Prsentation PowerPoint

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Bioinformatics of nuclear receptors Marc
Robinson-Rechavi Joint Center for Structural
Genomics UCSD, La Jolla, CA, USA marc_at_sdsc.edu
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Types of BLAST
query database program DNA DNA blastn protein
protein blastp translated DNA protein blastx p
rotein translated DNA tblastn translated DNA
translated DNA tblastx
blastn check close homologs, pseudogenes, close
non coding blastp compare NRs tblastn,
blastx compare NR to non annotated DNA genomes,
ESTs
http//www.ncbi.nlm.nih.gov/BLAST/
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pre-genome 48 NR genes, including 46 LBDs IHGSC
2001 60 NR genes, including 60 LBDs Venter et al
2001 59 NR genes, including 47
LBDs Robinson-Rechavi et al 2001, Maglich et al
2001 48 NR genes, including 46 LBDs 1 "?"
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Nuclear receptors in dbEST http//www.ncbi.nlm.nih
.gov/dbEST/
BLAST between 104 human NR alternative
transcripts and 5 484 645 EST sequences
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276 NR/EST tissues 83 normal 71 tumoral 31
disease 91 uncharacterized
55 NR/EST tissues with gt 10 000 EST 28 normal
24 tumoral 1 disease 2 uncharacterized
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Number of tissues where each NR is detected by
ESTs
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there is a vertebrate FXRb it became a pseudogene
in primates
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Some notes on phylogenetic reconstruction
1- Quality of the alignment do not trust
automatic alignment check by eye (SeaView,
Jalview, ...) remove partial sequences which
make you loose a lot of sites
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Some notes on phylogenetic reconstruction
1- Quality of the alignment do not trust
automatic alignment check by eye (SeaView,
Jalview, ...) remove partial sequences which
make you loose a lot of sites 2- Phylogenetic
reconstruction choose model substitution
matrix (Dayhoff, Grantham, JTT, ...) rate
heterogeneity between sites gamma law choose
method ML if you can, NJ if you must 3-
Confidence in results bootstrap (1000-2000
replicates) Likelihood tests (KH, SH, AU)
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recommended software PhyML http//atgc.lirmm.fr/p
hyml/ http//evolution.genetics.washington.edu/phy
lip/software.html
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Some notes on phylogenetic reconstruction
1- Quality of the alignment do not trust
automatic alignment check by eye (SeaView,
Jalview, ...) remove partial sequences which
make you loose a lot of sites 2- Phylogenetic
reconstruction choose model substitution
matrix (Dayhoff, Grantham, JTT, ...) rate
heterogeneity between sites gamma law choose
method ML if you can, NJ if you must 3-
Confidence in results bootstrap (1000-2000
replicates) Likelihood tests (KH, SH, AU) 4-
Draw tree NJplot, TreeView root it!
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Exemple of tree rooting with an outgroup
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Dedicated databases

