lecture 26: epigenetics in human health and disease Flashcards
1
Q
What is epigenetics?
A
- epigenetic modifications mediate changes in gene expression in the absence of change to underlying DNA sequence
- “the structural adaptation of chromosomal regions so as to register, signal, or perpetuate altered activity states” (Bird, 2007)
- chromosome-associated factors that regulate the activity of underlying DNA sequence
- cellular “memory”/’plastic’ in response to environment
2
Q
What are epigenetic modifications?
A
- epigenetic mechanisms are affected by these factors and processes:
- development (in utero, childhood)
- environmental chemicals
- drugs/pharmaceuticals
- ageing
- diet
- health endpoints
- cancer
- autoimmune disease
- mental disorders
- diabetes
- DNA methylation
- methyl group (an epigenetic factor found in some dietary sources) can tag DNA and activate or repress genes)
- Histone modification
- the binding of epigenetic factors to histone “tails” alters the extent to which DNA is wrapped around histones and the availability of genes in the DNA to be activated
- variant histones
3
Q
What is the histone code?
A
- “writer” enzymes that catalyse modification
- “eraser” enzymes that remove modification
- euchromatin (open)
- acetylation
- maybe methylation
- phosphorylation
- heterochromatin (closed)
- lots of methylation
4
Q
What is DNA methylation?
A
- occurs primarily at CpG in vertebrates
- On/Off switch at gene promoters, dimmer at gene enhancers
- CpG island = dense region of CpG sites
- most CpG sites (greater than 90%) are dispersed around the genome at low densities
- methylated CpG site → blocked transcription
- unmethylated CpG site → transcriptionally competent
5
Q
In what way is chromatin a dynamic equilibrium?
A
- open or active euchromatic → closed or inactive heterochromatin
- DNMTs, HDACs, HMTs, MBPs
- adding hypermethylated histone tails
- removing hyperacetylated histone tails
- adding more methylated CpG
- other way
- TETs, HATs, HDMs
- RNA pol complex can access the gene and transcribe it
6
Q
What is X-chromosome inactivation?
A
- all female mammals silence one X chromosome
- expression of non-coding RNA (Xist)
- change in histone posttranslational modification
- elevated H3K9, H3K27 methylation
- loss of histone acetylation
- incorporation of variant histone protein
- macroH2A
- association of chromatin modifying proteins
- e.g. MBD, ATRX
- methylation of CpG islands in DNA
7
Q
What is spatial heterogeneity?
A
- 1 genome: 1000s of epigenomes
- sum total of epigenetic modifications within a cell
- every cell has a distinct epigenome
- (cumulative environmental factors)n →
- genotype →
- (cumulative stochastic influence)n →
- all lead to epigenotype → gene expression → phenotype
8
Q
Is DNA methylation static?
A
- no it is highly dynamic
9
Q
What is epigenetics in foetal programming and DOHaD?
A
- environmental exposure (e.g. diet)
- sub optimal intrauterine environment
- stochastic factors
- genetic and sex specific effects
- disruption in epigenetic profile
- changes in gene expression
- metabolic/endocrine disruption
- modified tissue function/development
- foetal programming/maladaption?
- adverse birth outcome including low birth weight
- predisposition to early life and adult onset disease (e.g. T2D)
- adverse birth outcome including low birth weight
- disruption in epigenetic profile
10
Q
What are DNA methylation platforms?
A
- genome-wide DNA methylation analysis
- infinium Human Methylation27 (HM27) and HM450 bead arrays
- 27,000 CpG sites, 14,500 genes or 486,000 CpG sites - all genes
- 12 samples/array (~$500/Sample)
- 450 targets all regions
- 27 targets promoter regions
11
Q
How variable is the early human epigenome?
A
- unsupervised clustering of 27,000 DNA methylation values (HM27) from human placenta across gestation
- watch this
12
Q
What is a heatmap of most variable probes (1st vs 3rd trimester)?
A
- increasing methylation
13
Q
What does the blood of premature infants show?
A
- large-scale epigenetic differences
- analysis of epigenetic changes in survivors of preterm birth reveals the effect of gestational age and evidence for a long term legacy
14
Q
What is DNA methylation and T-development?
A
- HM450 array analysis
- HT-12 expression analysis
- genome-scale profiling reveals a subset of genes regulated by DNA methylation that programme somatic T-cell phenotypes in humans
15
Q
summary thus far
A
- clear evidence of highly dynamic early life DNA methylation profile in multiple tissues, both before and after birth
- overall increase in methylation level and number of variable CpG sites over time in blood and placenta
- evidence for increasing drift (increasing variance) between individuals over time
- supports a model of cumulative effects of environmental exposure on epigenetic profile during early life
16
Q
What is the relationship between epigenetics and the environment?
