From basic Concepts to Advanced applications

Molecular Evolution and Phylogeny

By Ofir Cohen

The Bioinformatics UnitG.S. Wise Faculty of Life Science

Tel Aviv University, Israel2011

http://ibis.tau.ac.il/twiki/bin/view/Bioinformatics/Phylogeny

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Darwin’s teachings– common descent and Tree-like evolution

Introduction – The tree concept

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Common Descent – Modern evidence

Introduction – The tree concept

"The unity of life is no less remarkable than its diversity" "The unity of life is no less remarkable than its diversity" THEODOSIUS DOBZHANSK

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What is a Phylogenetic Tree? Phylogenetic tree:

(hypothetical) historical pattern of evolutionary relationships among organisms

Introduction – The tree concept

Homo

Bos

Mus

Rattus0.011

0.025

0.012

0.011

Gallus

0.038

0.066

0.01

Root

Node

Leaf

Branch

(Greek: phylon = race and genetic = birth)

sps

Horizontal branch length –proportional to evolutionary distances (unit = substitution / site)

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Molecular evidence of HIV transmission in a

criminal case

Introduction - Anecdotes

Metzker, Michael L. et al. (2002) Proc. Natl. Acad. Sci. USA 99, 14292-14297

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Criminal investigation

August 1994 a nurse tests negative for HIV. breaks off a messy 10 year affair with a doctor. Three weeks later the doctor gives his ex-mistress a vitamin B-12 shot

In January 1995, the nurse tests positive for both HIV and hepatitis C.

The doctor’s office records from the day are missing (but eventually found). The doctor had withdrawn blood samples from a known HIV patient and a known hepatitis C patient

the same day as the vitamin B-12 shot. The nurse had never had contact with either patient

Introduction - Anecdotes

Circumstantial evidence that the doctor injected blood from a patient of his into this ex-girlfriend….

How can this be proved using a phylogenetic approach?

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HIV – short background

Extreme heterogeneity Within each patient there are many different viral

strains ("quasi-species")

Introduction - Anecdotes

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History of the virus:

gp120PATIENT

VICTIM

CONTROLS

©2002 National Academy of Sciences, U.S.A.

Introduction - Anecdotes

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History of the virus:

RT VICTIM

PATIENT

Introduction - Anecdotes

Source sequences that are paraphyletic (other sequences are nested within them)

with respect to the recipient sequences provide evidence for the direction of transmission.

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Phylogenetic analysis: Not only among organisms - Cancer

phylogenyA phylogeny of acute myeloid leukemia (AML) subtypes

Riester et al. 2010Liu et al. 2009

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Phylogenetic analysis: Not only in biology – Language evolution

Russell and Atkinson. 2003

Researchers learn the evolution of languages by treating them like genomes.

Instead of COGs (gene families), analyze COGNATES (words families)

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Comparative Genomics – "All life is one"

Compare homologues sequences

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Newick format with branch lengths

(A:0.3,((B1:0.1,B2:0.1):0.3,(C1:0.1,C2:0.1):0.5):0.3);

0.1

A

B1

C1

C2

B2http://tree.bio.ed.ac.uk/software/figtree/

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Alignment and phylogeny are mutually dependant

Inaccurate tree building

MSA

Sequence alignment

0.4

Phylogeny reconstruction

Unaligned sequences

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Multiple sequence alignment (MSA)Several advanced MSA programs are available.

Today we will use two: MAFFT – fastest and one of the most accurate PRANK – distinct from all other MSA programs because of its

correct treatment of insertions/deletions

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MAFFT Web server & download:

http://align.bmr.kyushu-u.ac.jp/mafft/online/server/ Efficiency-tuned variants

quick & dirty or slow but accurate

Nucleic Acids Research, 2002, Vol. 30, No. 14 3059-3066© 2002 Oxford University Press

MAFFT: a novel method for rapid multiple sequence alignment based on fast Fourier transform

Kazutaka Katoh, Kazuharu Misawa1, Kei-ichi Kuma and Takashi Miyata*

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Choosing a MAFFT strategy

qu

ick &

dirty slow

bu

t accurate

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Choosing a MAFFT strategy

qu

ick &

dirty slow

bu

t accurate

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Choosing a MAFFT strategy

qu

ick &

dirty slow

bu

t accurate

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Choosing a MAFFT strategy

L-INS-i

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--------------------------------XX-XXXXXXXXXXXXXXX-XXXXXXXXooooooooooo-------

