a.unicef (sept 2013) meaney€¦ · early environmental regulation of gene expression: how early...

11/15/2013

© Copyright 2013 Michael J. Meaney 1

Early Environmental Regulation of Gene Expression: How Early Experience Exerts a Sustained Influence on Neuronal Function

[email protected]

Michael J MeaneyDouglas Mental Health University Institute

McGill Universityand

Singapore Institute for Clinical Sciences

Summary

The experience of the child is “biologically embedded” and serves to influence health and capacity over the lifespan.

11/15/2013


Summary


This effect is apparent even at the level of theDNA of the individual; the activity of genes implicated in brain development and functionare directly regulated by the social environment.

Summary


This effect is apparent even at the level of theDNA of the individual; the activity of genes implicated in brain development and functionare directly regulated by the social environment.

This effect is potentially stable over time; theimprint of of childhood adversity on the genomeis apparent at later ages, providing a biological basis for an enduring effect on health and capacity.

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The development of an individual is an active process of adaptation that occurs within a social and economic context:

• To resource (food, shelter, safety) availability.

• To social interactions (e.g., parental signals).

The development of an individual is an active process of adaptation that occurs within a social and economic context:

• To resource (food, shelter, safety) availability.

• To social interactions.

This influence is apparent in the epigenetic mechanismsthat regulate genomic structure and function

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Early experience

AbuseFamily strifeEmotional neglectHarsh discipline

Health Risks

DepressionDrug abuseAnxietyDiabetesHeart diseaseObesity

Developmental Origins of Adult Disease

Early experience


Health Risks


Individual differencesin neural and endocrine responses to stress (defensive responses

How does family life influence health over the life span?

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Early experience


Health Risks


Individual differencesin neural and endocrine responses to stress

Poverty

Nutrient Supply

Mate Quality

Violence

Infection

Population density

Parentalinvestment

Defensive Strategies

Foraging/Metabolism

Reproductive Strategies

Environmental Parental Developmentalcondition signal outcome

Robert Hinde: Evolution has shaped the young to use parental signals as a ‘forecast’ of the quality of the environment

into which they have been born. For most species, there isno single, optimal phenotype.

Evolutionary biology - Maternal effects

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Health andHuman Capacity

Early experience


Individual differencesin neural and endocrine responses to stress

Adversity

Environmental regulation of phenotypic variation

What is the biological basis for ‘programming’ effectswhereby environmental signals acting over perinataldevelopment associate with stable changes in transcriptionand complex phenotypes (physiology, behaviour)?

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Summary

• Parental care affects the activity ofgenes in the brain that regulate stressresponses, neural development and reproduction.

• This parental effect involves a forma “plasticity” at the level of the DNA.

Epigenetics: Any functional change in the genome thatdoes not involve an alteration of DNA sequence.

If they ask you anything you don’t know,just say its due to epigenetics.

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Nucleosome core particle: ribbon traces for the 146-bp DNA phosphodiester backbones (brown and turquoise) and eight histone protein chains (Luger et al. Nature 1997).

+ -

Prevents TFbinding to DNA

TF binding involvesalteration of

chromatin structure

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+ -



chromatin structure

Acetyl group

B. Turner. Chromatin structure and gene regulation. 2001.

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+ -



chromatin structure

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C T A C G T A C T C G G A A T C T C GCH3CH3

CH3

C T A C G T A C T C G G A A T C T C G

Genetic code is defined by the sequence of four nucleotides that produce proteins.

RNAs, proteins

Epigenetic effects refer to modifications of the DNA that alter the activity of the gene, but not its function.

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SIN

SIN3a

CH3

CH3

MeCP2/MBD2

HDACMeCP2/MBD2

DNA Methylation can inhibit gene expressionby blocking transcription factor binding

HDAC HDAC: Histonedeacetylase

Methylated DNAbinding proteinCH3

CH3

CH3

CH3

HDAC


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+ -



chromatin structure

Every cell in your body has the same nuclear genes, but…?

Multiple phenotypes from a common genotype

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Maternal licking/grooming

Environmental epigenetics hypothesis: Environmentalevents activate intracellular signals that remodelthe epigenome, leading to sustained alterations inthe structure and function of the genome, and thus stable effects on gene transcription.

Parental care PhenotypeGeneexpression

Epigeneticmarks

Parental signals as a source of phenotypic plasticity?

