Bioinformatics Werkbespreking 2006-11-07

• 1 – PhyloPat: phylogenetic pattern analysis of eukaryotic genes (20 slides)

• 2 – Chicken-human immunogenomics project (9 slides)

PhyloPatphylogenetic pattern analysis

of eukaryotic genes

Tim Hulsen

2006-10-17

BeNeLux BioInformatics Conference 2006

Introduction (1)

• Phylogenetic patterns show presence/absence of genes over a certain set of species:e.g. for 10 species: 0011101011

• Very useful for all kinds of evolutionary analyses:– Origin of certain genes– Deletion of certain genes– Clustering of genes with similar patterns: likely

to have similar function / be in same pathway

Introduction (2)

• Earlier phylogenetic pattern initiatives:– Phylogenetic Pattern Search (PPS), incorporated into

COG (Natale et al., 2000)– Extended Phylogenetic Patterns Search (EPPS)

(Reichard & Kaufmann, 2003)– Incorporated into OrthoMCL-DB (Chen et al., 2006)

• All applied on proteins, not on genes! PhyloPat: phylogenetic pattern analysis of

eukaryotic genes

Method

• Genes: easier to check for lineage-specific expansions (no alternative transcripts or splice forms); less redundant

• Basis: Ensembl (EnsMart) database: 21 fully available genomes (i.e. no Pre! versions or low coverage genomes): S. cer. to H. sap.

• Make use of accurate Ensembl orthology pipeline (combination of BLAST,SW,MUSCLE and PHYML)

• Single linkage cluster algorithm: create orthologous groups containing ALL genes in Ensembl

Results

• 446,825 genes were clustered into 147,922 groups, using 3,164,088 orthologies from 21 species

• Species ordered from ‘low’ ( ) to ‘high’ ( ), i.e. approximate distance to human :

• Can be queried in several ways• Output in HTML, Excel or plain text format

Web interface

http://www.cmbi.ru.nl/phylopat

Pattern/ID Search

• Binary string:0=absent, 1=present, *=absent/presente.g. ‘00000********11111111’: must be absent in non-chordata , must be present in all mammals

• MySQL regular expression:e.g. ‘^0*1{10}0*$’ gives all genes that occur only in ten subsequent species

• Input list of Ensembl/EMBL IDs (PhyloPat contains EMBL to Ensembl mapping)

Output

Phylogenetic Tree

Oligo-/Polypresent Genes• Oligopresent: present in only one/two species (oligo=few),

e.g. ‘000000010000000000100’• These two species should be highly related

1. C. sav C. int 1737 div. 100 Mya(Boffelli et al.,

2004)

2. T. nig T. rub 1572 div. 85 Mya(Yakanoue et al., 2006)

3. A. gam A. Aeg 1058 div. 140 Mya(Service, 1993)

4. P. tro H. sap 887 div . 6 Mya(Glazko & Nei, 2003)

5. R. nor M. Mus 713 div. 20 Mya(Springer et al., 2003)

• Polypresent: present in all species, except for one/two (poly=many),

e.g. ‘111110111110111111111’• These two species should be related too; similar analysis possible

Omnipresent genes

• Omnipresent: present in all 21 species (omni=all): ‘111111111111111111111’

• Currently 1001 omnipresent groups

• Tend to have very general/important functions, mostly involved in transcription/translation

FatiGO analysis

• FatiGO: connection with GO terms, KEGG pathways, InterPro domains, etc. (El-Shahrour et al., 2004)

• Analysis of all human genes in output by just one mouse click

• e.g. omnipresent genes:

Other possibilities

• Anti-correlating patterns:

e.g. ‘001111100011000000000’

and ‘110000011100111111111’

could be completely different, or very similar (analogous)!

• Easy homology-inferred functional annotation (using information from other genes in the same lineage)

Case study: Hox genes (1)• Hox genes determine where limbs and other body

segments will grow in a developing embryo• Should exist mostly in vertebrates• Expansion in teleost fish species ( , 8-11);

seven Hox clusters instead of the mammalian four• Search Ensembl database for human genes with term

‘hox’ in annotation• 44 genes found -> enter in PhyloPat -> 32 groups found

(PP######)

Case study: Hox genes (2)PPID # genes per species phylogenetic pattern gene name(s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

