A 1-hour lecture to 4th-year undergraduate and/or MSc students in human genetics, focusing on exploring the role of DNA methylation in human complex disease.
Dna methylation ppt
definition of Dna methylation ppt
discovery of Dna methylation ppt
types of Dna methylation ppt
history of Dna methylation ppt
process of Dna methylation ppt
mechanism of Dna methylation ppt
methylation in cancer
cytosine methylation
genomic imprinting
Accurate DNA Methylation Analysis with Successful Bisulfite Conversion WebinarQIAGEN
Bisulfite conversion is a popular used method for DNA methylation analysis. It is the most convenient and effective way to map DNA methylation to individual bases. The efficiency of the bisulfite conversion has a huge impact on the reliability of the downstream analysis methods and complete conversion is a prerequisite for correct determination of methylation. However, standard methods require harsh conditions with long incubations times at high temperatures and low pH to achieve complete conversion. This harsh treatment can lead to DNA degradation, lowering the yields and sensitivity of the subsequent analysis.
This slidedeck:
• Explains the principle of bisulfite conversion
• Points out the challenges and critical factors for successful bisulfite conversion
• Describes how to overcome the challenges with QIAGEN’s EpiTect Fast Bisulfite Kits
• Gives general recommendations for planning successful bisulfite conversion experiments
Dna methylation ppt
definition of Dna methylation ppt
discovery of Dna methylation ppt
types of Dna methylation ppt
history of Dna methylation ppt
process of Dna methylation ppt
mechanism of Dna methylation ppt
methylation in cancer
cytosine methylation
genomic imprinting
Accurate DNA Methylation Analysis with Successful Bisulfite Conversion WebinarQIAGEN
Bisulfite conversion is a popular used method for DNA methylation analysis. It is the most convenient and effective way to map DNA methylation to individual bases. The efficiency of the bisulfite conversion has a huge impact on the reliability of the downstream analysis methods and complete conversion is a prerequisite for correct determination of methylation. However, standard methods require harsh conditions with long incubations times at high temperatures and low pH to achieve complete conversion. This harsh treatment can lead to DNA degradation, lowering the yields and sensitivity of the subsequent analysis.
This slidedeck:
• Explains the principle of bisulfite conversion
• Points out the challenges and critical factors for successful bisulfite conversion
• Describes how to overcome the challenges with QIAGEN’s EpiTect Fast Bisulfite Kits
• Gives general recommendations for planning successful bisulfite conversion experiments
Epigenetics definition, history of epigenetics, molecular basis of epigenetics, epigenetic modification, tools to study epigenetics, disease linked with epigenetics, DNA methylation demethylation and enzymes regulating DNA methylation
regulation of gene expression in eukaryotes is a complex mechanism involved many factors. out of many levels of regulations, chromosomal and transcription level of regulation are discussed in this slides.
The epigenetic regulation of DNA-templated processes has been intensely studied over the last 15
years. DNA methylation, histone modification, nucleosome remodeling, and RNA-mediated targeting regulate many biological processes that are fundamental to the genesis of cancer. Here, we
present the basic principles behind these epigenetic pathways and highlight the evidence suggesting that their misregulation can culminate in cancer. This information, along with the promising clinical and preclinical results seen with epigenetic drugs against chromatin regulators, signifies that it
is time to embrace the central role of epigenetics in cancer.
DNA SEQUENCING METHODS AND STRATEGIES FOR GENOME SEQUENCINGPuneet Kulyana
This presentation will give you a brief idea about the various DNA sequencing methods and various strategies used for genome sequencing and much more vital information related to gene expression and analysis
DNA Methylation: An Essential Element in Epigenetics Facts and TechnologiesQIAGEN
Check out this slide deck from Dr. Thorsten Singer and Dr. Ralf Peist to learn about DNA methylation in epigenetics, from its significance in cancer to strategies for studying it.
Epigenetics definition, history of epigenetics, molecular basis of epigenetics, epigenetic modification, tools to study epigenetics, disease linked with epigenetics, DNA methylation demethylation and enzymes regulating DNA methylation
regulation of gene expression in eukaryotes is a complex mechanism involved many factors. out of many levels of regulations, chromosomal and transcription level of regulation are discussed in this slides.
The epigenetic regulation of DNA-templated processes has been intensely studied over the last 15
years. DNA methylation, histone modification, nucleosome remodeling, and RNA-mediated targeting regulate many biological processes that are fundamental to the genesis of cancer. Here, we
present the basic principles behind these epigenetic pathways and highlight the evidence suggesting that their misregulation can culminate in cancer. This information, along with the promising clinical and preclinical results seen with epigenetic drugs against chromatin regulators, signifies that it
is time to embrace the central role of epigenetics in cancer.
DNA SEQUENCING METHODS AND STRATEGIES FOR GENOME SEQUENCINGPuneet Kulyana
This presentation will give you a brief idea about the various DNA sequencing methods and various strategies used for genome sequencing and much more vital information related to gene expression and analysis
DNA Methylation: An Essential Element in Epigenetics Facts and TechnologiesQIAGEN
Check out this slide deck from Dr. Thorsten Singer and Dr. Ralf Peist to learn about DNA methylation in epigenetics, from its significance in cancer to strategies for studying it.
Ibica2014 p(8) visualizing and identifying the dna methylationAboul Ella Hassanien
DNA methylation is an epigenetic mechanism that cells use to control
gene expression. DNA methylation has become one of the hottest topics in cancer
research, especially for abnormally hypermethylated tumor suppressor genes
or hypomethylaed oncogenes research. The analysis of DNA methylation data
determines the differential hypermethlated or hypomethylated genes that are candidate
to be cancer biomarkers. Visualization the DNA methylation status may
lead to discover new relationships between hypomethylated and hypermethylated
genes, therefore this paper applied a mathematical modelling theory called formal
concept analysis for visualizing DNA methylation status.
