genome duplication and fish models for toxicology · • genome duplication in vertebrates...
TRANSCRIPT
DIVISION OF COMPARATIVE MEDICINE
~\. National Institute of Environmental Health Sciences liilllr'J' Your Environment. Your Health. ,'~ National Toxicology Program ~ U.S. Department of Health and Human Services
Genome duplication and fish models for toxicology
John Postlethwait University of Oregon
NIH ' DIVISION OF COMPARATIVE MEDICINE
··-·• ~\. Nationa_l Institute of En~ir~nmental Hea~~S · -lilllr'/ Your Environment. Your Health. ciences
Genome duplication and fish models for toxicology Frank von Hippel N AZ U
Loren Buck N AZ U
Tom Titus UO
Pete Batzel UO
Jesse Gologergen Sivuqaq Island
Tiffany Immingen Sivuqaq Island
Ingo Braasch
The problem: People living a subsistence lifestyle in the Arctic are highly exposed to persistent organic pollutants.
Annie Alowa, Sivuqaq Islander
' ~""' ,;
t@~ a 1 VA ,. . ... -). _ . ..··~i ·t. • . .A. · . . • ··.•~--... ~. .. ,•· ·••' ... . . 1 11 . ·•. . . ' . . •' '• • \ . . . . .
. 'II,_ !'· . • ·•. -~ ... • • .• : ,.
~''-~--.. ----. \-:l·· 4_· ..... . -:
····: ... : . ··. "\ .:· . : ·- . ·~ :- - ---
aqumaps.org
organochlorines evaporate
move to colder climates
condense & fall
Biological magnification
The problem: Formerly used defense sites
Greenland
Canada Norway
SLIGlobal distillation Point sources Sources:
' ' ~,v- ,;
t@.,- ~ : 1 VA ,. . ·. -:·J. . • !
... ... . . . .. ··~ : 1.. a·· •· .A . ·· •
• \ I •• •• :• .' • · ••' .:• • • • . . . ' . . .. , . . . . . •. . . . . . . . ~.... . •. ·• .. · .. . :.:, ~- \ ... . ' ... ;.-.~. ... I . :l ... ... • 4_· . -:· ~ _;· ~,. fyl ______ ..:,
aqumaps.org
Global distillation
Biological magnification
The problem: Formerly used defense sites
Greenland
Canada Norway
SLI Point sources
9-spine stickleback
Teleost fish are convenient models for toxicology.
How to connect teleost genomes to human biology?
• Genome duplication complicates connectivity • Genome duplication in vertebrates
How to connect teleost genomes to human biology?
• Ohnologs gone missing and lineage-specific evolution • Application: St. Lawrence Island
How to connect teleost genomes to human biology? • Genome duplication in vertebrates
zebrafish
bony fish Susumo Ohno
Evolution by Gene Two rounds of genome Duplication (1970)duplication
Ohnologs: duplicates from genome duplication
jawed vertebrates
1 Hox cluster
4 Hox clusterse shark cartilaginous fish
How to connect teleost genomes to human biology? zebrafish
e shark
bony fish
cartilaginous fish
jawed vertebrates
coelacanth
xenopus
chicken
mouse
human
• Genome duplication in vertebrates
rayfin fish
Two rounds of genome duplication
lobefin fish
1 Hox cluster
4 Hox clusters
rayfin fish
teleost fish
How to connect teleost genomes to human biology?
Two rounds of genome duplication
zebrafish
coelacanth
e shark
bony fish
cartilaginous fish
jawed vertebrates
gar
xenopus
chicken
mouse
human
medaka
cavefish
goldeye
eel
• Genome duplication in vertebrates
lobefin fish
1 Hox cluster
4 Hox clusters
How to connect teleost genomes to human biology?
4 Hox clusters
1 Hox cluster
human
4 Hox clusters
• Genome duplication in vertebrates
zebrafish
coelacanth
bony fish
Two rounds of genome duplication
jawed vertebrates
gar
xenopus
chicken
mouse
medaka
cavefish
goldeye
eel
Teleost genome duplication
1R, 2R VGD
3R, TGD
e shark cartilaginous fish
teleost fish
rayfin 7-8 Hox clusters Amores…
Postlethwait 1998 Science
fish
Amores…Postlethwait 2011 Genetics
lobefin fish
4 Hox clusters
teleost fish 7-8 Hox clusters
Spotted gar: • Diverged before the TGD • Biology similar to zebrafish
• Genome duplication in vertebrates
zebrafish
coelacanth
e shark 4 Hox clusters
bony fish
cartilaginous fish
1 Hox cluster
Two rounds of genome duplication
jawed vertebrates
gar
xenopus
chicken
mouse
human
medaka
cavefish
goldeye
eel
Teleost genome duplication
How to connect teleost genomes to human biology?
