lecture 26: epigenetics in human health and disease Flashcards Preview

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Flashcards in lecture 26: epigenetics in human health and disease Deck (47):

What is epigenetics?

  • 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 


What are epigenetic modifications?

  • 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 


What is the histone code?

  • "writer" enzymes that catalyse modification 
  • "eraser" enzymes that remove modification 
  • euchromatin (open)
    • acetylation 
    • maybe methylation 
    • phosphorylation
  • heterochromatin (closed)
    • lots of methylation  


What is DNA methylation?

  • 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 


In what way is chromatin a dynamic equilibrium?

  • 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 


What is X-chromosome inactivation?

  • 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 


What is spatial heterogeneity?

  • 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 


Is DNA methylation static?

  • no it is highly dynamic 


What is epigenetics in foetal programming and DOHaD?

  • 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) 


What are DNA methylation platforms?

  • 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 


How variable is the early human epigenome?

  • unsupervised clustering of 27,000 DNA methylation values (HM27) from human placenta across gestation 
  • watch this 


What is a heatmap of most variable probes (1st vs 3rd trimester)?

  • increasing methylation 


What does the blood of premature infants show?

  • 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 


What is DNA methylation and T-development?

  • 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 


summary thus far

  • 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 


What is the relationship between epigenetics and the environment?

  • 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


What is the influence postnatal maternal care?

  • 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 


What are components of diet that can contribute a methyl group to DNA?

  • 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 


What is the effect of maternal diet on the neonatal epigenome?

  • a focus on folate 
  • maternal epigenetics and methyl supplements affect agouti gene expression in A/a mice 


What are other environmental epigenetic regulators as seen in animal studies?

  • 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 


What is seen in a methylation analysis of twins?

  • 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 


What is the PETS timeline?

  • 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 


What is genome-wide methylation data?

  • evidence of environmental effects 
  • clear evidence of variabilty within MZ twins 


What is the relationship between smoking and epigenetics?

  • maternal 
  • DNA methylation of 1,062 newborn cord bloods from the Norwegian Mother and Child Cohort Study (MoBa) 
  • replication in newborn epigenetic study - NEST 


What is replication of methylation at 26 CpG sites?


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?


What is the regional effect of maternal smoking AHRR intron 1 methylation in CBMCs?


Is there a difference between those who smoked early and those who smoked throughout pregnancy?

  • yes - prolonged exposure is necessary 


What is the difference between buccal epithelial and placent of smoking vs non-smoking mothers?


What is the difference in mean methylation between never smokers and smokers at birth and 18m?


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


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


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)


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 


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 


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 


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 


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 


What is DNA methylation as a biomarker for disease?

  • watch this


What is epigenetic deregulation in paediatric acute lymphoblastic leukaemia?

  • watch this


What are prognostic signatures in paediatric pre B-cell ALL?

  • LAT1 intra-genic DNA methylation is associated with relapsed paediatric ALL 


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 


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 


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


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


What is the prevailing model?

  • epigenetic variation, disease and ageing 
  • determined by:
    • genetic variation
    • environment/lifestyle (including diet)
    • stochastic factors 
    • telomere length
    • other?


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