Long-lasting alterations to DNA methylation and ncRNAs could underlie the eff...Ben Laufer
Fetal alcohol spectrum disorders (FASDs) are characterized by life-long changes in gene expression, neurodevelopment and behavior. What mechanisms initiate and maintain these changes are not known, but current research suggests a role for alcohol-induced epigenetic changes. We assessed alterations to adult mouse brain tissue by assaying DNA cytosine methylation and small noncoding RNA (ncRNA) expression, specifically the microRNA (miRNA) and small nucleolar RNA (snoRNA) subtypes. We found long-lasting alterations in DNA methylation as a result of fetal alcohol exposure, specifically in the imprinted regions of the genome harboring ncRNAs and sequences interacting with regulatory proteins. The findings of this study help to expand on the mechanisms behind the long-lasting changes in the brain transcriptome of FASD individuals.
Webinar Link: http://www.youtube.com/watch?v=fzdc0GIdCnA
Developmental Origins of Obesity: The Role of Epigeneticszferraro
This presentation will:
- Review the concept of ‘fetal programming’
- Demonstrate that early life nutritional events may serve as molecular memory of individual in utero experiences
- Show how changes persist following multiple rounds of cell division
- Highlight extrinsic (recapitulation) & Intrinsic (genetic) mechanisms that strongly suggest Intergenerational transmission of traits via epigenetics in humans
- Look at how to best move forward as a scientific and clinical community
this is done by me and my team mates of Wayamba University Sri Lanka for our project.From now we decided to allow download this file.I would be greatful if you could send your comments..
And I'm willing to help you in similar works.I'm in final year of my degree(.BSc Biotechnology)..
pubudu_gokarella@yahoo.com
It contains introduction on basic molecular biology followed by detailed description on discovery , mechanism of oncogene activation, their effect on tumerogenesis , name of important oncogenes , their detection and targeted therapies against oncogenes in treating cancer
Nutraceutical market, scope and growth: Herbal drug technologyLokesh Patil
As consumer awareness of health and wellness rises, the nutraceutical market—which includes goods like functional meals, drinks, and dietary supplements that provide health advantages beyond basic nutrition—is growing significantly. As healthcare expenses rise, the population ages, and people want natural and preventative health solutions more and more, this industry is increasing quickly. Further driving market expansion are product formulation innovations and the use of cutting-edge technology for customized nutrition. With its worldwide reach, the nutraceutical industry is expected to keep growing and provide significant chances for research and investment in a number of categories, including vitamins, minerals, probiotics, and herbal supplements.
Professional air quality monitoring systems provide immediate, on-site data for analysis, compliance, and decision-making.
Monitor common gases, weather parameters, particulates.
This pdf is about the Schizophrenia.
For more details visit on YouTube; @SELF-EXPLANATORY;
https://www.youtube.com/channel/UCAiarMZDNhe1A3Rnpr_WkzA/videos
Thanks...!
Comparing Evolved Extractive Text Summary Scores of Bidirectional Encoder Rep...University of Maribor
Slides from:
11th International Conference on Electrical, Electronics and Computer Engineering (IcETRAN), Niš, 3-6 June 2024
Track: Artificial Intelligence
https://www.etran.rs/2024/en/home-english/
(May 29th, 2024) Advancements in Intravital Microscopy- Insights for Preclini...Scintica Instrumentation
Intravital microscopy (IVM) is a powerful tool utilized to study cellular behavior over time and space in vivo. Much of our understanding of cell biology has been accomplished using various in vitro and ex vivo methods; however, these studies do not necessarily reflect the natural dynamics of biological processes. Unlike traditional cell culture or fixed tissue imaging, IVM allows for the ultra-fast high-resolution imaging of cellular processes over time and space and were studied in its natural environment. Real-time visualization of biological processes in the context of an intact organism helps maintain physiological relevance and provide insights into the progression of disease, response to treatments or developmental processes.
In this webinar we give an overview of advanced applications of the IVM system in preclinical research. IVIM technology is a provider of all-in-one intravital microscopy systems and solutions optimized for in vivo imaging of live animal models at sub-micron resolution. The system’s unique features and user-friendly software enables researchers to probe fast dynamic biological processes such as immune cell tracking, cell-cell interaction as well as vascularization and tumor metastasis with exceptional detail. This webinar will also give an overview of IVM being utilized in drug development, offering a view into the intricate interaction between drugs/nanoparticles and tissues in vivo and allows for the evaluation of therapeutic intervention in a variety of tissues and organs. This interdisciplinary collaboration continues to drive the advancements of novel therapeutic strategies.
Introduction:
RNA interference (RNAi) or Post-Transcriptional Gene Silencing (PTGS) is an important biological process for modulating eukaryotic gene expression.
It is highly conserved process of posttranscriptional gene silencing by which double stranded RNA (dsRNA) causes sequence-specific degradation of mRNA sequences.
dsRNA-induced gene silencing (RNAi) is reported in a wide range of eukaryotes ranging from worms, insects, mammals and plants.
This process mediates resistance to both endogenous parasitic and exogenous pathogenic nucleic acids, and regulates the expression of protein-coding genes.
What are small ncRNAs?
micro RNA (miRNA)
short interfering RNA (siRNA)
Properties of small non-coding RNA:
Involved in silencing mRNA transcripts.
Called “small” because they are usually only about 21-24 nucleotides long.