1R, 2R VGD
3R, TGD
How to connect teleost genomes to human biology?
4 Hox clusters What does duplication mean for toxicology?
4 Hox clusters
teleost fish 7-8 Hox clusters
Spotted gar: • Diverged before the TGD • Biology similar to zebrafish
• Genomics similar to tetrapods • Same duplication as human
• Amenable to laboratory culture • Can grow from embryos
• Genome duplication in vertebrates
zebrafish
coelacanth
e shark
bony fish
cartilaginous fish
1 Hox cluster
Two rounds of genome duplication
jawed vertebrates
gar
xenopus
chicken
mouse
human
medaka
cavefish
goldeye
eel
Teleost genome duplication
1R, 2R VGD
3R, TGD
What does duplication mean for toxicology?
How to connect teleost genomes to human biology?
1R, 2R VGD
Two rounds of genome duplication
chicken
mouse
human
jawed vertebrates
1 Hox cluster e shark cartilaginous fish
zebrafish
coelacanth
bony fish gar
xenopus
medaka
cavefish
goldeye
eel
Teleost genome duplication
• Genome duplication in vertebrates
3R, TGD
cytoplasm
nucleus
dioxin response element
dioxins
AHR
cyp1a1
ARNTAHR
Catchen…Postlethwait 2009 Genome Res. 19:1497
Arnt Arntl Tandem duplication
VGD1 Arnt Arntl
Arnt Arntl
VGD2 Arnt Arntl
Arnt2 Arntl2
TGD
arnt arntl1a
arntl1b
arnt2 arntl2zebrafish
human
cytoplasm
nucleus
dioxin response element
dioxins
AHR
cyp1a1
ARNTAHR
How do we know?
How to connect teleost genomes to human biology?
ARNT ARNTL
ARNT2 ARNTL2
What does duplication mean for toxicology?
100
,-----Hsa.ARNTI. ---Dre.amtl1a
---Dre.amtl1b r--------Hsa.ARNTL2
100 '--------Dre.amt12 ,__ ____ -Rfl .110703
,-----Hsa.ARNT2 ,...... __ --l
81
Dre.amt2
,--------Hsa.ARNT
~-----Dre.amt
How do we know?
gene tree How to connect teleost genomes to human biology?
arnt arntl1a
Catchen…Postlethwait 2009 Genome Res. 19:1497
Arnt Arntl Tandem duplication
VGD1 Arnt Arntl
Arnt Arntl
VGD2 Arnt Arntl
Arnt2 Arntl2
TGD zebrafish
human
Arnt4 Arntl4
Arnt3 Arntl3
ARNT ARNTL
ARNT2 ARNTL2
arnt2 arntl1b
arnt2 arntl2
arnt2b arntl2b
orthologs
100
,-----Hsa.ARNTI. ---Dre.amtl1a
---Dre.amtl1b r--------Hsa.ARNTL2
100 '--------Dre.amt12 ,__ ____ -Rfl .110703
,-----Hsa.ARNT2 ,...... __ --l
81
Dre.amt2
,--------Hsa.ARNT
~-----Dre.amt
How do we know?
gene tree How to connect teleost genomes to human biology?
TGD
arnt arntl1a
arntl1b
arnt2 arntl2
Catchen…Postlethwait 2009 Genome Res. 19:1497
Arnt Arntl Tandem duplication
VGD1 Arnt Arntl
Arnt Arntl
VGD2 Arnt Arntl
Arnt2 Arntl2
zebrafish
human
ARNT ARNTL
ARNT2 ARNTL2
orthologs
100
100
,-----Hsa.ARNTI. ---Dre.amtl1a
---Dre.amtl1b r--------Hsa.ARNTL2
'--------Dre.amt12 ,__ ____ -Rfl .110703
,-----Hsa.ARNT2 ,...... __ --l
81
Dre.amt2
,--------Hsa.ARNT
~-----Dre.amt
How do we know?
gene tree How to connect teleost genomes to human biology?
VGD1 Arnt Arntl
Arnt Arntl Arnt ArntlTandem duplication
TGD
arnt arntl1a
arntl1b
arnt2 arntl2
VGD2 Arnt Arntl
Arnt2 Arntl2
zebrafish
human
ARNT ARNTL
ARNT2 ARNTL2
orthologs
Catchen…Postlethwait 2009 Genome Res. 19:1497
100
,-----Hsa.ARNTI. ---Dre.amtl1a
---Dre.amtl1b
100 '--------Dre.amt12 ,__ ____ -Rfl .110703
,-----Hsa.ARNT2 ,...... __ --l
81
Dre.amt2
,--------Hsa.ARNT
~-----Dre.amt
human
TGD
arnt arntl1a
arntl1b
arnt2 arntl2zebrafish
ARNT ARNTL
ARNT2 ARNTL2
Arnt Arntl
VGD1 Arnt Arntl
Arnt Arntl
VGD2 Arnt Arntl
Arnt2 Arntl2
How do we know?