Case study: Hox genes (3)PPID(s) name cl.A cl.B cl.C cl.D first sp. positionPP053829,085049 HOX1 HOXA1 HOXB1 HOXD1 T. nigrov. anteriorPP053847,053833 HOX2 HOXA2 HOXB2 T. nigrov. anteriorPP053836,053845,053834 HOX3 HOXA3 HOXB3 HOXD3 T. nigrov. PG3PP053832,053844,075622 HOX5 HOXA5 HOXB5 HOXC5 T. nigrov. centralPP053849 HOX6 HOXA6 HOXB6 HOXC6 T. nigrov. centralPP053835,053854 HOX9 HOXA9 HOXB9 HOXC9 HOXD9 T. nigrov. posteriorPP053827,084287,053846 HOX10 HOXA10 HOXC10 HOXD10 T. nigrov. posteriorPP053858,053840,053824 HOX11 HOXA11 HOXC11 HOXD11 T. nigrov. posteriorPP053838,087941 HOX12 HOXC12 HOXD12 T. nigrov. posteriorPP053842,089685,053828 HOX13 HOXA13 HOXB13 HOXC13 HOXD13 T. nigrov. posterior

PP053853,053830,024984,053839 HOX4 HOXA4 HOXB4 HOXC4 HOXD4 A. gamb. centralPP027791 TLX TLX1 TLX2 TLX3 A. gamb.

PP070659 HOX7 HOXA7 HOXB7 G. acul. central

PP049478 HOX8 HOXB8 HOXC8 HOXD8 C. intest. central

PP022041 MSX MSX1 MSX2 C. eleg.

‘First’vertebrate

Non-vertebrate

Non-vertebrate

Non-vertebrate

Vertebrate

Conclusions

• PhyloPat: quick and easy tool for phylogenetic pattern search on complete Ensembl database

• Also usable for study of lineage-specific expansions of genes

• Just updated to Ensembl v41 (released last Thursday); 5 new species:

D.nov E.tel L.afr O.cun O.lat+ extra option: gene neighborhood

Gene neighborhood

Equal color = belonging to same orthologous group

Conservation of gene order = functionally related

Acknowledgements

Supervision:• Peter Groenen• Jacob de Vlieg

Fruitful discussions:• Wilco Fleuren• Erik Franck• Nanning de Jong• Arnold Kuzniar

supervisor

head of group

suggestions

suggestions

suggestions

suggestions

Where to find• Web interface:

http://www.cmbi.ru.nl/phylopat

(accessible through www.cmbi.ru.nl and www.nbic.nl)

• Publication:

Hulsen T., Groenen P.M.A., de Vlieg J.

BMC Bioinformatics 2006, 7: 398

http://www.biomedcentral.com/1471-2105/7/398

• Powered by Ensembl:

http://www.ensembl.org/info/about/ensembl_powered.html

Bioinformatics Werkbespreking 2006-11-07

• 1 – PhyloPat: phylogenetic pattern analysis of eukaryotic genes (20 slides)

• 2 – Chicken-human immunogenomics project (9 slides)

Goals:- study evolution of genes/proteins involved in immune system, from chicken to human- check for expansions and deletions in families- zoom in to interesting families

Chicken-human immunogenomics project (part of Biorange SP3.2.2)

In collaboration with Martien Groenen, HinriKerstens (Animal Sciences Group, Wageningen UR)

Proteins -> Genes

• Earlier initiatives: based on proteins (Protein World, IPI, ParAlign, MCL)

• Disadvantages:– large scale computations needed for

orthology determination– Difficult to study lineage-specific expansions

because of alternative transcripts, isoforms– Difficult to connect to WUR synteny data

• --> Genes: connect to PhyloPat tool

PhyloPat• PhyloPat: queries the orthologies of all complete

genomes within Ensembl database using phylogenetic patterns

• Advantages:– Usage of accurate orthology determination of

Ensembl (BLAST/SW, MUSCLE, PHYML), single linkage clustering by ourselves)

– No alternative transcripts, isoforms– Easy to connect to WUR synteny data– 26 species, from S.cer. to H.sap.

• Disadvantage:– Genome information sometimes incomplete (but Pre-

versions and low coverage genomes are not included)

Immunophyle

• Application to immune system: parse through PhyloPat set using IRIS database

• Take all HUGO IDs from IRIS database, input in PhyloPat -> 585 immunologic lineages containing 18,933 genes from 26 species

• Divided into immunologic 22 categories from IRIS database (adaptive immunity, innate immunity, inflammation, chemotaxis, etc.