• Nuclear Receptor Ressource, NURSA
• NucleaRDB
• Nurebase NRbase

http//www3.oup.co.uk/nar/database/c/
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Nuclear receptor ressource
http//nrr.georgetown.edu/NRR/nrrhome.htm
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Nuclear receptor ressource
http//www.nursa.org
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NucleaRDB
http//receptors.ucsf.edu/NR/
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NucleaRDB
The current NucleaRDB release contains 2225 point
mutations (PMs) extracted from 546 articles. 396
point mutations have not yet been checked
manually and therefore, you may found some false
positives.
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Nurebase
http//www.ens-lyon.fr/LBMC/laudet/nurebase/nureba
se.html
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Nurebase
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Nurebase
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Nurebase
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Nurebase
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Nurebase
tree in the WWW browser clicable gene names
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Nurebase
format organ stage tumor
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Nurebase
keyword breast tumor OR cervix tumor OR uterus
tumor -gt 16 human NRs keyword breast tumor AND
cervix tumor AND uterus tumor -gt human
ERRa keyword fetal AND tumor -gt 14 NRs,
probably relevant to cell proliferation
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Nurebase alternative transcripts
local BLAST
global CLUSTAL
real redundance
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alternative transcripts
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lt 100
take gaps into account
information on completeness is there a start?
a stop?
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Main transcript of human THRa (cDNA)
ID THA0HOMS1 standard DNA PRI 1233
BP. XX AC THA0HOMS1 XX SV THA0HOMS1.1 XX DT
09-MAY-1995 (Rel. 03, Created) DT 23-OCT-2001
(Rel. 03, Last updated, Version 1) XX DE
Thyroid hormone receptor alpha, complete
cds. XX KW NR1A1 TRa c-erbA-1 THRA KW
Thyroid hormone Triiodothyronine T3. XX OS
Homo sapiens (Homo sapiens) OC Eukaryota
Metazoa Chordata Craniata Vertebrata
Euteleostomi OC Mammalia Eutheria Primates
Catarrhini Hominidae Homo. XX RN 1 RX
MEDLINE 89127255 PUBMED 2464749. RA
Nakai,A., Sakurai,A., Bell,G.I. and
DeGroot,L.J. RT "Characterization of a third
human thyroid hormone receptor RT coexpressed
with other thyroid hormone receptors in several
tissues" RL Mol. Endocrinol. 2 (11), 1087-1092
(1988). XX DR NRBPROT AAA66021 THA_HOMS1. DR
EMBL M24748 . XX CC Draft entry and computer
readable sequence for 1 kindly submitted CC
by A. Nakai, 15-MAY-1989. XX FH Key
Location/Qualifiers FH FT source
1..1233 FT /organism"Homo
sapiens" FT /db_xref"taxon9606
" FT /map"17q11.2-q12" FT
/clone"lambda-Me2." FT
/tissue_type"skeletal muscle" FT
/dev_stage"foetus" FT CDS_pept
1..1233
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Alternative transcript of human THRa (cDNA)
ID THA1HOMS1 standard mRNA PRI 1473
BP. XX AC THA1HOMS1 XX SV THA1HOMS1.1 XX DT
12-SEP-1994 (Rel. 03, Created) DT 22-AUG-2002
(Rel. 03, Last updated, Version 1) XX DE
Thyroid hormone receptor alpha, isoform, complete
cds. XX KW NR1A1 TRa c-erbA-1 THRA KW
alternative transcript splice variant
alternative splicing. XX OS Homo sapiens (Homo
sapiens) OC Eukaryota Metazoa Chordata
Craniata Vertebrata Euteleostomi OC
Mammalia Eutheria Primates Catarrhini
Hominidae Homo. XX RN 1 RX MEDLINE
89195219 PUBMED 2539258. RA Miyajima,N.,
Horiuchi,R., Shibuya,Y., Fukushige,S.,
Matsubara,K., RA Toyoshima,K. and
Yamamoto,T. RT Two erbA homologs encoding
proteins with different T3 binding RT
capacities are transcribed from opposite DNA
strands of the same RT genetic locus" RL
Cell 57 (1), 31-39 (1989). XX DR NRBPROT
AAA35783 THA_HOMS1. DR EMBL M24899 . XX CC
On Sep 13, 1994 this sequence version replaced
gi341273. XX FH Key
Location/Qualifiers FH FT source
1..1473 FT /organism"Homo
sapiens" FT /db_xref"taxon9606
" FT CDS_pept 1..1473
keyword "TRa" and species "homo sapiens" -gt 8
cDNAs
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Alternative transcript of human THRa (cDNA)
THA5HOMS1 ----------------------------------
-------------------------- THA6HOMS1
--------------------------------------------------
---------- THA7HOMS1 -----------------------
------------------------------------- THA3HOMS1
----------------------------------------------
-------------- THA2HOMS1 -------------------
----------------------------------------- THA1HOMS
1 ------------------------------------------
------------------ THA4HOMS1
--------------------------------------------------
---------- THA0HOMS1 TGCCGGGGGGGCCAGTGTGCCCA
CCCCAGTCTCTTGGCGTGCTGGAGGGCATCCTGGATG

