Fri 1110 Jackson-Cook - Association of Genetic Technologists

6/6/2015
Studies of Discordant Identical
Twins Reveal the Presence of
Acquired Chromosomal
Instability & Epigenetic
Alterations
C. Jackson-Cook, PhD, FACMG
Virginia Commonwealth University
Departments of Pathology and
Human & Molecular Genetics
Richmond, VA
Twins
Identical
•1 in 250 births
Tia & Tamera Mowry
Fraternal
•Race (1 in 80-100)
•Heredity (♀ 1 in 58; ♂1 in 116)
Mary-Kate & Ashley Olsen
Michael & Ashton Kutcher
•Fertility Drugs (20%-40%)
•Age
•Number of Previous Pregnancies
Patricia & Giselle Bundchen
Hunter & Scarlett Johansson
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Types of Twins
Identical Twins
"Placentation" by Kevin Dufendach - Licensed under CC BY 3.0 via Wikimedia
Commons
Maria & Teresa Tapia separated by team VCUHS in 2011
Castillo-Fernandez et al. Genome Medicine 2014 6:60 doi:10.1186/s13073-014-0060-z
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Twin Studies to quantify genetic and
environmental influences on a trait
ACE Model
Qihua Tan et al. J Exp Biol 2015;218:134-139
Sources of Variation in
Twin Studies
Primary Study Aim
To measure the frequency of acquired
chromosomal abnormalities and chromosomespecific telomere lengths in twins and to
determine the extent to which the variation in
the frequency of these traits is determined by
genetics and/or environment.
PI: Jackson-Cook (NIH R01ES12074)
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Chromosomal changes
with aging
What is Known?
•Acquired sex chromosomal
changes seen
•Acquired chromosomal changes
are associated with age-related
human diseases
•Telomeres shorten with normal
cellular aging
Telomere (TTAGGG)n
A specialized structure at the ends of eukaryotyic chromosomes.
Maintain chromosomal integrity by preventing end-to-end fusion of
chromosomes.
Chromosomal changes
with aging
What is Not Known?
•Do most acquired chromosomal changes arise from environmental
insults, or are they “programmed” to occur as a result of an
individual’s genetic make-up?
•Are chromosomes with longer or shorter telomeres involved more
often in age-related chromosomal changes?
•Can telomere lengths or acquired chromosomal abnormality
frequencies be used as a screening tool for the early detection and
possibly enhanced treatment of health conditions?
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Twin Ascertainment
Mid-Atlantic Twin Registry
•220 twin pairs
•Ascertained through MATR
•Ages 6 to 80+
•Males (40%) and females (60%)
•Monozygotic & same sex dizygotic
(1) Blood sample
(b) Uncultured Blood Smear
(a) Cultured Lymphocytes
(1 ) Standard
GTG-banding
(2) FISH for aneuploidy
FISH for aneuploidy
(2) Buccal Smear
(3) FISH for chromosome-specific
telomere lengths
FISH for aneuploidy
Cytokinesis Block Micronucleus (MN)
and Cytome Assay
Cytokinesis Block MN/Cytome Assay
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Micronuclei frequency
Mean micronuclei frequency
35
Males
30
Females
25
20
15
10
5
0
7yrs9yrs
10yrs19yrs
20yrs29yrs
30yrs39yrs
40yrs49yrs
50yrs59yrs
60yrs69yrs
70yrs79yrs
80yrs+
Genetic vs environmental
influences
Decreased Frequency
Green, leafy vegetables (p<0.001)
Fruit (p=0.14)
Folate-enriched bread (p=0.035)
Vitamin D (males)(p=0.044)
34.8%
Arthritis (p=0.002)
Increased Frequency
Multi-vitamins (p=0.041)
Vitamin E (p=0.01)
65.2%
Tobacco (females)(p=0.001)
Allergies (p=0.001)
Migraines (p=0.026)
Frequency of Chromosome-Specific
Somatic Cell “Numerical” Changes
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Types of changes
acquired with aging
*
*
Acquired copy number changes
with aging
Acquired aneuploidy is a
complex trait
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Acquired CNVs with aging
Cancerfree
All
Solid
tumors
From: Machiela, et al., 2015, AJHG 96 (3): 487 - 497
(G=Gain; B=LOH; R=Loss)
Epigenetic Patterns in MZ Twins
From: Fraga, et al. (2005) Epigenetic differences arise during the
lifetime of monozygotic twins. PNAS 102: 10604-10609.
