Transcript Molecular basis of evolution.

Molecular basis of evolution.
Goal – to reconstruct the evolutionary history of all
organisms in the form of phylogenetic trees.
Classical approach: phylogenetic trees were
constructed based on the comparative morphology
and physiology.
Molecular phylogenetics: phylogenetic trees are
constructed by comparing DNA/protein sequences
between organisms.
Evolution of mankind.
Analysis of mitochondrial DNA proposes that Homo sapiens
evolved from one group of Homo erectus in Africa (African
Eve) 100,000 – 200,000 years ago.
American indians I,
25-35,000
Europeans
40-50,000
American indians II,
7-9,000
Africans
100,000
Asians
55-75,000
Adam appeared 250,000 years ago, much earlier!
Mechanisms of evolution.
- By mutations of genes. Mutations spread
through the population via genetic drift and/or
natural selection.
- By gene duplication and recombination.
Mutational changes of DNA sequences.
1. Substitution.
Thr Tyr Leu Leu
ACC TAT TTG CTG
3. Insertion.
Thr Tyr Leu Leu
ACC TAT TTG CTG
ACC TCT TTG CTG
Thr Tyr Leu Leu
ACC TAC TTT GCT G—
Thr Tyr Phe Ala
2. Deletion.
Thr Tyr Leu Leu
ACC TAT TTG CTG
4. Inversion.
Thr Tyr Leu Leu
ACC TAT TTG CTG
ACC TAT TGC TGThr Tyr Cys
ACC TTT ATG CTG
Thr Phe Met Leu
Synonymous and nonsynonymous
nucleotide substitutions.
Synonymous substitutions in codons do not change the
encoding amino acid, non-synonymous substitutions do.
ds/dn < 1 indicates positive natural selection.
ds, dn - # of synonymous substitutions per
non-synonymous site
Gene duplication and recombination.
New genes/proteins occur through gene duplication and
recombination.
Ancestral globin
duplication
Gene 1
+
Gene 2
globin
globin
hemoglobin
myoglobin
New gene
Duplication
Recombination
Measures of evolutionary distance
between amino acid sequences.
1. P-distance. Evolutionary distance is usually measures
by the number of amino acid substitutions.
p  nd / n
nd – number of amino acid differences between two
sequences; n – number of aligned amino acids.
Poisson correction for evolutionary
distance.
2. PC-distance. Takes into account multiple substitutions
and therefore is proportional to divergence time.
PC-distance can be expressed through the p-distance:
d   ln( 1  p )
Another method to estimate evolutionary
distances: amino acid substitution matrices.
3. Distance from amino acid substitution matrices.
Substitutions occur more often between amino acids of
similar properties.
- Dayhoff (1978) derived first matrices from multiple
alignments of close homologs.
- The number of aa substitutions is measured in terms of
accepted point mutations (PAM) – one aa substitution
per 100 sites.
- Dayhoff-distance can be approximated by gammadistance with a=2.25.
Fixation of mutations.
Not all mutations are spread through population. Fixation –
when a mutation is incorporated into a genome of species.
Majority of mutations are neutral (Kimura), do not effect the
fitness of organism.
Fixation rate depends on the size of population (N), fitness (s)
and mutation rate (μ):
r  Ns
Phylogenetic analysis.
- Phylogenetic trees are derived from multiple sequence
alignments. Each column describes the evolution of one
site.
- Each position/site in proteins/nucleic acids changes in
evolution independently from each other.
- Insertions/deletions are usually ignored and trees are
constructed only from the aligned regions.
Evolutionary tree constructed from rRNA
analysis.
The concept of evolutionary trees.
- Trees consist of nodes and branches, topology - branching
pattern.
- The length of each branch represents the number of
substitutions occurring between two nodes. If rate of
evolution is constant, branches will have the same length
(molecular clock hypothesis).
- The distance along the tree is calculated by summing up all
intervening branch lengths.
- Trees can be binary or bifurcating.
- Trees can be rooted and unrooted. The root is placed by
including a taxon which is known to branch off earlier than
others.
Accuracies of phylogenetic trees.
Two types of errors:
- Topological error
- Branch length error
Bootstrap test:
Resampling of alignment columns with
replacement; recalculating the tree; counting how
many times this topology occurred – “bootstrap
confidence value”. If it is close to 100% – reliable
topology/interior branch.
Estimation of species divergence time.
Assumption: rate constancy, molecular clock.
Find T1, if T2 is known.
T1
T2
A
B
D AC
D AB

;
2T1
2T2
T1 
D ACT2
D AB
C
Estimation of evolutionary rates in
hemoglobin alpha-chains.