THA5HOMS1
------------ATGGAACAGAAGCCAAGCAAGGTGGAATGTGGGTCAGA
CCCAGAGGAG THA6HOMS1 ------------ATGGAACAGAA
GCCAAGCAAGGTGGAGTGTGGGTCAGACCCAGAGGAG THA7HOMS1
------------ATGGAACAGAAGCCAAGCAAGGTGGAGTGTGGGT
CAGACCCAGAGGAG THA3HOMS1 ------------ATGGAAC
AGAAGCCAAGCAAGGTGGAGTGTGGGTCAGACCCAGAGGAG THA2HOMS
1 ------------ATGGAACAGAAGCCAAGCAAGGTGGAGTGT
GGGTCAGACCCAGAGGAG THA1HOMS1
------------ATGGAACAGAAGCCAAGCAAGGTGGAGTGTGGGTCAGA
CCCAGAGGAG THA4HOMS1 ------------ATGGAACAGAA
GCCAAGCAAGGTGGAGTGTGGGTCAGACCCAGAGGAG THA0HOMS1
GAATTGAAGTGAATGGAACAGAAGCCAAGCAAGGTGGAGTGTGGGT
CAGACCCAGAGGAG

THA5HOMS1 AACAGTGCCAGGTCACCAGATGGAAAGCGAAAAA
GAAAGAACGGCCAATGTTCCCTGAAA THA6HOMS1
AACAGTGCCAGGTCACCAGATGGAAAGCGAAAAAGAAAGAACGGCCAATG
TTCCCTGAAA THA7HOMS1 AACAGTGCCAGGTCACCAGATGG
AAAGCGAAAAAGAAAGAACGGCCAATGTTCCCTGAAA THA3HOMS1
AACAGTGCCAGGTCACCAGATGGAAAGCGAAAAAGAAAGAACGGCC
AATGTTCCCTGAAA THA2HOMS1 AACAGTGCCAGGTCACCAG
ATGGAAAGCGAAAAAGAAAGAACGGCCAATGTTCCCTGAAA THA1HOMS
1 AACAGTGCCAGGTCACCAGATGGAAAGCGAAAAAGAAAGAAC
GGCCAATGTTCCCTGAAA THA4HOMS1
AACAGTGCCAGGTCACCAGATGGAAAGCGAAAAAGAAAGAACGGCCAATG
TTCCCTGAAA THA0HOMS1 AACAGTGCCAGGTCACCAGATGG
AAAGCGAAAAAGAAAGAACGGCCAATGTTCCCTGAAA

THA5HOMS1
ACCAGCATGTCAGGGTATATCCCTAGTTACCTGGACAAAGACGAGCAGTG
TGTCGTGTGT THA6HOMS1 AGCAGCATGTCAGGGTATATCCC
TAGTTACCTGGACAAAGACGAGCAGTGTGTCGTGTGT THA7HOMS1
ACCAGCATGTCAGGGTATATCCCTAGTTACCTGGACAAAGACGAGC
AGTGTGTCGTGTGT THA3HOMS1 ACCAGCATGTCAGGGTATA
TCCCTAGTTACCTGGACAAAGACGAGCAGTGTGTCGTGTGT THA2HOMS
1 ACCAGCATGTCAGGGTATATCCCTAGTTACCTGGACAAAGAC
GAGCAGTGTGTCGTGTGT THA1HOMS1
ACCAGCATGTCAGGGTATATCCCTAGTTACCTGGACAAAGACGAGCAGTG
TGTCGTGTGT THA4HOMS1 ACCAGCATGTCAGGGTATATCCC
TAGTTACCTGGACAAAGACGAGCAGTGTGTCGTGTGT THA0HOMS1
ACCAGCATGTCAGGGTATATCCCTAGTTACCTGGACAAAGACGAGC
AGTGTGTCGTGTGT