Synthesized by first cutting up longer precursor sequences (like the 61nt one that Lee discovered).
Silence an mRNA by base pairing with some sequence on the mRNA.
Discovery of siRNA?
The first small RNA:
In 1993 Rosalind Lee (Victor Ambros lab) was studying a non- coding gene in C. elegans, lin-4, that was involved in silencing of another gene, lin-14, at the appropriate time in the
development of the worm C. elegans.
Two small transcripts of lin-4 (22nt and 61nt) were found to be complementary to a sequence in the 3' UTR of lin-14.
Because lin-4 encoded no protein, she deduced that it must be these transcripts that are causing the silencing by RNA-RNA interactions.
Types of RNAi ( non coding RNA)
MiRNA
Length (23-25 nt)
Trans acting
Binds with target MRNA in mismatch
Translation inhibition
Si RNA
Length 21 nt.
Cis acting
Bind with target Mrna in perfect complementary sequence
Piwi-RNA
Length ; 25 to 36 nt.
Expressed in Germ Cells
Regulates trnasposomes activity
MECHANISM OF RNAI:
First the double-stranded RNA teams up with a protein complex named Dicer, which cuts the long RNA into short pieces.
Then another protein complex called RISC (RNA-induced silencing complex) discards one of the two RNA strands.
The RISC-docked, single-stranded RNA then pairs with the homologous mRNA and destroys it.
THE RISC COMPLEX:
RISC is large(>500kD) RNA multi- protein Binding complex which triggers MRNA degradation in response to MRNA
Unwinding of double stranded Si RNA by ATP independent Helicase
Active component of RISC is Ago proteins( ENDONUCLEASE) which cleave target MRNA.
DICER: endonuclease (RNase Family III)
Argonaute: Central Component of the RNA-Induced Silencing Complex (RISC)
One strand of the dsRNA produced by Dicer is retained in the RISC complex in association with Argonaute
ARGONAUTE PROTEIN :
1.PAZ(PIWI/Argonaute/ Zwille)- Recognition of target MRNA
2.PIWI (p-element induced wimpy Testis)- breaks Phosphodiester bond of mRNA.)RNAse H activity.
MiRNA:
The Double-stranded RNAs are naturally produced in eukaryotic cells during development, and they have a key role in regulating gene expression .
Deep Behavioral Phenotyping in Systems Neuroscience for Functional Atlasing a...Ana Luísa Pinho
Functional Magnetic Resonance Imaging (fMRI) provides means to characterize brain activations in response to behavior. However, cognitive neuroscience has been limited to group-level effects referring to the performance of specific tasks. To obtain the functional profile of elementary cognitive mechanisms, the combination of brain responses to many tasks is required. Yet, to date, both structural atlases and parcellation-based activations do not fully account for cognitive function and still present several limitations. Further, they do not adapt overall to individual characteristics. In this talk, I will give an account of deep-behavioral phenotyping strategies, namely data-driven methods in large task-fMRI datasets, to optimize functional brain-data collection and improve inference of effects-of-interest related to mental processes. Key to this approach is the employment of fast multi-functional paradigms rich on features that can be well parametrized and, consequently, facilitate the creation of psycho-physiological constructs to be modelled with imaging data. Particular emphasis will be given to music stimuli when studying high-order cognitive mechanisms, due to their ecological nature and quality to enable complex behavior compounded by discrete entities. I will also discuss how deep-behavioral phenotyping and individualized models applied to neuroimaging data can better account for the subject-specific organization of domain-general cognitive systems in the human brain. Finally, the accumulation of functional brain signatures brings the possibility to clarify relationships among tasks and create a univocal link between brain systems and mental functions through: (1) the development of ontologies proposing an organization of cognitive processes; and (2) brain-network taxonomies describing functional specialization. To this end, tools to improve commensurability in cognitive science are necessary, such as public repositories, ontology-based platforms and automated meta-analysis tools. I will thus discuss some brain-atlasing resources currently under development, and their applicability in cognitive as well as clinical neuroscience.
Observation of Io’s Resurfacing via Plume Deposition Using Ground-based Adapt...Sérgio Sacani
Since volcanic activity was first discovered on Io from Voyager images in 1979, changes
on Io’s surface have been monitored from both spacecraft and ground-based telescopes.
Here, we present the highest spatial resolution images of Io ever obtained from a groundbased telescope. These images, acquired by the SHARK-VIS instrument on the Large
Binocular Telescope, show evidence of a major resurfacing event on Io’s trailing hemisphere. When compared to the most recent spacecraft images, the SHARK-VIS images
show that a plume deposit from a powerful eruption at Pillan Patera has covered part
of the long-lived Pele plume deposit. Although this type of resurfacing event may be common on Io, few have been detected due to the rarity of spacecraft visits and the previously low spatial resolution available from Earth-based telescopes. The SHARK-VIS instrument ushers in a new era of high resolution imaging of Io’s surface using adaptive
optics at visible wavelengths.
Observation of Io’s Resurfacing via Plume Deposition Using Ground-based Adapt...
The role of DNA methylation in complex diseases
1. The
role
of
DNA
methyla0on
in
complex
diseases
Jordana
Bell
Senior
Lecturer
Department
for
Twin
Research
and
Gene8c
Epidemiology
King’s
College
London
Dec
4th,
2014
3. Epigene0c
Mechanisms
Reproduced
from
Nature
441,
143-‐145
(11
May
2006)
DNA
methyla0on
Histone
modifica0ons
Regula0on
of
chroma0n
structure
and
gene
expression
‣
DNA
methyla0on
&
histone
modifica0ons
‣
Chroma0n-‐remodeling
complexes
‣
Non-‐coding
RNA-‐mediated
gene-‐silencing
‣
Transcrip0on-‐factor
binding
‣
Mechanisms
involved
in
genera0ng
and
maintaining
heritable
chroma0n
structure
and
aTachment
to
the
nuclear
matrix.