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Epigeneticmarks


Maternal licking/grooming

Source of tactile stimulation/nurturance: Enhances activity of endocrine systems (e.g., GH/IGF) that

promote somatic growth, suppresses those (glucocorticoids) that inhibit growth

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Variations in maternal care

��6�� 11 1� �� 6 ��0

50

100

150

200F

req

uen

cy c

ou

nt

% Licking/grooming

Stress responses

Neural development

Learning & memory

Metabolism

Reproduction (females)

Broad range of parental effects

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High LG Low LG


Epigeneticmarks


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Stress responses

Neural development

Learning & memory

Metabolism

Reproduction (females)

Broad range of parental effects

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GlucocorticoidReceptor

Hypothalamus

Pituitary

Adrenals

Hippocampus

Glucocorticoids

��

��

��

Stress

(-)

CRF/AVP

ACTH

��

CRF: corticotropin releasing factor. ACTH: adrenocorticotropin

Hypothalamus

Pituitary

Adrenals

High LG offspring

Glucocorticoids

��

��

��

(-)

CRF

ACTH

(-)

Hypothalamus

Pituitary

Adrenals

Low LG offspring

Glucocorticoids

��

��

��

(-)

CRF

ACTH

(-)

Individual differences in glucocorticoid receptorlevels lead to altered pituitary-adrenal responses

to stress

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High LG offspring show more modest HPA stress responses

Pre S10 S20 P20 P40 P60 P1200

200

400

600

800

1000High LGLow LG

Pla

sma

AC

TH

(p

g/m

l)

�

�

Pre 10 20 40 60 80 1000

10

20

30

40

50

60

70

80High LGLow LG

Cor

ticos

tero

ne (

µg/

dl)

� �

�

�

�

Disruption of hippocampal GR signaling eliminates this maternal effect

Hypothalamus

Pituitary

Adrenals

High LG offspring

Glucocorticoids

��

��

��

(-)

CRF

ACTH

(-)

Hypothalamus

Pituitary

Adrenals

Low LG offspring

Glucocorticoids

��

��

��

(-)

CRF

ACTH

(-)

Cross-fostering reveals evidence for direct, postnataleffects of maternal care

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Hippocampal GR(17) mRNA

1 2 3 4 5 6 7 8 9 1011 12 2 3 4 5 67 8 9

5’ 3’

DG CA1 CA2 CA30.00

0.05

0.10

0.15

High LG

Low LG

Rel

ativ

e O

DU

Hippocampal GR(17) mRNA

1 2 3 4 5 6 7 8 9 1011 12 2 3 4 5 67 8 9

5’ 3’

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1681 ccc1741 ctctgctagt gtgacacact t1cg2cgcaact c3cgcagttgg 4cggg5cg6cgga ccacccctg7c1801 ggctctgc8cg gctggctgtc accct9cgggg gctctggctg c10cgaccca11cg ggg12cgggct1861 c13cgag14cggtt ccaagcct15cg gagtggg16cg gggg17cgggag ggagcctggg agaa

DNA sites that regulate glucocorticoid receptor gene

NGFI-A

GR Promoter 17 Sequence

1 2 3 4 5 6 7 8 910 1112 2 3 4 5 6 78 9

Variable exon 1 region Constant region

5’ 3’

NGFI-A GR gene

GR mRNA

NGFI-A regulates GR gene expression

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5’ 3’0.0

0.2

0.4

0.6

0.8

1.0Low LG

High LGc-

Met

hyl

atio

n

*

NGFI-A site

5’…tgcgggggcgggg…3’NGFI-A

CH3CH3

Low/Low High/High Low/High High/Low0.0

0.2

0.4

0.6

0.8

c-m

eth

yla

tio

n

5' CpG region of NGFI-A/RE

5’…tgcgggggcgggg…3’NGFI-A

CH3 CH3CH3

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SIN

SIN

CH3

CH3

CH3

CH3MeCP2/MBD2

HDACCH3

CH3

MeCP2/MBD2

DNA Methylation can inhibit gene expressionby blocking transcription factors binding

HDAC HDAC: Histonedeacetylase

Methylated DNAbinding protein

HDAC


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High Low

0.0

0.2

0.4

0.6

0.8

1.0

0.0

0.2

0.4

0.6

0.8

1.0

An

tib

od

y/In

pu

t

HistoneAcetylation

NGFI-AAssociation

Increased methylation of the exon 17 GR promoter associates with decreased H3K9ac, reduced NGFI-A binding and GR expression