THA5HOMS1 GGGGACAAGGCAACTGGTTATCAC
TACCGCTGTATCACTTGTGAGGGCTGCAAGGGCTTC THA6HOMS1
GGGGACAAGGCAACTGGTTATCACTACCGCTGTATCACTTGTGAGGG
CTGCAAGGGCTTC THA7HOMS1 GGGGACAAGGCAACTGGTTA
TCACTACCGCTGTATCACTTGTGAGGGCTGCAAGGGCTTC THA3HOMS1
GGGGACAAGGCAACTGGTTATCACTACCGCTGTATCACTTGTG
AGGGCTGCAAGGGCTTC THA2HOMS1
GGGGACAAGGCAACTGGTTATCACTACCGCTGTATCACTTGTGAGGGCTG
CAAGGGCTTC THA1HOMS1 GGGGACAAGGCAACTGGTTATCA
CTACCGCTGTATCACTTGTGAGGGCTGCAAGGGCTTC THA4HOMS1
GGGGACAAGGCAACTGGTTATCACTACCGCTGTATCACTTGTGAGG
GCTGCAAGGGCTTC THA0HOMS1 GGGGACAAGGCAACTGGTT
ATCACTACCGCTGTATCACTTGTGAGGGCTGCAAGGGCTTC


THA5HOMS1 AAACACAACATTCCGCACTTCTGGCCCAAGCTGC
TGATGAAGGAGAGAGAAGT--GCAGA THA6HOMS1
AAACACAACATTCCGCACTTCTGGCCCAAGCTGCTGATGAAGGAGAGAGA
AGT--GCAGA THA7HOMS1 AAACACAACATTCCGCACTTCTG
GCCCAAGCTGCTGATGAAGGAGAGAGAAGT--GCAGA THA3HOMS1
AAACACAACATTCCGCACTTCTGGCCCAAGCTGCTGATGAAGGAGA
GAGAAGT--GCAGA THA2HOMS1 AAACACAACATTCCGCACT
TCTGGCCCAAGCTGCTGATGAAGGAGAGAGAAGT--GCAGA THA1HOMS
1 AAACACAACATTCCGCACTTCTGGCCCAAGCTGCTGATGAAG
GAGAGAGAAGT--GCAGA THA4HOMS1
AAACACAACATTCCGCACTTCTGGCCCAAGCTGCTGATGAAGG-------
---------- THA0HOMS1 AAACACAACATTCCGCACTTCTG
GCCCAAGCTGCTGATGAAGGTGACTGACCTCCGCATG

THA5HOMS1
GTTCGATTCTGT-ACAAGGGG-GCAGCGGCAGAAGGCCGGCCGGGCGGGT
CACTGGGCG- THA6HOMS1 GTTCGATTCTGT-ACAAGGGG-G
CAGCGGCAGAAGGCCGGCCGGGCGGGTCACTGGGCG- THA7HOMS1
GTTCGATTCTGT-ACAAGGGG-GCAGCGGCAGAAGGCCGGCCGGGC
GGGTCACTGGGCG- THA3HOMS1 GTTCGATTCTGT-ACAAGG
GG-GCAGCGGCAGAAGGCCGGCCGGGCGGGTCACTGGGCG- THA2HOMS
1 GTTCGATTCTGT-ACAAGGGG-GCAGCGGCAGAAGGCCGGCC
GGGCGGGTCACTGGGCG- THA1HOMS1
GTTCGATTCTGT-ACAAGGGG-GCAGCGGCAGAAGGCCGGCCGGGCGGGT
CACTGGGCG- THA4HOMS1 -----------------------
------------------------------------- THA0HOMS1
ATCGGGGCCTGCCACGCCAGCCGCTTCCTCCACATGAAAGTCGAGT
GCCCCACCGAACTC