Changes
to
the
genome
that
affect
gene
expression
without
changing
the
DNA
sequence
4. Gene
expression
Central
Dogma
A,C,T,G
A,C,U,G
Leu,
Pro,...
Transcrip0on
Start
Site
(TSS)
DNA
DNA
RNA
polymerase
pre-‐mRNA
mRNA
5’UTR 3’UTRORF
5’UTR 3’UTR
pre-‐mRNA
Transcrip0on
End
Site
(TES)
Transcrip0on
5. Epigene0c
regula0on
of
gene
expression
• Epigene0c
marks
regulate
gene
expression,
predominantly
by
changing
chroma0n
structure.
➡Euchroma0n?
➡Ac0ve
histone
marks?
➡No
promoter
DNA
methyla0on?
➡TF
promoter
binding?➡Enhancers
ac0ve?
➡No
RNA-‐mediated
silencing?
Transcript
(mRNA)
6. DNA
methyla0on
• 5-‐methylCytosine
(5mC)
• One
of
the
most
common
and
stable
epigene0c
marks
• Other
types:
5hmC,
5fC,
[4mC,
6mA
in
bacteria]
• CpG
dinucleo0des
&
CpG
islands
• Heritable
through
cell
division,
but
dynamic
or
REVERSIBLE
• Important
role
in
gene
regula0on,
XCI,
genome
stability
netic modifications. A methyl group is
n DNA is treated with bisulfite, unmethylated
thylated cytosines are protected.
Me
NH2
C
C
C
N
C
NO
NH2
C
C
C
N
C
NO
CH3
Unmethylated Methylated
C C
Bisulfite
Conversion
O
C
C
C
N
C
NO
U
NH2
C
C
C
N
C
NO
CH3
C
R A WIDE
ne of several epigenetic modifications. A methyl group is
cytosine C . When DNA is treated with bisulfite, unmethylated
d to uracil, but methylated cytosines are protected.
Me
Me
Me
Me
Me
Bisulfite
Conversion
O
C
C
C
N
C
NO
U
NH2
C
C
C
N
C
NO
CH3
C
R A WIDE
HYLATION
e of several epigenetic modifications. A methyl group is
cytosine C . When DNA is treated with bisulfite, unmethylated
d to uracil, but methylated cytosines are protected.
e
Me
Me
Me
Me
Me
NH2
C
C
C
N
C
NO
NH2
C
C
C
N
C
NO
CH3
Unmethylated Methylated
C C
Bisulfite
Conversion
O
C
C
C
N
C
NO
U
NH2
C
C
C
N
C
NO
CH3
C
SAM SAH
Methyltransferase
7. Origin
and
dynamics
of
epigene0c
varia0on?
Gene0c
influences
Reproduced
from
Mill
&
Heijmans
2013
Nature;
Reik
&
Kelsey
2014
Nature
-‐
Guo
et
al
&
Smith
et
al
2014
Egg Sperm
Embryo
Demethyla)on
Rapid
Turnover
Direct
environmental
exposures
In
utero
effects
Ageing
8. The
importance
of
DNA
methyla0on
in
normal
development
and
throughout
life
‣ Gene
expression
regula0on
‣ Reprogramming:
cell
lineage
&
0ssue
differen0a0on
• All
cells
contain
the
same
genes
–
cell
iden00es
depend
on
which
genes
are
expressed
and
repressed.
This
process
is
in
part
regulated
by
DNA
methyla0on.
‣ Imprin0ng
• Preferen0al
DNA
methyla0on
of
the
promoter
of
a
parent-‐of-‐origin
specific
copy
of
the
allele
to
silence
its
expression.
‣ X-‐chromosome
inac0va0on
‣ Genomic
stability
9. DNA
methyla0on
in
human
disease
• For
the
past
few
decades
DNA
methyla0on
changes
in
specific
genes
have
been
linked
to:
• Imprin0ng-‐related
disorders
• X-‐chromosome
abnormali0es
(eg
XO)
and
XCI
skewing
related
traits
• Cancers
10. Example
of
imprin0ng-‐related
disorders:
Prader-‐Willi
and
Angelman
syndrome
Imprinted
region
on
chromosome
15:
imprin0ng
is
not
the
cause
of
disease,
but
is
responsible
for
the
paTern
of
manifesta0on
of
the
disease
Reproduced
from
hTps://www.peds.ufl.edu
11. X-‐chromosome
inac0va0on
skewing
• Mechanism
of
dosage
compensa0on
• XCI
skewing
can
occur
(eg
preferen0al
inac0va0on
of
maternal
X)
• A
recessive
muta0on
carried
on
the
ac0ve
X
chromosome
can
have
profound
adverse
phenotypic
effects.
• XCI
skewing
has
been
associated
with
haemophilia,
Fragile-‐X
syndrome,
and
Duchenne
Muscular
Dystrophy.
