (negative) selection removal of deleterious variants

2 diversifying (positive) selection fixation of adaptive variants

Types of substitution rates: (for protein coding genes i.e. codons)

1 synonymous substitution rate (Ks or ds) rate of substitution for DNA changes that do notchange the encoded amino acids

2 non-synonymous substitution rate (Ka or dn) rate of substitution for DNA changes that do

change the encoded amino acids

The relative levels for these rates indicate the mode of selection for a gene

Neutral evolution (no selection): Ks

KaPurifying selection: Ks >> Ka

Diversifying selection: Ks

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Exercises

Compare synonymous and nonsynonymous substitution rates for:

1 the Drosophila alcohol dehydrogenase (Adh) gene dros-adh.meg

2 the human & mouse gene pair mammal.meg

Determine the mode of selection acting on each based on these rates

1 - load alignment into DnaSP2 - assign coding region

3 - calculate Ks and Ka and compare (which is higher)

4 - load alignment into Mega

5 - calculate ds and dn and compare (which is higher)

6 - do statistical test for difference between ds and dn

(see appendix II slide # 11)

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Kimuras derivation of the neutral theory noticed variation too high and change too rapid to be explained by natural selection

- based on amino acid sequence data (hemoglobin & cytochrome c) Kimura calculated an

average of 1 aa per 28 my in a 100 aa protein

- this is too high for natural selection based on Haldanes concept of the cost of selection

- if only individuals with high fitnesses for a number of diff traits survive, only a

very small fraction of the population will remain

e.g. moth melanism - 50% mortality due to bird predation

- if simultaneous effects at 10 loci 1 / 210 (1 out of 1,024) survivors - population likely to go

extinct before all 10 alleles fixed

- Haldane calculated that a 1 new allele per 300 generations can be substituted

- Kimura noted that in fact substitutions at the molecular level occurring much more rapidly

1 -1 sub per 28my per 100 aa2 - mammalian genome size 4 x 109 bp

3 - 100 aa = 300 bp and 20% nucleotide subs synonymous thus 1 aa sub 1.2 bp sub4 - time it would take for nucleotide substitution to occur in the genome is:

(28 x 106) / (4 x 109/300) / 1.2 = 1.8 years

- this is a much higher rate of substitution than 1 every 300 generations

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Statistical comparison of synonymous

& non-synonymous substitution rates

Depending on the values observed one may wish to test the following hypotheses:

ds > dn ds = dn ds < dn

To do this use the normal deviate or Z test (see lecture 8 slide #6)

Z = difference between ds & dn divided by the standard error of the difference

difference D = abs (ds dn)

Standard error of the difference sD = (se(ds)2 + se(dn)2)

Z = D / sD formula in MSExcel =abs(ds-dn)/sqrt(se(ds)^2+se(dn)^2)

Then use t-table with infinite degrees of freedom to evaluate P value

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