------------------ooooooooooooooXXXXX----XXXXXXXX---XXXXXXXooooooooooo-------

--------ooooooooooooooooooooooooXXXXX-XXXXXXXXXX----XXXXXXXoooooooooooooooooo

--------------------------------XXXXXXXXXXXXXXXX----XXXXXXX------------------

G-INS-i

XXXXXXXXXXX-XXXXXXXXXXXXXXX

XX-XXXXXXXXXXXXXXX-XXXXXXXX

XXXXX----XXXXXXXX---XXXXXXX

XXXXX-XXXXXXXXXX----XXXXXXX

XXXXXXXXXXXXXXXX----XXXXXXX

E-INS-i

oooooooooXXX------XXXX---------------------------------XXXXXXXXXXX-XXXXXXXXXXXXXXXooooooooooooo

---------XXXXXXXXXXXXXooo------------------------------XXXXXXXXXXXXXXXXXX-XXXXXXXX-------------

-----ooooXXXXXX---XXXXooooooooooo----------------------XXXXX----XXXXXXXXXXXXXXXXXXooooooooooooo

---------XXXXX----XXXXoooooooooooooooooooooooooooooooooXXXXX-XXXXXXXXXXXX--XXXXXXX-------------

---------XXXXX----XXXX---------------------------------XXXXX---XXXXXXXXXX--XXXXXXXooooo--------

qu

ick &

dirty slow

bu

t accurate

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MAFFT outputSaving the output Choose a format: Clustal, Fasta, or

click "Reformat" to convert to a selection of other formats

Save page as a text file

A colored view of the alignment

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PRANK

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Classical alignment errors for HIV env

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PRANK Web server: http://www.ebi.ac.uk/goldman-srv/webPRANK/

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PRANK output

If you need a different format – copy the results to the READSEQ sequence converter: http://www-bimas.cit.nih.gov/molbio/readseq/

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Downloadable PRANK http://www.ebi.ac.uk/goldman-srv/prank/prank/

PRANK: A command-line program interface PRANKSTER: A program with graphical user interface

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1. Download and unzip the sequence files from my homepage (Google “Ofir Cohen" and look for the workshop materials under "Teaching"). Open "fahA.fas" in Notepad – these are 65 protein sequences in FASTA format.

2. Run PRANKSTER, open the "fahA.fas" file, and run "Alignment""Make alignment"

3. While you wait: Copy the sequences into the MAFFT web server and run the "automatic" "moderate" strategy – which strategy did MAFFT choose for you? Click "Reformat", choose "phylip|phylip4", and save as "fahA.mafft.phylip"

4. When PRANKSTER finishes click FileSave, and save the MSA in Phylip format by the name "fahA.prank.phylip"

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Phylogeny reconstructionDifferent approaches (algorithms / programs): Distance based methods (e.g. neighbor-joining, as in ClustalW)

Fast but inaccurate Maximum parsimony (e.g. MEGA) Maximum likelihood methods (e.g. phyML, RAxML)

Accurate but slower Bayesian methods (e.g. MrBayes)

Most accurate but very slow

ABCDE

Guide tree

A

DCB

E

MSA

Pairwise distance table

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PhyMLThe most widely used maximum likelihood (ML) program Web server & download: http://www.atgc-montpellier.fr/phyml/

Accepts input MSA in PHYLIP format only:

• Interleaved: • Sequencial:

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1. Give "fahA.prank.phylip" or "fahA.mafft.phylip" as input to the phyML webserver (don't forget to choose "Amino-acids" and enter your email)

2. Run it with the local installation of "phyml.bat"

• You should end up with a file: "fahA.prank.phylip_phyml_tree.txt"

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RAxML Web server: http://phylobench.vital-it.ch/raxml-bb/ Similar maximum likelihood (ML) methodology as phyML, but much faster

Faster results Better results in same run-time

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1. Give "fahA.prank.phylip" or "fahA.mafft.phylip" as input to the RAxML webserver (don't forget to tick "Protein sequences" and enter your email)

• Save the resulting tree file as: "fahA.prank.phylip.raxml"

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FigTree: tree visualization and figure creation

Manipulate a node

Manipulate a clade

Manipulate a taxon

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1. Open "fahA.prank.phylip_phyml_tree.txt" in FigTree

2. Play around with the different options and make a pretty figure!

1. Find out how to color specific clades, as below

2. Try each of the three options under "Layout"

3. Export a figure in PDF format(File Export Graphic…)

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Final Questions…

Thanks for your attention