• X-‐chromosome
inac0va0on
(XCI)
is
the
random
silencing
of
one
X
chromosome
in
order
for
female
development
to
proceed
X-Chromosome Inactivation
E. Heard, March 17th 2014
Xa' Xa' X' Xi'
Xist""
RNA'
One of the two X chromosomes must be silenced during e
embryogenesis in order for female development to proc
RNA'Pol'II'
Ac' H3K4'
me2'
Ac' Ac' Ac'
H3K4'
me3'
PcH3K2
me3
Ac6ve'X'chromosome' Inac6
5V
12. Epigene0cs
of
cancer
• More
recently,
genome-‐wide
scans
confirm
gross
epigene0c
abnormali0es
in
cancer
0ssue
• Cancer
is
defined
by
uncontrolled
division
of
abnormal
cells
Reproduced
from
Nephew
and
Huang
2003
Cancer
LeT
Progressive changes in promoter methylation at C
sites during cancer initiation and progression
Nephew"&,Huang,"Cancer"LeI."2003;190:125
• Progressive
epigene0c
changes
in
cancer
ini0a0on
and
progression:
13. DNA
methyla0on
in
human
disease
• For
the
past
few
decades
DNA
methyla0on
changes
in
specific
genes
have
been
linked
to:
• Imprin0ng-‐related
disorders,
eg
Angelman
syndrome
and
Prader-‐Willi
syndrome
(15q),
Beckwith-‐Wiedemann
syndrome
(11)
• X-‐chromosome
abnormali0es
(eg
XO)
and
XCI
skewing
related
traits
• Cancers
• Recent
technologies
allow
for
DNA
methyla0on
assays
throughout
genome,
and
have
the
poten0al
to
iden0fy
disease-‐related
changes
of
modest
effects
across
a
wide
range
of
traits
[EWAS].
16. Consequences
Genes
Complex
Phenotypes
DNA
methyla2on
varia2on
Causes
and
Consequences
of
DNA
methyla0on
varia0on
in
human
popula0ons
Environment
Causes
1. Prior
to
disease
onset
(causal
&
biomarker)
2. Consequence
of
disease
(disease
progression)Gene
Expression
Time
17. Consequences
Complex
Phenotypes
DNA
methyla2on
varia2on
Causes
and
Consequences
of
DNA
methyla0on
varia0on
in
human
popula0ons
Causes
External
Environment
Technical
covariates
Cell
heterogeneity
Internal
Environment
Age
Sex Complex
Phenotypes
Germline
Soma0c
indels
etc
Hormones
Longitudinal
stability
Genes Environment
Gene
Expression
Time
18. Consequences
DNA
methyla2on
varia2on
Gene0c
and
environmental
impacts
on
DNA
methyla0on
variability,
and
their
consequences
Causes
External
Environment
Germline
Genes Environment
Complex
Phenotypes
Gene
Expression
1 2
3
19. TwinsUK
Cohort
Department
for
Twin
Research,
King’s
College
London
~13,000
volunteer
adult
twins:
6,000
monozygo0c
(MZ)
twins
www.twinsuk.ac.uk
Established
in
1992 Same-‐sex
adult
(age
range:
16-‐101)
twin
pairs
20. The
EpiTwin
Project1,2
Aim:
To
iden)fy
Differen)ally
Methylated
Regions
(DMRs)
in
common
complex
disease
www.epitwin.eu
Pain
sensi2vity
Type
2
Diabetes
Depression
Heart
Disease
IVF
Breast
Cancer
Telomeres
Allergy
Asthma
Osteoporosis
Hypertension
Osteoarthri2s
Muscle
mass
Lipids
Psoriasis
Colon
cancerEczema
Obesity
Alcohol
use
10M
methyla2on
sites,
5000
individuals
Differen2ally
Methylated
Regions
(DMRs)
in
Disease
Epigenome-‐wide
Associa2on
Scan
(EWAS)
Bone
mineral
density
Autoimmune
disease
Epigene2c
profiles
of
5,000
UK
Twins
using
DNA
methyla2on
sequencing
and
Illumina
450k
profiles
in
whole
blood.
Select
disease-‐discordant
MZ
twins
1Bell
et
al.
Nat
Commun
2014;
2Davies,
Krause,
Bell
et
al.
Genome
Bio
2014;
21. The
MuTHER
Study1,2
Mul2ple
Tissue
Human
Expression
Resource
TwinsUK(Resource!
!!
Adipose((subcutaneous!fat)!!
Whole(Skin((
Lymphoblastoid(cell(lines((LCL)!
Lymphocytes,!Skeletal!Muscle!
(
850!female!
!twins!
Punch!Biopsies!&!
!blood!samples!!
Mul$%centre+collabora$on+
Kings&College&London&
Wellcome&Trust&Sanger&Ins7tute&
University&of&Oxford&
University&of&Geneva&
University&of&Cambridge&
&&
Expression
Gene0cs
Epidemiology
DNA
methyla2on
Illumina
HT12
Array
Illumina
450k
Array3
~2mln
genotyped
and
imputed
SNPs
Clinical
&
Lifestyle
longitudinal
data
1Nica
et
al.
Plos
Gene0cs
2012
2Grundberg
et
al.
Nature
Gene0cs
2013;
3Grundberg
et
al.
American
Journal
of
Human
Gene0cs
2013;
Adipose
Skin
Lymphoblastoid
cell
lines
Lymphocytes,
Skeletal
Muscle
850 female
twins
Punch
biopsies
& blood
from
P
Deloukas
22. Consequences
DNA
methyla2on
varia2on
Gene0c
and
environmental
impacts
on
DNA
methyla0on
variability
Causes
External
Environment
Germline
Genes Environment
Complex
Phenotypes
Gene
Expression
1
23. DNA
methyla0on
heritability
MZ DZ
Genes
DNA
methyla0on
In
twins,
heritability
es0mates
compare
the
degree
of
phenotypic
similari0es
between
groups
of
MZ
and
DZ
twins.
Heritability
(H)
=
propor0on
of
the
phenotypic
variance
that
is
aTributable
to
gene0c
effects.