How to connect teleost genomes to human biology?
gene tree
Tree shows duplication after human-fish divergence.
co-orthologs
Tandem duplication
Catchen…Postlethwait 2009 Genome Res. 19:1497
ARNTLarntl1b
arntl1a
conserved syntenies
human-fish
show duplicated chromosome segments
How do we know?
How to connect teleost genomes to human biology? Tree shows duplication after
arnt arntl1adivergence.
arnt2 arntl1b
arnt2 arntl2 TGD
arnt2b arntl2b
zebrafish
VGD1 Arnt Arntl VGD2 Arnt Arntl
Arnt Arntl Arnt2 Arntl2
Arnt4 Arntl4
Arnt3 Arntl3
Arnt Arntl Tandem duplication ARNT ARNTL
human ARNT2 ARNTL2How do we know?
Catchen…Postlethwait 2009 Genome Res. 19:1497
♦
+ +-++t++ + +♦ + -· ++++ + ♦ + + ++ + ♦++ + +
+ + ·- -*• + + ♦ + ♦
+ * + + ♦ ++ ++
+ + + ... +
+ + + + ++-I+ ♦++ + + + ++ +
+ ++ ..... + + + ..... + ++
+ -++- ++++ * ++ ♦
+ ++ ++ + +
++ +
. +- _ __.._ ··- - -- -➔ -++- .. -+- ·+-·- ·- .... - - .,_.,... __ +-·
++ ♦ ♦ + +++ -· ** + + + +
+ - +
0 + + + HI-++ + ++
♦
+ + + + + + ++• + + + + + +
+ + + + + + + + - + + + + ............ +++++ + ·- .... + + + ... + ♦ ♦- +
How do we know? ARNT
How to connect teleost genomes to human biology?Human
chromosomes Duplicated ARNTL chromosome
segments in arnt arntl1aARNTL2 humanARNT2 genome arnt2 arntl1b
ARNT2 ARNTL2
arnt2 arntl2 Genes on human chromosome 11
Arnt Arntl Arnt2 Arntl2
Arnt4 Arntl4
Arnt3 Arntl3
TGD
arnt2b arntl2b
zebrafish
human
VGD1 Arnt Arntl VGD2 Arnt Arntl
Arnt Arntl Tandem duplication ARNT ARNTL
How do we know?
Catchen…Postlethwait 2009 Genome Res. 19:1497
♦
+ +-++t++ + +♦ + -· ++++ + ♦ + + ++ + ♦++ + +
+ + ·- -*• + + ♦ + ♦
+ * + + ♦ ++ ++
+ + + ... +
+ + + + ++-I+ ♦++ + + + ++ +
+ ++ ..... + + + ..... + ++
+ -++- ++++ * ++ ♦
+ ++ ++ + +
++ +
. +- _ __.._ ··- - -- -➔ -++- .. -+- ·+-·- ·- .... - - .,_.,... __ +-·
++ ♦ ♦ + +++ -· ** + + + +
+ - +
0 + + + HI-++ + ++
♦
+ + + + + + ++• + + + + + +
+ + + + + + + + - + + + + ............ +++++ + ·- .... + + + ... + ♦ ♦- +
How do we know? ARNT
How to connect teleost genomes to human biology?Human
chromosomes Duplicated ARNTL chromosome How do
ARNTL2 ARNT2
Genes on human chromosome 11
VGD1 Arnt Arntl VGD2 Arnt Arntl
Arnt Arntl Tandem duplication
How do we know?
Arnt Arntl Arnt2 Arntl2
Arnt4 Arntl4
Arnt3 Arntl3
Catchen…Postlethwait 2009 Genome Res. 19:1497
ARNT ARNTL
ARNT2 ARNTL2
TGD
arnt arntl1a
arnt2 arntl1b
arnt2 arntl2
arnt2b arntl2b
zebrafish
human
segments in human genome
• Genome duplication complicates connectivity
fish genes relate to human genes?
• Genome duplication complicates connectivity • Genome duplication in vertebrates
How to connect teleost genomes to human biology?
• Ohnologs gone missing and lineage-specific evolution • Application: St. Lawrence Island
Regulatory duplication
• Genome duplication complicates connectivity
Protein coding Functional domains
nonfunctionalization Gene loss: most frequent fate
Alan Force
Force…Postlethwait 1999 Genetics
• Genome duplication complicates connectivity
Protein coding Functional domains
Regulatory
nonfunctionalization neofunctionalization subfunctionalization
duplication
Gene loss: most New functions evolve Reciprocal loss of frequent fate from ancestral genes ancestral functions
Alan Force
Force…Postlethwait 1999 Genetics
•. -L
..... -• -L
• -L ••
•-/ .