0

0.1

0.2

0.3

0.4

GR

IR

(R

OD

-IO

D)

GRExpression

High Low High Low

Effects are reversed with intra-hippocampal infusion of HDAC inhibitor

Low/Veh Low/TSA High/Veh High/TSA

Basal Stress Stress + 40 Stress + 60 Stress + 900

10

20

30

40

50

Co

rtic

ost

ero

ne

( µ

g/d

l)

Time (min)

Low/Veh

Low/TSAHigh/Veh

High/TSA

Glucocorticoid stress response

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GGAGCCTGGGAGAACCAAGCCTCGGAGTGGGCGGGGGCGGGAGCH3CH3

GGAGCCTGGGAGAACCAAGCCTCGGAGTGGGCGGGGGCGGGAGCH3

Offspring of Low LG mothers

Offspring of High LG mothers

Lower levels of GR in hippocampus -Increased stress response

High levels of GR in hippocampus -Modest stress response

cAMP

5-HT7 receptor

5-HTTactile stimulation

(maternal LG)

PKA CBP

Summary of in vivo and in vitro studies

NGFI-A (egr-1, zif-268, krox-24)

(T4 - T3)

SP1

CBPNGFI-ASP1

CpGme Exon 17 promoter

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C T A C G T A C T C G G A A T C T C G

CH3

CH3

CH3

cAMP

NGFI-A

5-HT7 rec

5-HT

GR gene

PKA CBPSP1

CBPSP1

Do comparable processes occur in humans?

• Post-mortem studies of hippocampus.

• Samples from suicide victims/controls.

• QSBB (Gustavo Turecki) – forensic phenotyping.

• Human exon 1F promoter (Turner & Muller, 2005)

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Human glucocorticoid receptor gene

A B CD E F G H I J 2 3 4 5 67 8 9


5’ 3’

Exon 1F is the human ortholog of the rat exon 17 GR promoter(70% homology) and contains an NGFI-A consensus sequence.

Human glucocorticoid receptor gene

A B C DE F G H I J 2 3 4 5 67 8 9


5’ 3’

Control Suicide0.0

0.5

1.0

1.5GRtotal

GR

mR

NA

Control Suicide0.0

0.3

0.6

0.9

1.2 GR1F

GR

1-F

var

ian

ts

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Suicide vs abuse - GR expression

GRtotal

GR

mR

NA

/GA

PD

H (

log

co

nc.

)

Control Suicide Suicide- Abuse + Abuse

0.0

0.4

0.8

1.2

*

GR

-1F

mR

NA

/GA

PD

H (

log

co

nc.

)


0.0

0.5

1.0

1.5

*

GR1F

GR

1-F

CpG

Met

hyla

tion

(%)


0

20

40

60*

Suicide vs abuse - CpG methylation

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Hypothalamus

Pituitary

Adrenals

Controls

Glucocorticoids

��

��

��

(-)

CRF

ACTH

(-)

Hypothalamus

Pituitary

Adrenals

Maltreatment

Glucocorticoids

��

��

��

(-)

CRF

ACTH

(-)

Childhood maltreatment associates with increased centralCRF levels and greater HPA and autonomic responses to stress

(DeBellis et al 1994; Heim et al 2000; Lee et al 2005)

Childhood adversity and NR3C1 promoter methylation in DNAfrom periperhal samples

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Early experience


Health Risks


Individual differencesin neural and endocrine responses to stress (defensive responses

How does family life influence health over the life span?

Childhood adversity and NR3C1 promoter methylation in DNAfrom periperhal samples

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345 genes

Secure:

TBCD (23.89%: Body)

CEBPE (19.70%: TSS200)

Attachment at 18 months (Secure Vs Insecure)associates with epigenetic variation (6-8 yrs)

612 probes≥. 5% (p<0.05)

Insecure:

ABR (21.35%: Body)

SHANK2 (9.4%: 1st Exon)

DRD4 (5.76%: Body)

Microtubule formation LTP

Synaptic density

Reference 1: ADHDgene Database (I)Reference 1: ADHDgene Database (I)

64

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213 Genes in the ADHDgene database (24 Genes are hot genes)

36 Probes (25 Genes) of ADHD database genes are found in the 2907 DMPs list.

213 Genes in the ADHDgene database (24 Genes are hot genes)

36 Probes (25 Genes) of ADHD database genes are found in the 2907 DMPs list.

Reference 1: ADHDgene Database (II)Reference 1: ADHDgene Database (II)