Here,
the
phenotype
is
DNA
methyla0on
at
1
CpG-‐site
in
the
genome.
Twin-‐based
studies
can
es0mate
the
narrow-‐sense
heritability
(h2),
which
measures
the
propor0on
of
trait
variance
that
is
due
to
addi0ve
gene0c
effects.
h2
=
2(rMZ
-‐
rDZ)
24. DNA
methyla0on
heritability
across
0ssues
1Grundberg
et
al.
American
Journal
of
Human
Gene0cs
2013;
2
Gordon
et
al.
2012Genome
Research;
3
Bell
et
al.
2012
PLoS
Gene0cs;
Genes
DNA
methyla0on
‣
Blood
samples
from
240
female
twins
‣
377,000
methyla0on
sites
‣
Assess
DNA
methyla0on
heritability
‣
Adipose
0ssue
from
518
female
twins1
‣
424,000
methyla0on
sites
‣
Assess
DNA
methyla0on
heritability
_
__
_
MZ DZ Singleton
0.980
0.985
0.990
0.995
Whole Blood Illumina 450k Adipose Tissue Illumina 450k
➡
Mean
CpG-‐site
heritability
is
0.183
(blood)
and
0.191
(adipose).
10%
CpGs
with
>50%
heritability.
➡
Overlap
of
heritable
probes,
consistent
with
previous
blood(-‐related)
es0mates
in
newborns2.
Correla0on
Zygosity Zygosity
Correla0on
25. DNA
methyla0on
QTLs
(meQTLs)
Genes
DNA
methyla0on
‣
Blood
samples
from
188
female
twins
‣
370,000
methyla0on
sites
&
>3M
genotypes
‣
Genotype-‐methyla0on
associa0ons
‣
Adipose
0ssue
from
649
female
twins1
‣
424,000
methyla0on
sites
&
>3M
genotypes
‣
Genotype-‐methyla0on
associa0ons
0 1
Methyla0on
Unmethylated Methylated
DNA
methyla2on
level
met-‐QTLs
Individuals
AA
AG
GG
Genotype
1
Grundberg
et
al.
AJHG
2013;
26. DNA
methyla0on
QTLs
(meQTLs)
Genes
DNA
methyla0on
‣
Blood
samples
from
188
female
twins
‣
370,000
methyla0on
sites
&
>3M
genotypes
‣
Genotype-‐methyla0on
associa0ons
Distance from the CpG site (kb)
ProportionofmeQTL(1kbbins)
0.000.020.040.060.080.100.12
−100 −50 0 50 100
Distance
to
CpG
(kb)
ProbabilitythatSNPismeQTL
Genome-wide meQTLs
(FDR = 5%)
cis-meQTL (7e-5) 14,206
trans-meQTL (8e-10) 460
➡Cis
meQTLs
are
<5kb
away
from
CpGs
~3.9%
of
probes
‣
Adipose
0ssue
from
649
female
twins1
‣
424,000
methyla0on
sites
&
>3M
genotypes
‣
Genotype-‐methyla0on
associa0ons
Genome-wide meQTLs1
cis-meQTL 36,139
~10%
of
probes
from th
jects an
of metQ
was sign
restricte
or metQ
TVS me
and 52%
whole b
have be
among
metQTL
the rep
those e
sites and
is show
Distance from the probe (kb)
Frequency
−100 −80 −60 −40 −20 0 20 40 60 80 100
02,0004,0006,0008,00010,000
Figure 4. Distribution of Top SNPs Associated with the Probe
We performed metQTL analysis by associating methylation levels
with common sequence variants (MAF > 0.05) located close to the
Distance
to
CpG
(kb)
ProbabilitythatSNPismeQTL
➡Cis
meQTLs
are
<5kb
away
from
CpGs
1
Grundberg
et
al.
AJHG
2013;
27. DNA
methyla0on
QTLs
(meQTLs)
across
0ssues
Genes
DNA
methyla0on
‣
Blood
samples
from
188
female
twins
‣
370,000
methyla0on
sites
&
>3M
genotypes
‣
Genotype-‐methyla0on
associa0ons
Distance from the CpG site (kb)
ProportionofmeQTL(1kbbins)
0.000.020.040.060.080.100.12
−100 −50 0 50 100
Distance
to
CpG
(kb)
ProbabilitythatSNPismeQTL
Genome-wide meQTLs
(FDR = 5%)
cis-meQTL (7e-5) 14,206
trans-meQTL (8e-10) 460
➡Cis
meQTLs
are
<5kb
away
from
CpGs
~3.9%
of
probes
‣
Adipose
0ssue
from
649
female
twins
‣
424,000
methyla0on
sites
&
>3M
genotypes
‣
Genotype-‐methyla0on
associa0ons
Genome-wide meQTLs1
cis-meQTL 36,139
~10%
of
probes
from th
jects an
of metQ
was sign
restricte
or metQ
TVS me
and 52%
whole b
have be
among
metQTL
the rep
those e
sites and
is show
Distance from the probe (kb)
Frequency
−100 −80 −60 −40 −20 0 20 40 60 80 100
02,0004,0006,0008,00010,000
Figure 4. Distribution of Top SNPs Associated with the Probe
We performed metQTL analysis by associating methylation levels
with common sequence variants (MAF > 0.05) located close to the
Distance
to
CpG
(kb)
ProbabilitythatSNPismeQTL
➡Cis
meQTLs
are
<5kb
away
from
CpGs
1
Grundberg
et
al.
AJHG
2013;
3
Shi
et
al.