A
- the inherent sensitivity of epigenetic processes to subcellular environmental cues, that is a hallmark of differentiation and development, also render epigenetic profile sensitive to external environmental influence - Novakovic et al, 2013
17
Q
What is the influence postnatal maternal care?
A
- epigenetic programming by maternal behaviour
- low licking and grooming → stuff → decreased GR expression → high corticosterone levels, high anxiety, low licking or grooming
- high licking and grooming → increased GR expression → low corticosterone levels, low anxiety, high licking
18
Q
What are components of diet that can contribute a methyl group to DNA?
A
- folate
- vitamin B2
- vitamin B6
- vitamin B12
- choline
- all one carbon donors → SAM-e → primary methyl donor in all eukaryotes → addition of a methyl group
19
Q
What is the effect of maternal diet on the neonatal epigenome?
A
- a focus on folate
- maternal epigenetics and methyl supplements affect agouti gene expression in A/a mice
20
Q
What are other environmental epigenetic regulators as seen in animal studies?
A
- maternal care and stress
- endocrine disruptors
- folate and micronutrients
- assisted reproduction
- superovulation and/or embryo culturing
- alcohol exposure
- smoking
- pollution/heavy metals/particulates
- many others emerging
21
Q
What is seen in a methylation analysis of twins?
A
- classical twin model
- monozygotic (MZ) twins share 100% of genetic material
- dizygotic (DZ) twins share ~50% of genetic variation
- MZ correlation greater DZ correlation (genetic influence)
- within pair MZ differences (environmental/stochastic influence)
- discordant monozygotic twins
- insights into non-genetic mechanisms in disease
22
Q
What is the PETS timeline?
A
- 18-20 weeks - recruitment, diet, stress, lifestyle, conception questionnaires
- 28 weeks
- maternal questionnaires (as above if needed)
- maternal blood → serum/plasma storage
- birth - baby measurements, birth data, (questionnaire data)
- cord blood → serum/plasma (-70C), WBC/CBMC (LN2)
- placenta → multiple biopsies in RNA later (-70C)
- cord tissue → biopsy (-70C), HUVECs (LN2)
- buccal swabs → DNA (-70C)
- 18mth followup
- questionnaires (diet, lifestyle, general), baby measurements
- peripheral blood → serum/plasma (-70C)
- buccal swabs → DNA (-70C)
- PETS = the peri/post-natal epigenetic twins study
23
Q
What is genome-wide methylation data?
A
- evidence of environmental effects
- clear evidence of variabilty within MZ twins
24
Q
What is the relationship between smoking and epigenetics?
A
- maternal
- DNA methylation of 1,062 newborn cord bloods from the Norwegian Mother and Child Cohort Study (MoBa)
- replication in newborn epigenetic study - NEST
25
What is replication of methylation at 26 CpG sites?
26
What are outstanding questions?
* how reproducible are the data in other cohorts?
* what is the size of any 'regional' effect?
* is there any tissue specificity?
* dosage and timing effects?
* functional relevance? is there an effect on expression?
* is there any evidence for stability postnatally – (epigenetic 'legacy') in the absence of continued exposure?
* do genetic factors contribute to inter-individual methylation variation?
27
What is the regional effect of maternal smoking AHRR intron 1 methylation in CBMCs?
28
Is there a difference between those who smoked early and those who smoked throughout pregnancy?
* yes - prolonged exposure is necessary
29
What is the difference between buccal epithelial and placent of smoking vs non-smoking mothers?
30
What is the difference in mean methylation between never smokers and smokers at birth and 18m?
31
What role does genetics play in regulating the early life epigenetic profile?
* CBMCs
* 11/16 MZ cluster - 69%
* 5/10 DZ cluster - 50%
* HUVECs
* 6/13 MZ cluster - 46%
* 0/8 DZ cluster - 0%
* placenta
* 7/8 MZ cluster - 88%
* 3/6 DZ cluster - 50%
* contribution of genetic variation to transgenerational inheritance of DNA methylation
* the effect of geneotype and in utero environment on interindividual variation in neonate DNA methylomes
* GeMes, Clusters of DNA methylation under genetic control, can inform genetic and epigenetic analysis of disease
32
Summary thus far
* variation in DNA methylation in MZ twins at birth highlights the importance of environment in specifying neonatal epigenetic profile
* clear evidence now exists confirming a genetic contribution to the human epigenetic profile
* known carcinogens (e.g. smoking) can induce stable epigenetic change even *in utero*
33
What is evidence for epigenetic disruption in complex phenotypes/disease in humans?