• Connected to GO, InterPro, KEGG, etc. by FatiGO

Immunophyle

• http://www.cmbi.ru.nl/immunophyle

CategoriesCat :: Category (# lineages, # genes) Lin Sc Ce Ag Aa Dm Cs Ci Tn Tr Ol Ga Dr Xt Gg Md Dn Bt Cf Et La Rn Mm Oc Mm Pt Hs Total

All :: All immunologic lineages (585,18933) 585 54 156193211214219239 876 830 824 8551015686 685 969 7401121948 870 802107011631131 818 1087115718933

InImm :: Innate Immunity (272, 8640) 272 17 51 77 89 81 81 93 351 355 339 351 420 304 295 466 355 566 435 416 384 517 571 539 384 535 568 8640

Inflm :: Inflammation (117, 4568) 117 13 34 45 57 43 53 55 202 200 194 197 237 179 150 227 200 267 221 215 197 263 302 271 194 265 287 4568

Chmtx :: Chemotaxis (54, 2374) 54 4 12 18 24 18 22 28 107 118 112 122 125 96 69 157 90 135 121 112 103 124 147 132 86 141 151 2374

Phago :: Phagocytosis (17, 890) 17 1 4 9 10 10 8 10 46 43 41 47 50 32 31 42 34 51 45 46 42 49 58 44 33 51 53 890

Compl :: Complement (33, 958) 33 0 3 13 7 7 11 19 45 41 43 43 54 37 31 50 36 58 45 48 34 60 62 55 43 54 59 958

Cy_Ch :: Cytokines and Chemokines (109, 2947) 109 2 11 14 20 18 18 18 122 119 124 119 148 92 120 144 106 219 173 143 133 175 187 195 143 190 194 2947

AdImm :: Adaptive Immunity (140, 4983) 140 17 44 37 40 48 59 62 212 207 204 219 253 158 170 246 188 330 260 225 223 276 315 324 244 303 319 4983

ClRsp :: Cellular Response (63, 2358) 63 6 26 20 23 22 36 41 106 101 102 100 119 78 96 116 93 138 112 105 104 124 148 148 111 137 146 2358

HmRsp :: Humoral Response (34, 1087) 34 3 9 8 8 9 8 9 48 46 48 45 47 37 40 49 43 60 58 55 50 61 65 76 68 65 72 1087

BMImm :: Barrier and Mucosal Immunity (18, 713) 18 0 1 10 9 15 2 4 20 25 17 27 24 18 11 33 30 68 42 38 32 48 58 47 25 52 57 713

Devlp :: Development of Immune System (50, 2044) 50 5 18 23 25 23 22 29 109 90 89 96 124 64 72 106 74 109 108 103 86 114 122 116 92 108 117 2044

AgPrc :: Antigen Processing (31, 830) 31 3 8 9 11 11 10 12 34 31 36 38 56 22 25 39 40 49 35 37 36 39 40 63 35 54 57 830

PtSig :: Immune Pathway or Signalling (224, 8245) 224 13 63 70 87 81 93 102 400 381 382 390 480 301 296 446 302 454 415 371 344 459 508 489 337 480 501 8245

Recpt :: Receptor (118, 3506) 118 2 18 16 20 18 18 24 148 151 150 158 187 125 124 165 141 205 191 170 154 227 240 226 156 231 241 3506

IndIm :: Induced by Immunomodulator (86, 3487) 86 7 23 28 25 40 29 32 172 163 159 175 200 129 122 171 130 224 184 154 154 198 218 197 151 193 209 3487

ImDef :: Involved in Immunodeficiency (30, 1013) 30 4 8 15 12 9 18 28 44 44 41 38 64 35 42 48 34 61 45 45 43 54 56 58 52 56 59 1013

AutIm :: Involved in Autoimmunity (19, 530) 19 0 1 12 6 1 5 3 23 24 26 23 32 18 20 25 19 29 30 28 22 29 30 31 29 31 33 530ExpIT :: Expressed Primarily in Immune Tissues (134,

3970)134 11 22 36 27 31 32 42 157 160 153 173 186 137 118 249 155 238 201 179 170 257 274 260 158 261 283 3970

Other :: Other (43, 1843) 43 9 28 32 31 37 23 25 99 82 80 90 105 86 78 84 78 98 82 75 74 93 92 96 73 94 99 1843