Epigenetics
Wikipedia
The study of heritable changes in phenotype (appearance) or gene
expression caused by mechanisms other than changes in the
underlying DNA sequence, hence the name epi- (Greek: επί- over,
above) -genetics. These changes may remain through cell divisions
for the remainder of the cell's life and may also last for multiple
generations. However, there is no change in the underlying DNA
sequence of the organism;[1] instead, non-genetic factors cause the
organism's genes to behave (or "express themselves") differently.[2]
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6/6/2015
.
.
Breaker
.
.
or
Breaker
.
.
Breaker
Main
Power
Supply
.
Brighter
.
Dimmer
Breaker
Genotypic variation
important in
gene expression and
modulating effects
of “environmental”
exposure
From Szyf M, Weaver I, Meaney M (2007) Reprod Toxicol 24: 9-19.
Discordance in MZ twins
Zwijnenburg PJG, et al. (2010) Am J Med Genet 153B (6): 1134-1149.
From: Dongen, et al. (2012)
Nat Rev Genet 13 (9): 640-653
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Concordant or Discordant Twins
From: Bell JT, Spector TD (2011) Trends in Genetics 27: 116-125.
Epigenetic Changes in animals
Weaver,et al. (2004) Maternal
nurturing induced epigenetic
changes in glucocorticoid
receptor gene expression in
offspring; reversible with
cross-fostering
•Diet
•Environment
•Maternal nurturing
•Stress
How are traumatic events experienced in childhood biologically remembered to cause morbidity and mortality later in life?
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Epigenetic, Telomere & Chromosome Changes in Adult
Twins Having Child Adversity
Multi PIs: C Jackson-Cook and TP York (NIH R01 AG037986)
Jerry Sandusky, former Penn State football coach, was found guilty of
45 counts of childhood sexual abuse in 2012
Childhood Sexual Abuse
•1 in 4 girls; 1 in 6 boys
(Botash, 1997)
•Psychological and physiological impact
(Brown et al., 2009; Miller et al. 2009)
•Adults who were abused die nearly
20 years earlier
(Miller et al., 2009)
Biological Cascade
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Hypothesis
There are differences in the frequency and genomic location of epigenetic and DNA‐based changes that are present in adult twins who have a history of childhood sexual abuse, when compared to their genetically identical co‐twins, who were not abused.
Ascertainment of Discordant Identical Twins
Kendler, et al. 2003
Population-based
sample
female adult twins
n=1942
Returned health
questionnaire
n = 1411
No response
n = 531
MZ Pairs both
providing CSA info
n= 326
Discordant
n = 74
Age at abuse: 10.2 ± 3.5
Current age: 40-55
Contacted 40 pairs
n = 22 pairs
Concordant
n=252
Positive controls (Concordant Pairs)
Negative controls (Healthy Twins)
Aim 1: Is there a difference in the frequency of acquired
chromosomal changes present in adult identical twins
discordant for a history of CSA?
Cytokinesis Block MN/Cytome Assay
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35
30
25
20
15
0
5
10
Micronucleus Frequency
40
45
50
Micronuclei Frequencies Higher
in Abused Twins
Reference
CSA nonexposed
CSA exposed
p<0.001
p=0.009
4
5
5
0
Difference between co-twins more
pronounced with age
p=0.0006
3
5
3
0
2
5
2
0
1
5
0
5
1
0
M
icro
n
u
cle
u
sF
re
q
u
e
n
cy
4
0
CSA Discordant Exposed
CSA Discordant Nonexposed
35
40
45
50
55
60
65
70
Age at Blood Draw
Biological age of CSA+ twins 9.9 years older than CSA- identical co-twin
Is the observed increase in MN frequency due to more of the same
chromosomal abnormalities or are there differences in the chromatin
present in MN for the CSA+ compared to CSA- identical co-twins?