ADRA1A FGF10

ANK3 GNAO1

ARRB2 HK1

ATXN1 HLA‐DRB1

BAIAP2 ITGA11

BAIAP2 MEIS2

BAIAP2 MEIS2

BAIAP2 NCKAP5

BAIAP2 PTPRG

BDNF PTPRG

CACNA1C SH3BP5

CDH13 SH3BP5

CHRNA4 SLC6A3

CHRNA4 UNC5B

COMT UNC5B

DBH ZNF423

DRD4 ZNF423

EMP2 ZNF423

Child attention problems and co-methylationChild attention problems and co-methylation

Child attention problems (SDQ) associated with differential methylation of 1747 probes (957 genes).

30 probes (22 genes) associated with child attention problems (-0.3 < r >0.3) show co-methylation (-0.6 < r >0.6).

rs= -0.36, p=0.02 rs= 0.36, p=0.02 rs= -0.64, p<0.001

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7 Genes(GO) + 94 Genes(Pathway)--123 Probes--

7 Genes(GO) + 94 Genes(Pathway)--123 Probes--

Hierarchical clustering of pair-wise correlation

DMPs

DM

Ps

Pair-wise Correlation

20 40 60 80 100 120

20

40

60

80

100

120-0.8

-0.6

-0.4

-0.2

0

0.2

0.4

0.6

0.8

1

The Most highly correlated cluster (30 probes, AvgCorr=0.8)

67Co-methylation Clusters

The ‘partial’ mediating role of maternalpsychopathology

The ‘partial’ mediating role of maternalpsychopathology

Maternaldepression(36 months)

Maternalemotionalneglect


Child negativeemotionality (36

moths)p = 0.01

p = 0.008p = 0.001

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The ‘partial’ mediating role of maternal psychopathology


Maternalemotionalneglect


Child negativeemotionality (36

moths)p = 0.01

p = 0.008p = 0.001

(Parenting)

Do effective treatment programs that target mothers affect DNA methylation profiles

in the children?

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90 Unique Samples Infinium 450K Array Blood Samples 27 years old Variables:

methy_grp (0= control/no treatment ;1=intervention) ETXGROUP: 1 and 2 = different forms of no treatment

versus 3 and 4 which represent prenatal intervention and combined pre and postnatal intervention, respectively

CHILDGENDER 1=male 2=female Child Abuse at age 4 and age 15

90 Unique Samples Infinium 450K Array Blood Samples 27 years old Variables:

methy_grp (0= control/no treatment ;1=intervention) ETXGROUP: 1 and 2 = different forms of no treatment

versus 3 and 4 which represent prenatal intervention and combined pre and postnatal intervention, respectively

CHILDGENDER 1=male 2=female Child Abuse at age 4 and age 15

Child abuse is physical, sexual or emotional mistreatment or neglect

Principal Component Analysis

Component Fstat p

Gender (3.0%) 7.32 .003 x 10-8

Abuse/4 yrs (3%) 5.80 .002 x 10-2

Abuse/15yrs 1.67 ns

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Principal Component Analysis

Component Fstat p

Gender (3%) 7.32 .003 x 10-8

Abuse/4 yrs (3%) 5.80 .002 x 10-2

Abuse/15yrs 1.67 ns

Treatment (8%) 3.07 .002 x 10-4

Conclusions

• The function of the genome is regulated by epigenetic signals that are subject to environmental regulation.

• These epigenetic signals reflect the quality of thedearly environment, and guide the developmentand function of the brain.

• These effects are potentially stable, but aresubject to modification, potentially over the lifespan.

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Family

Parent-child

Family

Parent-child

Transgenerationalinfluences

Family

Parent-child

Family

Parent-child


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Family

Parent-child

Family

Parent-child


Control Perfume Snake0

1

2

3

4

Response to snake odours

Ton

gu

e-F

lick

Offspringare significantlylarger and withlonger tails

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Defense to snake predation in skink lizards

Most frequent prey

• smaller• shorter tails• less reactive to

snake cues

If mother has been exposed to the scent of apredatory snake then offspring are larger, withlonger tails and ….

Nutrient Supply

Mate Quality

Violence

Infection

Population density

Parentalinvestment

Defensive Strategies

Foraging/Metabolism

Reproductive Strategies

Environmental Parental Developmentalcondition signal outcome

Evolutionary biology - Maternal effects

a.unicef (sept 2013) meaney€¦ · early environmental regulation of gene expression: how early...

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