--L ••
--L •• t ••
neofunctionalization New functions evolve from ancestral genes
nonfunctionalization Reciprocal loss of ancestral functions
Gene loss: most frequent fate
subfunctionalization
Protein coding
• Genome duplication complicates connectivity
duplication
Species Retained TGD pairs
Codingge n e s
%retained
Zebrafish 2,228 26,459 8.4
Stickleback 2,156 20,787 10.4
Medaka 1,910 19,699 9.7
Te t r a od o n 1,853 19,602 9.5
Functional domains
Most teleost genes are 1:1 orthologs with human
Alan ForceWe don’t yet know the relative importance of these fates.
Force…Postlethwait 1999 Genetics
• Genome duplication complicates connectivity Species Retained TGD Coding %retained
pairs genes Zebrafish 2,228 26,459 8.4
Stickleback 2,156 20,787 10.4
Medaka 1,910 19,699 9.7
Tetraodon 1,853 19,602 9.5
What does this mean for toxicology?
Problem: All can occur in a lineage-specific fashion.
nonfunctionalization neofunctionalization subfunctionalization Gene loss: most New functions evolve Reciprocal loss of frequent fate from ancestral genes ancestral functions
Alan ForceWe don’t yet know the relative
Forceimportance of these fates.
…Postlethwait 1999 Genetics
What does this mean for toxicology?
cytoplasm
nucleus
dioxin response element
dioxins
AHR
cyp1a1
ARNTAHRAHR
promotes expression of oxygen-responsive genes.
What about AHR?
Amphioxus
neofunctionalization subfunctionalization
To see if this is subfunctionalization or neofunctionalization, must look at a pre-duplication outgroup.
nonfunctionalization neofunctionalization subfunctionalization
heterodimer with ligand- heterodimer with CLOCK, bound AHR, promotes promotes expression of expression of xenobiotic circadian rhythm genes metabolism genes
ARNT ARNTL
ARNT2 ARNTL2 heterodimer with hypoxia- heterodimer with inducible factor 1alpha, CLOCK & HIF1alpha
Ascidians Appendicularians http://marineinvaders.myspecies.info/taxonomy/term/70 evodevogenomics-unibarcelona.weebly.com/oikopleura www2.le.ac.uk/departments/geology/people/goodall-r/personal/images/lancelet.
Dre16
Gga2
Dre22
Gga7
What does this mean for toxicology?
nonfunctionalization subfunctionalizationneofunctionalization
cytoplasm
nucleus
dioxin response element
dioxins AHR
cyp1a1
AHR ARNT
What about AHR?
AHR
Dre16 Dre22
one AHR gene
three AHR-related genes
three AHR-related genes
Conserved syntenies show that chicken and zebrafish AHR-related genes are orthologs
How did AHR-related genes evolve?Karchner 2005 Biochem J. 392:153
VGD2 ahr2 ahr1b
ahr1a ahr1b’ahr ahr2 ahr1
ahr2 ahr1b
ahr1a ahr1b’
AHR2 AHR1B
AHR ahr1b’
ahr1b’AHR
ahr1bahr1a
How did AHR-related genes evolve?
cytoplasm
nucleus
dioxin response element
dioxins AHR
cyp1a1
AHR ARNT
TGD zebrafish
human
chicken
nonfunctionalization subfunctionalizationneofunctionalization
Zebrafish three AH Rs • AHR2 primary mediator of toxicity ~ - =----• AHRl A deficient in TCDD binding and • transactivation activity • AHRl B functiona l but no known toxicological
roles
How did AHR-related genes evolve?
cytoplasm
nucleus
dioxin response element
dioxins AHR
cyp1a1
AHR ARNT
Robert Tanguay Oregon State University 2018 10 25
nonfunctionalization subfunctionalizationneofunctionalization
VGD2 ahr2 ahr1b
ahr1a ahr ahr2 ahr1
ahr2 ahr1b
ahr1a
AHR2 AHR1B
AHR
AHR
TGD zebrafish
human
chicken
The ‘primary mediator of toxicity’ for teleosts is not the ortholog of human AHR
Zebrafish ortholog of human AHR is an ‘incipient pseudogene’. Karchner 2005 Biochem J. 392:153
Zebrafish three AH Rs • AHR2 primary mediator of toxicity • AHRl A deficient in TCDD binding and • transactivation activity • AHRl B functiona l but no known toxicological
roles
How did AHR-related genes evolve?