-
Spectral Karyotyping
Telophase
Cyt B Block
Metaphase
•100 MN/twin
Anaphase
-
2N
Diploid
Interphase
Cell
2N-1
Metaphase
Anaphase
Telophase
Cyt B Block
-
2N-1
Metaphase
Anaphase
Telophase
Cyt B Block
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% Total Observations
No significant difference in the pattern of
chromatin present in MN was detected
between CSA+ and CSA- twins
25
20
15
10
5
0
1
2
3
4
5
6
7
8
9 10 11 12 13 14 15 16 17 18 19 20 21 22 X Mix
Chromosomes
CSA
No CSA
Aim 2: Is there is a difference in the telomere lengths of adult
identical twins who are discordant for a history of CSA?
Scoring
•Telomere average of
20 homologs/10 metaphase spreads
Difference in Mean Telomere Intensity
Telomere intensity scores in women experiencing CSA involving
intercourse tended to be less than those of their
unexposed identical co-twins
p=0.06
p=0.803
p=0.98
p=0.98
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Co-twin Difference in Telomere Signal
Intensity
6/6/2015
Aim 3: Do identical twin pairs discordant for CSA have
more epigenetic differences than concordant CSAidentical pairs?
HumanMethylation450
Figure from Illumina Website
Age: 55
A: 30
B: 19.5
Age: 68
A: 15
B: 15
Conclusions
•MN frequency significantly increased in CSA+ twins
compared to CSA- identical co-twins
 Difference between co-twins increases with age
 No clear difference in pattern of chromosomes present in MN
•Significant chromosome-specific telomeric attrition noted
in twins experiencing intercourse compared to their CSAidentical co-twins
 Telomeres with greatest difference: 1q, 3p & 3q, 4q, 5p, 6p, 8p, 11q, 13q
 No clear trend for attrition for other forms of CSA
•Differences in methylation patterns are acquired between
co-twins with greater divergence seen in twins discordant
for CSA
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Acquired traits associated with
trisomy 21
•
How early do the biological changes arise that are
associated with premature aging?
•
Do biological changes result from alterations
involving genes not only on chromosome 21, but
also genes or gene networks localized to other
chromosomes?
•
Could early recognition of symptoms or
genetic/epigenetic biomarkers for aging allow for
the development of more effective interventions or
treatments?
•
Can knowledge gained about alterations leading to
premature aging in Ds be exploited to inform
scientists about the biological cascade of factors
influencing aging in all people?
Challenges to research
Chromosome 21
Environment
Genes not associated with chromosome 21
Mitochondrial genes
Studies of cells/DNA/RNA from individuals with
mosaicism can help to recognize
effects of trisomy 21
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Studies of cells from individuals with
mosaicism provide unique vision
Chromosome 21
Environment
Genes not associated with chromosome 21
Mitochondrial genes
Studies of cells from individuals with
mosaicism provide unique vision
Chromosome 21
Environment
Genes not associated with chromosome 21
Mitochondrial genes
Studies of cells from individuals with
mosaicism provide unique vision
Chromosome 21
Environment
Genes not associated with chromosome 21
Mitochondrial genes
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Studies of cells from individuals with
mosaicism provide unique vision
Chromosome 21
Environment
Genes not associated with chromosome 21
Mitochondrial genes
•May provide tool(s) for recognizing
“at risk” individuals
•Reversibility of epigenetic marks gives promise for treatment
“Nature” versus “Nurture”
Francis Galton
“Nature” modulated by “Nurture”
The (TTAGGG)n
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Acknowledgements
Grant: RO1 AG037986
Timothy P. York
Jenni Brumelle
Kaitlyn Dochelli
Virginia Anne Hagood
Kenneth S. Kendler
Lindon J. Eaves
Ananda Amstadter
Grant: R01 ES12074
Kimberly Jones
Trisomy 21/DS Mosaicism Project
Kellie Archer
Kelly Rafferty
Chariyawan Charalsawadi
Discordant Identical Twins to Identify Acquired
Epigenetic Alterations
Roos, et al., (2014) Epigenomics 6(3): 299-309.
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