2014
Nat
Commun
➡
30%
of
CpGs
with
me-‐QTLs
overlap
across
0ssues
Blood
(14,206)
Adipose2 Lung3
1,722
5,7226,758
8,45171,116
4,255
20,131
28. DNA
methyla0on
QTLs
(meQTLs)
1
UK10K
project.
2
Grundberg
et
al.
2013
AJHG;
3
Shi
et
al.
2014
Nat
Commun
Genes
DNA
methyla0on
‣
Whole
blood
samples
from
188
female
twins
‣
370,000
methyla0on
sites
&
>3M
genotypes
‣
Genotype-‐methyla0on
associa0ons
Distance from the CpG site (kb)
ProportionofmeQTL(1kbbins)
0.000.020.040.060.080.100.12
−100 −50 0 50 100
Distance
to
CpG
ProbabilitythatSNPismeQTL
Genome-wide meQTLs
(FDR = 5%)
cis-meQTL (7e-5) 14,206
trans-meQTL (8e-10) 460
➡Cis
meQTLs
are
located
near
CpGs
~3.9%
of
probes
➡
CpGs
with
me-‐QTLs
overlap
across
0ssues
Blood
14,206
Adipose2 Lung3
1,722
5,7226,758
8,45171,116
4,255
20,131
A
propor0on
(4%-‐10%)
of
CpG-‐sites
exhibit
strongly
heritable
effects
with
evidence
for
meQTLs,
predominantly
in
cis.
At
many
of
these
regions
gene0c
effects
are
shared
across
0ssues
(30%
of
regions).
5-‐25%
of
meQTLs
are
also
eQTLs,
depending
on
study.
29. Consequences
DNA
methyla2on
varia2on
Gene0c
and
environmental
impacts
on
DNA
methyla0on
variability
Causes
External
Environment
Germline
Genes Environment
Complex
Phenotypes
Gene
Expression
2
30. Environmental
Epigene0cs
in
Humans
Tobacco
smoking
Snuff
Diet
Stress
Alcohol
consump0on
Exercise
Pathogen
infec0on
UV
radia0on
Sunlight
Genome-‐wide
studies Candidate-‐gene
studies
Air
pollu0on
Lead
&
arsenic
Pes0cides
Benzene
PAHs
Organic
chemicals
Season
of
concep0on
…
1
Nat
Rev
Genet
13:97
2011;
Nat
Rev
Genet
8:
253
(2007);
Many
recent
references
(2014)
32. • Tobacco
Smoking
is
a
major
risk
factor
in
disease
• Mul0ple
EWAS
for
smoking
in
whole
blood:
>25
smoking
differen0ally
methylated
regions
(s-‐DMRs)
iden0fied
and
replicated,
with
top
hits
in
AHRR,
F2RL3,
GFI1,
2q37
• S-‐DMRs
also
observed
in
newborns
from
mothers
who
smoked
during
pregnancy
• Few
studies
in
samples
other
than
blood,
or
on
gene
expression
Environment
DNA
methyla0onSmoking
impacts
the
epigenome
➡Systemic
impacts
of
smoking
on
DNA
methyla0on
and
expression?
33. Environment
DNA
methyla0onSmoking
impacts
the
epigenome
Smoking influences DNA methylation and gene expression in
adipose tissue, and effects are conserved across tissues.
‣
Blood
samples
from
306
female
twins2
‣
26
current-‐
,
94
ex-‐,
186
non-‐smokers
‣
377,000
methyla0on
sites
‣
smoking-‐EWAS
‣117,000
exons
RNA-‐seq
‣
smoking-‐TWAS
(Transcriptome-‐WAS)
‣
Analyses
account
for:
‣Methyla0on:
chip,
posi0on
on
chip,
BS
conversion
‣RNAseq:
mean
GC,
primer
index
‣age,
BMI,
family,
zygosity
‣no
probes
with
SNPs
&
mul0ple
alignments
1
Grundberg
et
al.
2013
AJHG;
2
Tsaprouni
et
al.
Epigene0cs,
in
press;
‣
Analyses
account
for:
‣Methyla0on:
chip,
posi0on
on
chip,
BS
conversion,
blood
cell
subtypes
(FACS)
‣RNAseq:
mean
GC,
primer
index
‣age,
BMI,
family,
zygosity
‣no
probes
with
SNPs
&
mul0ple
alignments
‣
Adipose
0ssue
from
349
female
twins1
‣
35
current-‐,
128
ex-‐,
186
non-‐smokers
‣
396,000
methyla0on
sites
‣
smoking-‐EWAS
‣119,000
exons
RNA-‐seq
‣
smoking-‐TWAS
(Transcriptome-‐WAS)
39. F2RL3
1
Tsaprouni
et
al.
Epigene0cs,
in
press;
Environment
DNA
methyla0on
GFI1
2q37.1
AHRR
4 regions overlap: tissue shared effects
Adipose
Blood1
Robust
markers
of
smoking
in
blood
and
fat
40. Methyla2on
markers
of
smoking
in
3
previously
iden2fied2
genes.
Current
smoker
Never
smoked
12 MZ pairs
‣
Illumina
450k
profiles
(1.7%
CpGs)
‣
Blood
(correc0on1
for
cell
heterogeneity)
Chromosome
ATAD3B
2q37.1 AHRR
GNG12 KLHL29
PCGF3
AHRR INTS1
KCNQ1 ETS2
1""""""""""""""""""""""""""""2"""""""""""""""""""""""""3"""""""""""""""""""""4"""""""""""""""""""""5"""""""""""""""""""6""""""""""""""""""7"""""""""""""""""8""""""""""""""""9""""""""""""""10"""""""""""""11""""""""""""12"""""""""""13""""""""""14"""""""""15""""""""16""""""17"""""18"""""19""""20""""21"""22"
-‐log(P-‐value)
FDR
25%
P=2e-‐6
FDR
90%
P=1e-‐4
1
Houseman
et
al.