cytoplasm
nucleus
dioxin response element
dioxins AHR
cyp1a1
AHR ARNT
Robert Tanguay Oregon State University 2018 10 25
nonfunctionalization subfunctionalizationneofunctionalization
VGD2 ahr2 ahr1b
ahr1a ahr1b’ahr ahr2 ahr1
ahr2 ahr1b
ahr1a ahr1b’
AHR2 AHR1B
AHR ahr1b’
ahr1b’AHR
ahr1bahr1a
TGD zebrafish
human
chicken
Study of ahr1b may reveal sub-functions of human AHR gene
subfunctionalization
VGD2 ahr2 ahr1b
ahr1a ahr1b’ahr ahr2 ahr1
ahr2 ahr1b
ahr1a ahr1b’
AHR2 AHR1B
AHR ahr1b’
ahr1b’AHR
ahr1bahr1a
How did AHR-related genes evolve?
cytoplasm
nucleus
dioxin response element
dioxins AHR
cyp1a1
AHR ARNT
TGD zebrafish
human
chicken
nonfunctionalization subfunctionalizationneofunctionalization
Postlethwait 2007 J Exp Zool B Mol Dev Evol. 308:563
• Ohnologs gone missing and lineage-specific evolution
• Genome duplication complicates connectivity • Genome duplication in vertebrates
How to connect teleost genomes to human biology?
• Ohnologs gone missing and lineage-specific evolution • Application: St. Lawrence Island
St. Lawrence Island AlaskaRussia
• Application: St. Lawrence Island
Savunga
St. Lawrence Island
Northeast Cape
Alaska Russia
air strip
Main operations of FUD
Suqitughneq River
12 10
14
11 13
8 7
6
5 4
2
3 • Application: St. Lawrence Island
1985 - 2014, $120M spent on remediation
air strip
Main operations of FUD
Suqitughneq River
12 10
14
11 13
8 7
6
5 4
2
3 • Application: St. Lawrence Island
1985 - 2014, $120M spent on remediation
Do upstream and downstream fish differ?
w en
+f 0 0) 0 0) ,-
.s ~ IIl 0 e:. ro § C: 0 ~ lO
::E
0
2 Estuary
i i 3 4 5 0 7 8
Site number
=> significantly more PCBs in downstream fish
Alaska blackfish
12 10
14
11 13
8 7
6
5 4
2
3
Are PCBs from global distillation or the FUD?
Do upstream and downstream fish differ?
von Hippel 2018 Environmental Pollution 234:279
0000 00 00 00 0
0 0 0 000 00000000 0 0000000000000 000000000 0 000000000 000 0 0 0000 0000000 000000000000 0 0000 0000
Are PCBs from global distillation or the FUD? 3
Greater => Upstream from global distillationupstream => Downstream PCBs from FUD + distillation. Greater light heavydown stream
6
7 8 12
10 11 13
14 Main operations of FUD
Danielle Dillon What are the biological effects?
What are the biological effects? sex genotype PCR Gene expression differences?
4 females from highly contaminated sites 4 females from less contaminated sites
BIOO NEXTflex qRNA-seq
Strand-specific: guards against gDNA
Unique molecular indexes: guards against PCR duplicates
Tom Titus UO
■ --
brca2 paired-end reads Aligned 9-spine reads to 3-spine genome
3-spine genome 3’UTR unannotated
polymorphisms
9-spine PCBs low strand specific
wrong way => genomic contamination
9-spine PCBs high
Pete Batzel
■ --I I
3-spine genome
brca2
DESeq2: Finds differentially expressed genes
paired-end reads
Pete Batzel
Aligned 9-spine reads to 3-spine genome
9-spine PCBs low
9-spine PCBs high
compared to
Differentially expressed genes Females: contamin/uncontam 2312 Males: contamin/uncontam 642
What do these genes do?