InKil :: Innate NK Killing (33, 1015) 33 1 4 9 9 6 6 8 31 37 28 32 35 44 23 38 49 70 49 52 50 74 88 72 38 80 82 1015

RlDis :: Related to Disease (91, 3141) 91 6 14 32 28 32 25 34 151 143 133 144 176 115 128 159 131 190 159 153 133 170 184 184 145 182 190 3141

Coagl :: Coagulation (51, 2624) 51 5 25 36 44 33 31 33 132 123 122 124 154 114 81 124 108 141 123 121 109 145 162 139 106 138 151 2624

Example: Toll-like receptors

GeneGoMetaCore,canonicalpathway

Example: Toll-like receptors

Lineage Sc Ce Ag Aa Dm Cs Ci Tn Tr Ol Ga Dr Xt Gg Md Dn Bt Cf Et La Rn Mm Oc Mm Pt Hs HUGO

IP406 0 0 0 0 0 0 0 0 0 0 0 0 2 1 1 2 2 2 0 1 2 2 3 2 3 3TLR1/6/10

IP308 0 0 0 0 0 0 0 1 1 1 1 1 0 2 1 0 1 1 0 0 1 1 1 1 1 1TLR2

IP197 0 0 0 0 0 0 0 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1TLR3

IP430 0 0 0 0 0 0 0 0 0 0 0 0 0 1 1 1 1 0 1 1 1 1 1 1 1 1TLR4

IP289 0 0 0 0 0 0 0 2 2 2 3 1 1 1 1 0 1 1 1 0 1 0 1 0 0 1TLR5

IP359 0 0 0 0 0 0 0 1 1 1 2 4 1 1 1 1 0 1 2 2 1 2 2 1 2 2TLR7/8

IP550 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 1 1 0 1 1TLR9

IP458 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 1 1 1 1 1 1 1 0 1 1IRAK1

IP475 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 1 1 1 1 1 0 1 1 1 1 1IRAK2

IP397 0 0 0 0 0 0 0 0 0 0 1 3 0 0 1 1 1 1 1 1 1 1 1 1 1 1IRAK3/IRAK-M

IP321 0 0 0 0 0 0 0 1 1 1 1 1 0 1 1 1 1 1 1 1 1 1 1 1 1 1IRAK4

IP539 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 1 1 1 1 1 1 1 1 1IL4

IP421 0 0 0 0 0 0 0 0 0 0 0 0 1 1 1 1 1 1 1 1 1 1 1 0 1 1IL6

IP294 0 0 0 0 0 0 0 1 1 1 1 2 1 0 1 1 1 1 1 1 0 0 1 1 1 1IL8

IP078 0 5 0 0 0 1 2 3 3 3 4 1 3 4 5 3 4 4 3 4 4 4 4 4 4 4LBP

IP484 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 1 1 1 1 1 1 1 1 1 1LTA

IP057 0 1 1 0 0 1 1 1 1 1 0 1 1 1 1 0 1 1 1 1 1 1 1 0 1 1TOLLIP

IP045 0 1 1 1 1 2 2 4 4 3 4 4 3 3 3 1 4 4 3 3 4 4 4 2 4 4NFKB1,NFKB2,NFKBIA

IP132 0 0 1 0 1 1 0 1 1 1 1 1 1 1 1 2 2 1 1 1 1 1 2 1 1 1TRAF6

IP059 0 1 1 1 1 1 1 7 7 5 6 4 4 0 3 0 3 2 2 0 2 3 2 0 2 3JUN/JUNB/JUND

IP145 0 0 0 1 0 1 1 1 1 1 2 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1MAP3K7/TAK1

IP158 0 0 0 0 0 1 1 2 2 2 2 3 2 2 2 2 2 2 2 1 2 2 2 2 2 2MAP3K7IP2

IP222 0 0 0 0 0 0 0 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 0 1MAP3K14

IP101 0 1 1 1 1 0 0 5 4 5 5 4 1 3 4 3 3 3 3 3 3 3 3 3 3 3MAP4K4

IP434 0 0 0 0 0 0 0 0 0 0 0 0 0 1 1 0 1 1 0 1 1 1 1 0 1 1MAP2K3

Check ImmunoPhyle for each gene involved in the TLR pathway:

Green: ‘first’ occurrence Red: deletion

Current/future directions

• Connect to literature (CoPub?)

• Connect to expression data, protein interaction data

• Zoom in to families: immunology expertise needed!