Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas
-
Upload
mgavery -
Category
Technology
-
view
937 -
download
1
description
Transcript of Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas
![Page 1: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/1.jpg)
Mackenzie Gavery & Steven Roberts
University of Washington
School of Aquatic and Fishery Sciences
Seattle, WA USA
Exploring the role of DNA methylation as a source of phenotypic variation in
Crassostrea gigas
![Page 2: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/2.jpg)
Background: oysters
Results: characterization of DNA methylation in Crassostrea gigas
Discussion: functional role
Outline
![Page 3: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/3.jpg)
Oyster: Biology
egg sperm
free swimming larvae (2 weeks)
2-3 years
juvenile
![Page 4: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/4.jpg)
Oysters: Economic value
(FAO Fishery Statistics, 2006)
Major Producers of Crassostrea gigas
![Page 5: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/5.jpg)
Oysters: Threats
![Page 6: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/6.jpg)
DNA methylationAn epigenetic mechanism found in plants and
animals
In animals: occurs primarily in a CpG context
Function: gene regulation
Can be affected by environmental factors
Me
C
GC
G
![Page 7: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/7.jpg)
DNA methylation: invertebratesOnly a handful of species have been evaluated
Model invertebrates lack DNA methylation
Most: 30 – 60 % methylation
Primarily in exonic regions
Important regulatory functions – honey bee (e.g. Kucharski et al., 2008; Elango et al., 2009; Lyko et al., 2010)
![Page 8: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/8.jpg)
Characterization of DNA methylation in oysters
Describe distribution of methylation
Elucidate functional significance
![Page 9: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/9.jpg)
Results
in silico analysis
Genome wide methylation analysis
![Page 10: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/10.jpg)
in silico approachPrinciple:
Methylated cytosines are highly mutable
C T
Methylated regions of DNA are depleted of CpG dinucleotides over evolutionary time (CpG to TpG)
m
CpG observed
CpG expectedCpG O/E low = methylated
![Page 11: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/11.jpg)
in silico approachPrinciple:
Methylated cytosines are highly mutable
C T
Methylated regions of DNA are depleted of CpG dinucleotides over evolutionary time (CpG to TpG)
m
CpG observed
CpG expectedCpG O/E high = unmethylated
![Page 12: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/12.jpg)
DNA metabolism
cell cycle and proliferation
RNA metabolism
protein metabolism
death
other metabolic processes
cell organization and biogenesis
other biological processes
transport
stress response
developmental processes
cell-cell signaling
signal transduction
cell adhesion
0.45 0.50 0.55 0.60 0.65 0.70CpGO/E
Results: in silico
Gavery & Roberts, 2010
![Page 13: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/13.jpg)
DNA metabolism
cell cycle and proliferation
RNA metabolism
protein metabolism
death
other metabolic processes
cell organization and biogenesis
other biological processes
transport
stress response
developmental processes
cell-cell signaling
signal transduction
cell adhesion
0.45 0.50 0.55 0.60 0.65 0.70CpGO/E
Results: in silico‘h
ouse
keep
ing’
‘indu
cibl
e’
Gavery & Roberts, 2010
=methylated CpG
![Page 14: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/14.jpg)
Summary of Results:Genes with differing regulatory requirements have
different levels of DNA methylation
‘housekeeping’, ubiquitously expressed = methylated
‘inducible genes’ = unmethylated
![Page 15: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/15.jpg)
Results
in silico analysis
Genome wide methylation analysis
![Page 16: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/16.jpg)
MBD-seq
YY
YMBD
MBD
MBD
Methyl-binding domain isolated - genome sequencing
![Page 17: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/17.jpg)
CpG O/E(modified from Gavery and Roberts 2010)
Predicted degree of DNA methylation
MBD-seq: Results
![Page 18: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/18.jpg)
CpG O/E(modified from Gavery and Roberts 2010)
Predicted degree of DNA methylation
Mea
sure
d de
gree
of D
NA
met
hyla
tion
En
rich
men
t le
vel
in M
BD
lib
rary
(unp
ublis
hed)
MBD-seq: Results
![Page 19: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/19.jpg)
CpG O/E(modified from Gavery and Roberts 2010)
Predicted degree of DNA methylation
Mea
sure
d de
gree
of D
NA
met
hyla
tion
En
rich
men
t le
vel
in M
BD
lib
rary
(unp
ublis
hed)
MBD-seq: Results
![Page 20: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/20.jpg)
CpG O/E(modified from Gavery and Roberts 2010)
Predicted degree of DNA methylation
Mea
sure
d de
gree
of D
NA
met
hyla
tion
En
rich
men
t le
vel
in M
BD
lib
rary
(unp
ublis
hed)
MBD-seq: Results
![Page 21: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/21.jpg)
Summary of Results:Experimental analysis confirms in silico results
Genes with differing regulatory requirements have different levels of DNA methylation
‘housekeeping’ ubiquitously expressed = methylated
‘inducible genes’ = unmethylated
![Page 22: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/22.jpg)
Discussion:
Why?