DAVID 6.7: Database for Annotation, Visualization and Integrated Discovery
DESeq2: Finds differentially expressed genes
Annotation Cluslllr1 Enrichment Sc:otr.15.17 k, Cowit P _Value BenJamlnl
UP _KEYWORDS Cell cycle RT - 135 1.9E-20 S.2E-l8
UP _KEYWORDS Cell division 9 1 1.2E-17 l.7E-l5
UP _KEYWORDS Mitosis ..... - 69 3.7E-16 3.0E-14
GOTERM_BP _DIRECT cell division RT - 83 1. l E-13 2.6E-10
What do these genes do? DAVID 6.7: brca2 (fancd1) = down cdc23 = down fbxo5 = down cell cycle genes h2ax = down haus6 = down mad2l1 = down rab11fip4a = down llgl2 = down ccnb1 = down most tgf betas down
Annotation Cluslllr1 Enrichment Sc:otr.15.17 k, Cowit P _Value BenJamlnl
UP _KEYWORDS Cell cycle RT - 135 1.9E-20 S.2E-l8
UP _KEYWORDS Cell division RT - 9 1 1.2E-17 l.7E-l5
UP _KEYWORDS Mitosis RT - 69 3.7E-16 3.0E-14
GOTERM_BP _DIRECT cell division RT - 83 1. l E-13 2.6E-10
GOTERM_BP _DIRECT mitotic nuclear division RT • 63 1.SE-11 2.3E-8
Annotation Clusta, 2 Enrichment Scorw: 11.1, Ill,, Cowit P _Value B■nJ■mlnl
UP _KEYWORDS
GOTERM_BP _DIRECT
- - --~-------- ii
ii
'UI 1 ""'-'" Q RE-14
E-8
What do these genes do? brca2 (fancd1) = down cdc23 = down fbxo5 = down h2ax = down haus6 = down
DNA replication genes
mcm3,4,6,7 cdc45 hsc70 pcna rbbp4 tert terf1, terf2
mad2l1 = down rab11fip4a = down llgl2 = down ccnb1 = down most tgf betas down
Annotation Cluslllr1 Enrichment Sc:otr.15.17 k, Cowit P _Value BenJamlnl
UP _KEYWORDS Cell cycle RT - 135 1.9E-20 5.2E-l8
UP _KEYWORDS Cell division RT - 91 1.2E-17 l.7E-l5
UP _KEYWORDS Mitosis RT - 69 3.7E-16 3.0E-14
GOTERM_BP _DIRECT cell division RT - 83 1. lE-13 2.6E-10
GOTERM_BP _DIRECT mitotic nuclear division RT • 63 1.5E-11 2.3E-8
Annotation Clusta, 2 Enrichment Scorw: 11.1, Ill,, Cowit P _Value B■nJ■mlnl
UP _KEYWORDS
GOTERM_BP _DIRECT
GOTERM_BP _DIRECT
D A replication
Gl/5 transit ion of mitot ic cell cycle
D A replication ini iation
ii
ii
i
38
36
19
1.5E-15 9.8E-l4
3.lE-11 3.7E-8
1. 3E-10 l.2E-7
Annotation Cluslllr 3 Enrichment Score: 8.IM k, Cowit . P _ Value BenJ■mlnl
UP _KEYWORDS
UP _KEYWORDS
AT 67
58
4 . 4E-9 l.8E-7
1 .8E-8 5.4E-7
What do these genes do? brca2 (fancd1) = down cdc23 = down fbxo5 = down h2ax = down haus6 = down mad2l1 = down rab11fip4a = down llgl2 = down ccnb1 = down most tgf betas down
DNA repair genes mcm3,4,6,7 cdc45 hsc70 pcna rbbp4 tert terf1, terf2
Fanconi anemia genes = down
Annotation Cluslllr1 Enrichment Sc:otr.15.17 k, Cowit P _Value BenJamlnl
UP _KEYWORDS Cell cycle RT - 135 1.9E-20 5.2E-l8
UP _KEYWORDS Cell division RT - 9 1 1.2E-17 l.7E-l5
UP _KEYWORDS Mitosis RT - 69 3.7E-16 3.0E-14
GOTERM_BP _DIRECT cell division RT - 83 1. l E-13 2.6E-10
GOTERM_BP _DIRECT mitotic nuclear division RT • 63 1.5E-11 2.3E-8
Annotation Clusta, 2 Enrichment Scorw: 11.1, Ill,, Cowit P _Value B■nJ■mlnl
UP _KEYWORDS
GOTERM_BP _DIRECT
GOTERM_BP _DIRECT
D A replication
Gl/5 transition of mitotic cell cycle
D A replication ini iation
ii
ii
i
38
36
19
1.5E-15 9.8E-l4
3. l E-11 3.7E-8
1. 3E-10 l.2E-7
Annotation Cluslllr 3 Enrichment Score: 8.IM k, Cowit . P _ Value BenJ■mlnl
UP _KEYWORDS
UP _KEYWORDS
GOTERM_BP _DIRECT
D Adamaqe
D A repair
D A repair
• • •
67
58
46
4 . 4E-9 l.8E-7
1 .8E-8 5.4E-7
1 .9E-5 5.7E-3
Annotation Cluster• Enrichment Scan: 8.39 Ila. Cowit P _ Yalu■ BenJ■mlnl
UP _KEYWORDS Centromere RT ii 36 1.0E-8 3.6E-7
UP _KEYWORDS Chromosome 8 3.7E-7
GOTERM_BP _DIRECT sister chromatid cohesion 8 6.9E-6
UP _KEYWORDS Kinetochore 7 2.8E-6
GOTERM_CC_DIRECT kinetochore RT ii 20 1.7E-4 9.4E-3
What do these genes do? brca2 (fancd1) = down cdc23 = down fbxo5 = down h2ax = down haus6 = down mad2l1 = down rab11fip4a = down llgl2 = down ccnb1 = down most tgf betas down
mcm3,4,6,7 cdc45 hsc70 pcna rbbp4 tert terf1, terf2
Fanconi anemia genes = down ercc4 msh6 pola1, pola2 prim1
What about duplicated genes?