* uniequivocal evidence for:
* imprinting disorders (BWS, SRS)
* ICF syndrome
* all adult cancers
* evidence emerging for:
* immune related (T1D, MS, atopy, asthma, arthritis)
* neurological (bipolar, schiz, MD\< eating disorders, Alzheimers, Parkinsons)
* musculoskeletal (osteoporosis)
* metabolic (type II diabetes, obesity)
* cardiovascular (foetal programming)
34
What is epigenetic disruption in complex human disease?
* personalised epigenomic signatures that are stable over time and cobary
* genome wide survey reveals predisposing diabetes type 2-related DNA methylation variations in human peripheral blood
35
Is there replication of T2D findings?
* genome-wide DNA methylation analysis of human pancreatic islets from Type 2 Diabetic and Non-Diabetic donors identifies candidate genes that influence insulin secretion
* epigenome-wide association study reveals longitudinally stable DNA methylation differences in CD4+ T cells from children with IgE-mediated food allergy
* genome-scale case control analysis of CD4+ T cell DNA methylation in juvenile idiopathic arthritis reveals potential targets involved in disease
36
What kind of disease is cancer?
* epigenetic
* CpG island methylation → methylation spreading
* tumour suppressor gene expression potential → gene silencing
* widespread hypomethylation (for example at late-replicating LADs)
* mislocalisation of DNMT1
37
What is utilising archived clinical material for DNA methylation analysis?
* bone marrow is taken for patient diagnosis
* excess bone marrow is archived
* suitable for methylation analysis (Wong, 2008)
* retrospective cohort of more than 600 patients with clinically annotated samples, including outcome
38
What defines paediatric pre-B cell acute lymphoblastic leukaemia?
* a distinct DNA methylation signature
* genome wide analysis of 17 matched pairs of ETV6-RUNX1 subtype paediatric ALL cases (infinium 27k methylation microarray - targeting gene promoters)
* heatmap shows 115 probes with leukaemic specific methylation
* associated with genes previously implicated in leukaemia, other malignancies and haematopoietic development
* A 15 gene signature accurately defines ALL
* validated on a subset of 85 mixed ALL patients
* represents a pan-ALL biomarkers
* the diagnostic potential of a subset of the markers was investigated
39
What is DNA methylation as a biomarker for disease?
* watch this
40
What is epigenetic deregulation in paediatric acute lymphoblastic leukaemia?
* watch this
41
What are prognostic signatures in paediatric pre B-cell ALL?
* LAT1 intra-genic DNA methylation is associated with relapsed paediatric ALL
42
What is other recent cancer research?
* stability of gene expression and epigenetic profiles highlights the utility of patient-derived paediatric acute lymphoblastic leukaemia xenografts for investigating molecular mechanisms of drug resistance
* integrated genomic analysis of relapsed childhood acute lymphoblastic leukaemia reveals therapeutic strategies
* hypermethylation and down-regulation of DLEY2 in paediatric acute myeloid leukaemia independent of embedded tumour suppressor miR-15a-16-1
* optimised DNA extraction for methylome profiling using neonatal dried blood
43
summary 3
* mounting evidence links distinct epigenetic change to complex phenotypes in humans
* in some instances evidence of epigenetic change exists prior to phenotypic onset
* all human cancers show a disrupted epigenetic profile
* epigenetic profiling has utility at multiple levels in paediatric cancers
* diagnosis, disease monitoring, prognostication
44
What are caveats?
* general lack of reproduction
* differences in phenotyping
* different analytical approaches
* generally insufficient sample size
* technological and $$$ limitations
* general lack of assessment in appropriate target tissue
* lack of longitudinal analysis - cause vs effect?
* efect sizes are often questionable
45
What is the relationship between methylation and alzheimer's?
* methylomic profiling implicates cortical deregulation of ANK1 in Alzheimer's disease
* Alzheimer's disease: early alterations in brain DNA methylation at ANK1, BIN1, RHBDF2 and other loci
46
What is the prevailing model?
* epigenetic variation, disease and ageing
* determined by:
* genetic variation
* environment/lifestyle (including diet)
* stochastic factors
* telomere length
* other?
47
What are conclusions?
* epigenetic variation defines who we are
* mounting evidence links environmentally induced epigenetic change in utero to altered phenotype in animal models
* the in utero period is critical in determining the overall epigenome in humans
* the early life epigenome is highly dynamic and sensitive to environmental influence
* also regulated by underlying genetic effects in a tissue specific manner
* preliminary data link altered DNA methylation to complex diseases such as allergy and cancer in children AND to specific exposures in utero