![Page 23: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/23.jpg)
DNA metabolism
cell cycle and proliferation
RNA metabolism
protein metabolism
death
other metabolic processes
cell organization and biogenesis
other biological processes
transport
stress response
developmental processes
cell-cell signaling
signal transduction
cell adhesion
0.45 0.50 0.55 0.60 0.65 0.70CpGO/E
‘hou
seke
epin
g’‘in
duci
ble’
Gavery & Roberts, 2010
Discussion: Functional Role
![Page 24: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/24.jpg)
DNA metabolism
cell cycle and proliferation
RNA metabolism
protein metabolism
death
other metabolic processes
cell organization and biogenesis
other biological processes
transport
stress response
developmental processes
cell-cell signaling
signal transduction
cell adhesion
0.45 0.50 0.55 0.60 0.65 0.70CpGO/E
‘hou
seke
epin
g’‘in
duci
ble’
Gavery & Roberts, 2010
Conventional transcription of genesrequired for essential functioning
Discussion: Functional Role
![Page 25: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/25.jpg)
DNA metabolism
cell cycle and proliferation
RNA metabolism
protein metabolism
death
other metabolic processes
cell organization and biogenesis
other biological processes
transport
stress response
developmental processes
cell-cell signaling
signal transduction
cell adhesion
0.45 0.50 0.55 0.60 0.65 0.70CpGO/E
‘hou
seke
epin
g’‘in
duci
ble’
Gavery & Roberts, 2010
Discussion: Functional Role
Increased variation in environmental response genes
![Page 26: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/26.jpg)
DNA metabolism
cell cycle and proliferation
RNA metabolism
protein metabolism
death
other metabolic processes
cell organization and biogenesis
other biological processes
transport
stress response
developmental processes
cell-cell signaling
signal transduction
cell adhesion
0.45 0.50 0.55 0.60 0.65 0.70CpGO/E
‘hou
seke
epin
g’‘in
duci
ble’
Gavery & Roberts, 2010
Discussion: Functional Role
TFTFIncreased variation in environmental response genes
a) alternative splicing
![Page 27: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/27.jpg)
DNA metabolism
cell cycle and proliferation
RNA metabolism
protein metabolism
death
other metabolic processes
cell organization and biogenesis
other biological processes
transport
stress response
developmental processes
cell-cell signaling
signal transduction
cell adhesion
0.45 0.50 0.55 0.60 0.65 0.70CpGO/E
‘hou
seke
epin
g’‘in
duci
ble’
Gavery & Roberts, 2010
Discussion: Functional Role
Increased variation in environmental response genes
a) alternative splicingb) sequence variation
![Page 28: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/28.jpg)
DNA metabolism
cell cycle and proliferation
RNA metabolism
protein metabolism
death
other metabolic processes
cell organization and biogenesis
other biological processes
transport
stress response
developmental processes
cell-cell signaling
signal transduction
cell adhesion
0.45 0.50 0.55 0.60 0.65 0.70CpGO/E
‘hou
seke
epin
g’Discussion: Functional Role
Gavery & Roberts, 2010
Increased variation in environmental response genes
a) alternative splicingb) sequence variationc) transient methylation
‘indu
cibl
e’
![Page 29: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/29.jpg)
DNA metabolism
cell cycle and proliferation
RNA metabolism
protein metabolism
death
other metabolic processes
cell organization and biogenesis
other biological processes
transport
stress response
developmental processes
cell-cell signaling
signal transduction
cell adhesion
0.45 0.50 0.55 0.60 0.65 0.70CpGO/E
‘hou
seke
epin
g’Discussion: Functional Role
Gavery & Roberts, 2010
Increased variation in environmental response genes
a) alternative splicingb) sequence variationc) transient methylation
‘indu
cibl
e’
Conventional transcription of genesrequired for essential functioning
![Page 30: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/30.jpg)
The distribution of DNA methylation may function to promote variation in environmental response genes Planktonic larvae
Sessile
Variable environments
Discussion: Functional Role
![Page 31: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/31.jpg)
Conclusions/Future Directions:
Oysters have a functioning DNA methylation system
Ubiquitously expressed and inducible genes have different levels of methylation – indicating a functional roleFuture work:
Test the hypothesis that the DNA methylation system functions to enhance random variation in aquatic invertebrates
Investigate epigenetic effects of synthetic estrogens in oysters
![Page 32: Exploring the role of DNA methylation as a source of phenotypic variation in Crassostrea gigas](https://reader034.fdocuments.in/reader034/viewer/2022052619/5565a849d8b42a083a8b4d14/html5/thumbnails/32.jpg)
Acknowledgements
Samuel White (UW, SAFS)
Joth Davis (Taylor Shellfish Farms)
US Environmental Protection Agency
Graduate School, UW
email: [email protected]: students.washington.edu/mgavery