chromosome behavior genes
Gene baseMean log2FC pvalue padj
camklga 27.345 -2.275 0.003 0.037
camklgb 60.386 1.587 0.010
fabpl0a 5935.662 2.457 0.000
fabpl0b 32.048 -1.766 0.001
fam65a 91.364 1.367 0.002
fam65b 64.561 -1.156 0.009
kcnj2a 97.195 -2.508 0.001
kcn '2b 18.028 ,-.~-Ji .r'_ ~-- --"'."-·- ,. '
. . . .. " "" .' ,;. . ........ . .. " ' ..,._,. ' -
!►....;~. ·_ ' . ._.. -_ -·
Water at North East Cape is toxic to stickleback genome.
What about duplicated genes?
246 TGD gene pairs had at least one copy that was differentially expressed. 21 pairs had both copies differentially expressed.
4 of those 21 pairs had one copy up, other copy down in contaminated fish.
nonfunctionalization subfunctionalizationneofunctionalization
225 TGD pairs had only one copy DE. Study of how they differ with toxicants may give insight into mechanisms.
neofunctionalization subfunctionalization TGD pairs provide that important advantage for analysis of toxicity.
DNA repair pathways: low expression in contaminated sites.
Estrogenic effect on males.
DNA replication: low in contaminated sites. Cell cycle genes: low expression in contaminated sites.
Water at North East Cape is toxic to stickleback genome.
• contaminants persist, • accumulate in fish, • alter endocrine pathways, • alter expression of genome stability genes.
Even after site remediation:
Health of indigenous people is put at risk.
• Genome duplication complicates connectivity • Genome duplication in vertebrates
How to connect teleost genomes to human biology?
• Ohnologs gone missing and lineage-specific evolution • Application: St. Lawrence Island
NIH ' DIVISION OF COMPARATIVE MEDICINE
··-·• ~\. Nationa_l Institute of En~ir~nmental Hea~~S · -lilllr'/ Your Environment. Your Health. ciences
Genome duplication and fish models for toxicology Frank von Hippel N AZ U
Loren Buck N AZ U
Tom Titus UO
Pete Batzel UO
Jesse Gologergen Sivuqaq Island
Tiffany Immingen Sivuqaq Island
Ingo Braasch
What are the biological effects?
[vitellogenin] in males (µg/g)
600
400
800
200
0
Suqi River
Tapi River
FUD
Ninespine stickleback
sex genotype PCR
=> estrogenic effect on Suqi males
Sq2 Sq4 Sq5 Sq6 Sq7 Sq11 Sq13 Tapi
What is the genetic response?
: . Mlfouplolo Ind Placental mommolo: 3 homolog1
,-., AHR2. Chicken ENSMGAG00000002128, Turkey
ENSAPLG00000003921 , Duck ENSFALG00000003870, Flycalcher
ENSOANG000000135-48, Pllltypul
Eu1eleol1eomorpha: 42 homolog1
ENSPNAG00000005817, Recl,belllod piranha ENSAMXG00000043051 , Cave fllh
ENSIPUG00000002619, Channel Clttllh ENSPNAG00000015192. Recl,bellled piranha
OOleoglonttonnH: 4 homolog1
EutlleolleOmOrphl: 44 homologa
1hr2, Red-bellied pln1nh1 ahr2, Cove lllh
■hr2. Channel catfish lhr2. Zebrllflah
Teleoat fl111H : 8 homologo ENSPNAG00000003288, Recl,belllod piranha
ahr2, Spotted gar ENSLACG00000018685, Coel1<1nlh
Cheroclphy-: 2 homologl ~ ~--- lhr1a. Zebr1fl1h
lhr11, Northern plkl Oateoglo11tt0nnff: 2 homolog1
lhr1 I. Spotted gar Placental mammala: 14 homotog1
r Homlnlnes: 3 homolog1 AC019117.2, Humon AHR, Hurrn,n f ENSPPYG0000001TT58, orangutan
ENSNLEG00000016550, Gibbon Old World monkeyl: 10 homolog1 New Wor1d monkeys: 4 homologI
0 ENSTSYG00000010830, Tarsler Wet noae lemura: 3 homologa
ENST8EG00000007210, Tree Shrew Rodents: 9 homolog1
Leurul1therl1n m1mma11: HS h0mok>g1 Placental■ : 5 homologa
Marauplala: 3 homok>ga ENSOANG00000009e11 . Ptatypu1
ENSACAG00000023372, Anole lizard e AHR, Chicken
• • , ENSMGAG00000010174, Tur1cey • S ENSAPLG00000008789. Duck L. Perching birds: 2 homologa
ENSPSIG00000010051 , Chlneoe eottohell lurtle ahr, Xenopua
.... ahr1 ■, Coelecanth Lamprey: 3 homologa
ver1e1>r,1eo: 5 homolog1 Perching blrdo : 2 homologo
ENSMGAG00000002118, Tur1cey AHR1B, Chicken
ENSPSIG00000018138, ChlneH aott1hell lurtle r ENSMODG00000027484, Opo11um • ENSOCUG00000028055, Rabbit
Ohr! b, Coelocanth
EuleleOateornO<ph■ : 33 homolog1
lhr1 b, Red-bellied pln1nha lhrl b, Cave fllh
lhr1b, Channel c■ttloh ahr1 b, Zebmllh
• Aatan bonytongue: 2 homolog1
Euleleoateomorphl: 45 homologo
ENSPNAG0000000528 I , Recl,bellled piranha ENSAMXG00000035088, Cava floh
■hr! b, P■romormyropo klng1ieyN ENSLBEG000000111998, B■ ltan wra11e
What does this mean for toxicology? What about AHR?
