The evolution of the immune system in chicken and higher

vertebrates

@ Organon, Oss

2005-09-20

Tim Hulsen

Biorange Project SP3.2.2

• Chicken immunosystem project is part of WP1, “Translational Medicine through Comparative Genomics and Integration”

• Partners:– Animal Breeding and Genetics Group,

Wageningen UR (Prof. dr. Martien Groenen)– Avian Cytokines Group, Institute for Animal

Health, Compton (UK) (Prof. dr. Pete Kaiser)

• Jack Leunissen (WUR) also part of WP1

M. Groenen: chicken sequencing

Kaiser: chicken immune system

Introduction

• Goal: developing an insight in the recent evolution of the immune system

• Usage of a more distant species: chicken (recently sequenced)

• Support by experimental data

Overview

1. Find IS-related proteins

2. Determine orthologies

3. Pfam annotation

4. Panther annotation

5. Zooming in

Step 1: Find IS-related proteins

• IRIS: “Immunogenetic Related

Information Source”• number of immune genes: 1562 (out of 21389 in

LocusLink)• percentage of genome related to immunity:

7.30% • 1562 LocusLink proteins mapped to our Protein

World set: 1381 proteins

Step 1: Find IS-related proteins

Step 1: Find IS-related proteins

• GO: Gene Ontology• collaborative effort to address the need for

consistent descriptions of gene products in different databases

• Checked human GO annotation for certain terms: “immunology”,”cytokine”,etc.

• 1515 proteins in human Potein World set

Step 1: Find IS-related proteins

• Result:

– 1381 proteins through IRIS

– 1515 proteins through GO

– 1929 proteins total

414

548

967

IRIS only

GO only

IRIS & GO

Step 2: Determine orthologies

• Study evolution from chicken (Gg) to rat (Rn), mouse (Mm) and human (Hs):

– Hs<->Mm– Hs<->Rn– Hs<->Gg– Mm<->Rn– Mm<->Gg– Rn<->Gg

• Two methods: Best Bidirectional Hit (BBH) and PhyloGenetic Tree (PGT)

Best Bidirectional Hit (BBH)

• Very easy and quick• Human protein (1) SW best

hit in mouse/rat (2)• Mouse/rat protein (2) SW

best hit in human (3)• If 3 equals 1, the human and

mouse/rat protein are considered to be orthologs

Step 2: Determine orthologies

BBH Hs Mm Rn Gg

Hs 1929 1145 1046 704

Mm - - 863 345

Rn - - - 488

Gg - - - -

PhyloGenetic Tree (PGT) Human

Human, mouse, rat, chicken

Hs, Mm, Rn, Gg

Z>20 RH>0.5*QL

~25,000 groups

PHYLOME

SELECTION OF HOMOLOGS

ALIGNMENTS AND TREES

PROTEOME

PROTEOMES

TREE SCANNING

LIST Hs-Mm pairsHs-Rn pairsHs-Gg pairsMm-Rn pairsMm-Gg pairsRn-Gg pairs

Step 2: Determine orthologies

PGT Hs Mm Rn Gg

Hs 1929 2301 1819 1129

Mm - - 2873 2087

Rn - - - 2142

Gg - - - -

Step 3: Pfam annotation

• Pfam: “Protein families database of

alignments and HMMs”• collection of protein families and domains• Pfam contains multiple protein alignments and

profile-HMMs of these families• 75% of protein sequences have at

least one match to Pfam• 1700 IS-related proteins mapped to 584 Pfam

families (2814 mappings)

Step 3: Pfam annotation

BBH Hs Mm Rn Gg PGT Hs Mm Rn Gg

Hs 1929 1145 1046 704 Hs 1929 2301 1819 1129

Mm - - 863 345 Mm - - 2873 2087

Rn - - - 488 Rn - - - 2142

Gg - - - - Gg - - - -

BBHpfam Hs Mm Rn Gg

PGTpfam Hs Mm Rn Gg

Hs 1776 1069 974 639 Hs 1776 2135 1700 1040

Mm - - 795 312 Mm - - 2846 2070

Rn - - - 442 Rn - - - 2125

Gg - - - - Gg - - - -

Step 3: Pfam annotation

62%11%

13%

14%

Hs

Hs+Mm

Hs+Mm+Rn

Hs+Mm+Rn+Gg

Step 4: Panther annotation

• PANTHER: “Protein ANalysis THrough Evolutionary Relationships”• designed to classify proteins (and their genes) in

order to facilitate high-throughput analysis• proteins have been classified according to

families and subfamilies, molecular functions, biological processes, pathways

• contains over 6683 protein families, divided into 31,705 functionally distinct protein subfamilies

• 1872 IS-related proteins mapped to 970 Panther families (4667 subfamilies, 14737 mappings)

Step 4: Panther annotation

BBH Hs Mm Rn Gg PGT Hs Mm Rn Gg

Hs 1929 1145 1046 704 Hs 1929 2301 1819 1129

Mm - - 863 345 Mm - - 2873 2087

Rn - - - 488 Rn - - - 2142

Gg - - - - Gg - - - -

BBHpanther Hs Mm Rn Gg

PGTpanther Hs Mm Rn Gg

Hs 1872 1125 1029 688 Hs 1872 2266 1793 1118

Mm - - 846 339 Mm - - 2729 1970

Rn - - - 481 Rn - - - 2008

Gg - - - - Gg - - - -

Step 4: Panther annotation

39%

15%

20%

26%

Hs

Hs+Mm

Hs+Mm+Rn

Hs+Mm+Rn+Gg

Step 5: Zooming in

• Which families are ‘new’ in human?

• Which orthologs have a different domain structure through evolution?

• Which human proteins don’t have orthologs in the other species?

• Any other interesting stuff

Future directions

• Include paralogs in our analysis (makes possible checking which families only exist in mouse/rat/chicken)

• Combine our findings with research at WUR: synteny between human and chicken

• Take a look at ratio of non-synonymous to synonymous substitutions (dN/dS)

Credits

• NV Organon:– Peter Groenen– Wilco Fleuren

• Wageningen UR:– Martien Groenen– Hindrik Kerstens

• Compton (UK):– Pete Kaiser