THA5HOMS1 -TCCACCCGGAAGGACAGCAGCTTCTCGGAAT
GCATGTTGTTCAG----------GGTCC THA6HOMS1
-TCCACCCGGAAGGACAGCAGCTTCTCGGAATGCATGTTGTTCAG-----
-----GGTCC THA7HOMS1 -TCCACCCGGAAGGACAGCAGCT
TCTCGGAATGCATGTTGTTCAG----------GGTCC THA3HOMS1
-TCCACCCGGAAGGACAGCAGCTTCTCGGAATGCATGTTGTTCAG-
---------GGTCC THA2HOMS1 -TCCACCCGGAAGGACAGC
AGCTTCTCGGAATGCATGTTGTTCAG----------GGTCC THA1HOMS
1 -TCCACCCGGAAGGACAGCAGCTTCTCGGAATGCATGTTGTT
CAG----------GGTCC THA4HOMS1
--------------------------------------------------
------GTCC THA0HOMS1 TTCCCCCCACTCTTCCTCGAGGT
CTTTGAGGATCAGGAAGTCTAAAGCCTCAGGCGGCCA

THA5HOMS1
GCAGGTCCG-GCAGCT--TGAGCAGCAGCTTGGTGAAGCGGGAAGTCTCC
AAGGGCCGGT THA6HOMS1 GCAGGTCCG-GCAGCT--TGAGC
AGCAGCTTGGTGAAGCGGGAAGTCTCCAAGGGCCGGT THA7HOMS1
GCAGGTCCG-GCAGCT--TGAGCAGCAGCTTGGTGAAGCGGGAAGT
CTCCAAGGGCCGGT THA3HOMS1 GCAGGTCCG-GCAGCT--T
GAGCAGCAGCTTGGTGAAGCGGGAAGTCTCCAAGGGCCGGT THA2HOMS
1 GCAGGTCCG-GCAGCT--TGAGCAGCAGCTTGGTGAAGCGGG
AAGTCTCCAAGGGCCGGT THA1HOMS1
GCAGGTCCG-GCAGCT--TGAGCAGCAGCTTGGTGAAGCGGGAAGTCTCC
AAGGGCCGGT THA4HOMS1 GCAGGTCCG-GCAGCT--TGAGC
AGCAGCTTGGTGAAGCGGGAAGTCTCCAAGGGCCGGT THA0HOMS1
GAGGGTGTGCGGAGCTGGTGGGGAGGAGCCTGGAGAGAAGGGGCAG
AGCTGGGGGCTGAG

keyword "TRa" and species "homo sapiens" -gt 8
cDNAs
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Other databases of interest
Classification of species www.ncbi.nlm.nih.gov/Ta
xonomy/tax.html/ Gene interactions and
pathways www.genome.ad.jp/kegg/ Genome
ressources www.ensembl.org/ Human genetic
disorders www3.ncbi.nlm.nih.gov/Omim/ OMIM
(genetic diseases) www3.ncbi.nlm.nih.gov/Omim/ N
ematode genome and biology www.wormbase.org/ Droso
phila genome flybase.bio.indiana.edu/ Transcrip
tion factors (fly) www.fruitfly.org/cis-analyst/
Endocrine disruption e.hormone.tulane.edu/ Hormon
e levels www.il-st-acad-sci.org//data2.html Liga
nd binding gong.rockefeller.edu/ligbase/ Pharmac
ology and nomenclature www.iuphar.org/ Promoters
www.epd.isb-sib.ch/ Transcription
factors transfac.gbf.de/TRANSFAC/
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Conclusions

• nuclear receptors have conserved sequences
  allowing "easy" datamining
• before doing an experiment, check if the result
  is already available
• but keep in mind the limitations of
  bioinformatics
• specialized ressources exist

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Acknowledgements
lab of Vincent Laudet Stéphanie Bertrand, Jorge
Duarte, Tarik Ourjdal , Gilles Parmentier, Emma
Ribes, David Ruau, Raquel Tavares with Guy
Perrière and Simon Penel (PBIL) support from
programmes "Bioinformatique" CNRS (2000-2002) and
inter-EPST (2002-2004)