nonfunctionalization neofunctionalization subfunctionalization
cytoplasm
nucleus
dioxin response element
dioxins AHR
cyp1a1
AHR ARNTAHR
Karchner 2005 Biochem J. 392:153
: . Mlfouplolo Ind Placental mommolo: 3 homolog1
,-., AHR2. Chicken
ENSA~~~~ri.,1!:' Turkey
ENSFALG00000003370, Flye«1.:'1er ENSOANG000000135-48, Pllltypul
Eu1eleol1eomorpha: 42 homologo
ENSPNAG00000005817, Recl,belllod piranha ENSAMXG00000043051 , Cave fllh
ENSIPUG00000002619, Channel cattl1h ENSPNAG00000015192, Recl,bellled piranha
001eoglonttormH: 4 homolog1
EU11leol1e0m0rphl: 44 homologa
1hr2, Red-bellied pln1nh1 ahr2, Cove lllh
■hr2. Channel catfish lhr2. Zebrllflah
Teleoat fl111H : 8 homologo ENSPNAG00000003289, Recl,t,elllod piranha
ahr2, Spotted gar ENSLACG00000018685, Coelacan1h
Cheroclphy-: 2 homologa ~ ~--- lhr1a. Zebr1fl1h
lhr11, Nor1hem pike Oateoglo11ttOrmH: 2 hornololl-'
lhr1 a, Spotted gar Placental mammala: 14 homotog1
r Homlnlnes: 3 homolog1 AC019117.2, Humon AHR, Hu'""n f ENSPPYG0000001TT58, Ofangu1an
ENSNLEG00000018550, Gibbon Old World monkeyl: 10 homolog1 New Wor1d monkeys: 4 homolog1
0 ENSTSYG00000010830, Taroler Wet noae lemura: 3 homologa
ENST8EG00000007210, Tree Shrew Rodents: 9 homolog1
Leurul1therl1n m1mma11: HS h0mok>g1 Placental■ : 5 homologa
Marauplala: 3 homok>ga ENSOANG00000009e11 , PlatypUI
ENSACAG00000023372, Anole lizard e AHR, Chicken
• • , ENSMGAGOOOOOOI0174, Tur1cey • S ENSAPLG00000008789. Duck L. Perching birds: 2 homologa
ENSPSIG00000010051 , Chlneoe eottohell turtle ahr, Xenopua
.... ahr1 ■, Coelecanth Lamprey: 3 homologa
ver1e1>r,1eo: 5 homolog1 Perching blrdo: 2 hornologo
ENSMGAG00000002118, Tur1cey AHR1B, Chicken
ENSPSIG00000018136, C:~!oeH aottahetll tun.Ml r ENSMODG000000274&ii, Opo11um • ENSOCUG0000..'!128055, Rabbit
Ohr1 b, Coelocanth
EuteleOateornorpha: 33 homolog1
lhr1 b, Red-bellied pln1nha lhrl b, Cave fllh
ahr1b, Channel cattloh ahr1 b, Zebralllh
• Aalan bonytongue: 2 homolog1
Euteleoateomorph1: 45 homologo
ENSPNAG0000000528 I , Recl,bellled piranha ENSAMXG00000035088, Cava floh
ahr1 b, Pan1mormyropo klng1ieyN ENSLBEG000000111998, Ballan wraaae
What does this mean for toxicology?
nonfunctionalization subfunctionalizationneofunctionalization
cytoplasm
nucleus
dioxin response element
dioxins AHR
cyp1a1
AHR ARNT
zebrafish and chicken have ahr2
zebrafish ahr1a appears to be ortholog of human AHR
zebrafish and chicken have ahr1b
Karchner 2005 Biochem J. 392:153