P-distance
PC-distance
Gamma-distance
Human/cow
0.121
0.129
0.134
Human/kangaroo
0.186
0.205
0.216
Human/carp
0.486
0.665
0.789
Estimate the evolutionary rate of divergence between human
and cow (time of divergence between these groups is ~90
millions years).
Methods for phylogenetic trees
construction.
Set of
related
sequences
Multiple
sequence
alignments
Strong
sequence
similarity?
Yes
Maximum
parsimony
methods
No
Recognizable
sequence
similarity?
Yes
Distance
methods
No
Maximum
likelihood
methods
Analyze
reliability of
prediction
1. Distance methods. Calculating branch
lengths from distances.
A
B
C
a  b  20;
A
-----
20
30
B
-----
-----
44
a  c  40;
b  c  44;
C
-----
-----
-----
a  8; b  12; c  32.
a
c
b
Neighbor-joining method.
NJ is based on minimum evolution principle (sum of branch length
should be minimized).
Given the distance matrix between all sequences, NJ joins sequences
in a tree so that to give the estimate of branch lengths.
1. Starts with the star tree, calculates the sum of branch lengths.
C
d AB  a  b;
B
b
d AC  a  c;
c
a
d
D
d AD  a  d ;
d AE  a  e;
S  abcd e 
e
A
(d AB  d AC  d AD  d AE  d BC  d BD  d BE  d CD  d CE  d DE ) /( N  1)
E
Neighbor-joining method.
2. Combine two sequences in a pair, modify the tree.
3. Treat cluster CDE as one sequence “X”, calculate average distances
between “A” and “X”, “B” and “X”, calculate “a” and “b”.
C
B
d AX  (d AC  d AD  d AE ) / 3;
c
b
d BX  (d BC  d BD  d BE ) / 3;
d
a
A
D
e
a  b  d AB ; a  x  d AX ; b  x  d BX .
E
4. Treat AB as a single sequence, calculate c, d and e.
5. Calculate the sum of branch lengths, S.
5. Repeat the cycle and calculate S for other pair, choose the lowest S.
Classwork I
Given a multiple sequence, construct distance matrix (p-distance) and
calculate the branch lengths.
APTHASTRLKHHDDHH
ALTKKSTRIRHIPD-H
DLTPSSTIIR-YPDLH
Classwork II: NJ tree using MEGA.
1. Go to CDD webpage and retrieve alignment of cd00157 in
FASTA format.
2. Import this alignment into MEGA and convert it to MEGA
format http://www.megasoftware.net/mega3/mega.html .
3. Construct NJ tree using different distance measures with
bootstrap.
4. Analyze obtained trees.
2.1 Maximum parsimony: definition of
informative sites.
Maximum parsimony tree – tree, that requires the smallest
number of evolutionary changes to explain the differences
between external nodes.
Site, which favors some trees over the others.
1
2
3
4
5
6
7
A
A
A
A
A
G
G
G
G
C
A
A
A
C
T
G
C
C
T
T
*
T
T
T
T
G
G
C
C
*
Site is informative (for nucleotide sequences) if there are at
least two different kinds of letters at the site, each of which
is represented in at least two of the sequences.
2. Maximum parsimony.
Site 3
1.G
3.A 1.G
G
A
2.C
A
A
4.A 3.A
Tree 1.
2.C
2.C 1.G
A
4.A
A
4.A
Tree 2.
3.A
Tree 3.
Site 3 is not informative, all trees are realized by the same number of
substitutions.
Advantage: deals with characters, don’t need to compute distance matrices.
Disadvantage:
- multiple substitutions are not considered
- branch lengths are difficult to calculate
- slow
2.3 Maximum parsimony method.
1.
Identify all informative sites in the alignment.
2.
Calculate the minimum number of substitutions at each
informative site.
3.
Sum number of changes over all informative sites for each
tree.
4.
Choose tree with the smallest number of changes.
Maximum likelihood methods.
• Similarity with maximum parsimony:
- for each column of the alignment all possible trees are
calculated
- trees with the least number of substitutions are more likely
• Advantage of maximum likelihood over maximum parsimony:
- takes into account different rates of substitution between
different amino acids and/or different sites
- applicable to more diverse sequences
Classwork: maximum marsimony.
1.
2.
3.
Search the NCBI Conserved Domain Database for pfam00127.
Construct maximum parsimony tree using MEGA3.
Analyze this tree and compare it with the phylogenetic tree from
the research paper.