BMC
Bioinforma0cs
2012;
2
Shenker
et
al.
HMG
2012;
Besingi&Johansson
HMG
2013;
Philibert
et
al.
Clin
Epigenet
2013;
Sun
et
al.
Hum
Genet
2013;
Zeilinger
et
al.
PLoS
One
2013;
EllioT
et
al.
Clin
Epigenet
2014;
(ALPPL2)
Markers
iden0fied
in
12
smoking-‐discordant
MZ
pairs
Environment
DNA
methyla0on
41. Methyla2on
markers
of
smoking
in
3
previously
iden2fied2
genes.
Current
smoker
Never
smoked
12 MZ pairs
‣
Illumina
450k
profiles
(1.7%
CpGs)
‣
Blood
(correc0on1
for
cell
heterogeneity)
Chromosome
ATAD3B
2q37.1 AHRR
GNG12 KLHL29
PCGF3
AHRR INTS1
KCNQ1 ETS2
1""""""""""""""""""""""""""""2"""""""""""""""""""""""""3"""""""""""""""""""""4"""""""""""""""""""""5"""""""""""""""""""6""""""""""""""""""7"""""""""""""""""8""""""""""""""""9""""""""""""""10"""""""""""""11""""""""""""12"""""""""""13""""""""""14"""""""""15""""""""16""""""17"""""18"""""19""""20""""21"""22"
-‐log(P-‐value)
FDR
50%
P=2e-‐6
FDR
90%
P=1e-‐4
1
Houseman
et
al.
BMC
Bioinforma0cs
2012;
2
Shenker
et
al.
HMG
2012;
Besingi&Johansson
HMG
2013;
Philibert
et
al.
Clin
Epigenet
2013;
Sun
et
al.
Hum
Genet
2013;
Zeilinger
et
al.
PLoS
One
2013;
EllioT
et
al.
Clin
Epigenet
2014;
(ALPPL2)
Markers
iden0fied
in
12
smoking-‐discordant
MZ
pairs
Environment
DNA
methyla0on
Smoking-‐DMRs
in
adipose
0ssue:
adipose-‐specific
and
0ssue-‐shared
effects
Smoking
influences
not
only
methyla0on,
but
also
gene
expression
Smoking
should
be
included
as
a
covariate
in
EWAS
Use
of
methyla0on
markers
as
a
proxy
of
smoking
in
large-‐scale
epidemiological
studies
43. Consequences
DNA
methyla2on
varia2on
Epigenome-‐wide
Associa0on
Scans
(EWAS)
Considera0ons
Causes
External
Environment
Germline
Genes Environment
Complex
Phenotypes
Gene
Expression
Functional annotation
EWAS study design
& power
Methylation assay:
coverage vs sensitivity
Time
Cause or Consequence?
Validation & ReplicationCell and Tissue
specificity
44. Consequences
DNA
methyla2on
varia2on
Epigenome-‐wide
Associa0on
Scans
(EWAS)
Considera0ons:
Study
design
Causes
External
Environment
Germline
Genes Environment
Complex
Phenotypes
Gene
Expression
vs
vs
Case-‐control
T0 T1 T2
Discordant
MZ
twins
Longitudinal
samples
45. EWAS
Methods
and
considera0ons
• EWAS
Study
designs:
tailored
to
research
ques0on
• Tissue
specificity:
disease
relevance/cell
heterogeneity
• Bias
and
covariates:
approaches
to
correct
for
artefacts
• Sta0s0cal
analysis:
test
sta0s0c/signal
distribu0on
• Mul0ple
tes0ng
correc0on
• Valida0on
using
independent
assay
• Replica0on
in
an
independent
sample
• Longitudinal
follow
up
to
establish
reversibility
• Func0onal
follow
up
to
establish
func0onal
relevance
46. EWAS
Examples
• Type
1
Diabetes,
SLE
• Type
2
Diabetes
• Obesity
• Alzheimer’s
disease
• Bipolar
disorder
• Depression
• Au0sm
• Pain
sensi0vity
See
further
reading
list
47. Conclusions
• Epigene0cs
represents
a
set
of
regulatory
mechanisms
that
can
be
stable,
heritable,
yet
also
reversible
• Epigene0c
changes
can
be
under
gene0c
control,
but
also
influenced
by
environmental
factors
• Epigene0c
changes
have
been
linked
to
complex
disease
• Epigene0cs
can
complement
and
extend
previous
genome-‐wide
gene0c
studies
to
help
us
understand
molecular
mechanisms
in
complex
disease.
48. Further
Reading
• DNA
methyla0on
assays
• Laird
Nat
Rev
Genet.
2010
Mar;11(3):191-‐203.
doi:
10.1038/nrg2732
• Heritability
• Visscher
et
al
Nature
Reviews
Gene0cs
12,
529-‐541
doi:10.1038/nrg3000
• EWAS
overviews
• Rakyan
et
al
Nature
Reviews
Gene0cs
12,
529-‐541
doi:10.1038/nrg3000
• Tsai
et
al.
Epigenomics.
2012
Oct;4(5):511-‐26.
doi:
10.2217/epi.12.45.
• Some
EWAS
examples:
• Bell
et
al.
Nature
Commun
2014;
Liu
et
al.
Nature
Biotech
2013;
Rakyan
et
al.
PLoS
Gene0cs
2011;
Bell
et
al.
PLoS
Gene0cs
2012;
Dempster
et
al.
Hum
Molec
Genet
2011;