Finding Orthologous Groups

René van der Heijden

What is this lecture about?

• What is ‘orthology’?• Why do we study gene-ancestry/gene-trees

(phylogenies)?• Several approaches to find orthologous genes• High-resolution orthology• Steps involved• Things to think about (homework)

Homology

Genes are homologous if and only if theyderive from the same ancestral gene

• Sufficient sequence similarity proofs homology

• Very dissimilar sequences:PSI blast, HMM searches

Homologous genes tend to have similar functions

The usual range

Homologous genes tend to have similar functions

Accurate function prediction requires something better than

homology

OrthologyOrthology

Duplications, Speciations, and Orthology

Evolution results in:• Growing number of genes

– Gene duplications– Horizontal gene transfer– De novo generation

• Growing number of species

The fate of gene duplicates:• Perish• Find a new functional niche

Tendency for functional expansionTendency for functional expansion

Duplications, Speciations, and Orthology

Two genes in two species are orthologous ifthey derive from one gene

in their last common ancestor

• Orthologous genes are likely to have the same function

• Much stronger than “tend to have similar function”

Duplications, Speciations, and Orthology

primalancestor

present genes

evolutionary distance

Homologs, Orthologs,and Paralogs

• Homologous: one common ancestral gene

• Orthologous: separated by a speciation event

• Paralogous: separated by a duplication event

• Orthologs and Paralogs must be Homologs

Are there homologous genes whichare not orthologous nor paralogous?

The view on orthology and The view on orthology and paralogy is relative to a paralogy is relative to a

certain speciationcertain speciation

Inparalogs and Outparalogs

• Both, In- and Outparalogous genes are separated by a gene duplication event

• For Inparalogs, the duplication event is not followed by speciation(s)

• Outparalogs are separated by a duplication event, followed by speciation(s)

• Inparalogs are recent paralogs• Outparalogs are more ancient paralogs

Are Inparalogs Orthologs ?Are Inparalogs Orthologs ?Depends on your definition:

Yes: two genes are orthologous ifthey derive from one gene

in their last common ancestorNo: two genes are orthologous if

they are only separated by cell division events

Reading Gene-Trees

Although genes spec1,1 and spec2,1 are closer relatives, their distance is larger than that between spec1,1 and spec3,1

The tree suggests at least 2 gene losses

In-, and Outparalogs, Orthologs, and Co-orthologs

More examples

www = What, Why, and hoW?

• What: Orthologous genes are separated by cell division only

• Why:Orthologous genes are likely to have the same function

• How:Yes, how can orthologous relations be established ?

Several approaches

• The COG approach

• InParanoid

• Tree-based methods

COG approach

• Based on blast hits

• Establishment and extension of triangles:

COG approachII

Extension oforthologous groups

InParanoid I

• Method denotes– IN- and OUTparalogs– For TWO species

• Find all hits from species A on B

• Find all hits from species B on A

• Find all bi-directional best hits (BBH)– These for putative orthologs

InParanoid II

• Find all hits from A on A

• Find all hits from B on B

• Find all InParalogs– These are all hits better than the orthologs– Better => more recently split

InParanoid III

• Putative orthologous pairs are curated by an outgroup species C

• InParalogs are given a confidence value

• Bootstrapping is used to give confidence values for orthologous pairs

Genes with promiscuous domains

• Gene A may hit on gene B because of a shared domain X

• Gene B may hit on gene C because of a shared domain Y

• Promiscuous domains require (manual) curation

Tree-based methods

1. Get all homologous genes

2. Make multiple alignments

3. Generate phylogenetic gene trees

4. Analyze trees

• Uncertainty in multiple alignment?• Different methods for distance calculations• Superpose a trusted species tree?• How to assess a level of accuracy?

The Phylogenetic Gene-Tree• Multiple alignment for all genes

• Distance matrix calculation– Kimura correction– PAM model– Categories model

• Large trees: distance-based methods– Neighbor Joining

Uncertainty in trees

• Evolutionary noise– Differing rates of evolution

– Convergent evolution (low complexity, coiled coils)

– Promiscuous domains (recombination, fusion, fission)

• Use of heuristic methods– Multiple alignment

– Tree making

Analyze trees … but don’t trust them fully

• Rigid analysis suggests many duplications and losses• Presume scp branch is wrongly placed!

If this is correct …. this can’t be

Three orthologous groups suggesting 15 gene losses

Considering one wrongly placedgene leaves only 2 gene losses

Analyze trees … but don’t trust them fully

• And if we accept wrong placement of branches …

High-res versus Low-res

• Many,

• Complete, and

• Closely related

genomes

Challenge:Automatic Orthology assignment

Things to think about (homework)

• Select a partner• Collect a gene tree (and some copies)• Carefully deduce which nodes are duplications and

which are speciations• Denote which genes are orthologous to each other

(orthologous groups)• Select interesting parts to predict what

– The COG procedure would say– InParanoid would say– What would have happened if some genes (or species) where

not involved in the analysis

Homework: also think about …