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![Schematic figure of chromatin and histone structure.
Muhonen P , and Holthofer H Nephrol. Dial. Transplant.
2009;24:1088-1096
© The Author [2009].](https://image.slidesharecdn.com/epigeneticsroleindiseases-171205083624/85/Epigenetics-role-in-diseases-4-320.jpg)
![Schematic figure of epigenetic regulation mechanism.
Muhonen P , and Holthofer H Nephrol. Dial. Transplant.
2009;24:1088-1096
© The Author [2009].](https://image.slidesharecdn.com/epigeneticsroleindiseases-171205083624/85/Epigenetics-role-in-diseases-40-320.jpg)
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Epigenetics role in diseases
The document discusses epigenetics and chromatin structure. It defines epigenetics as heritable changes in gene expression that do not involve changes to the underlying DNA sequence. This includes histone modifications and DNA methylation. It describes chromatin as being made up of nucleosomes, which consist of DNA wrapped around histone octamers. The structure of chromatin affects gene expression, with euchromatin typically being gene-rich and actively transcribed, while heterochromatin is gene-poor and transcriptionally silent. Histone modifications like acetylation and methylation also influence gene expression. DNA methylation typically silences genes and occurs most frequently at CpG islands. These epigenetic changes play important roles in development and diseases like
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Epigenetics role in diseases
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- 3. Chromatin Walther Flemmingfirst used the term Chromatin in 1882. Chromatin structure: DNA wrapping around nucleosomes – a “beads on a string” structure. In non-dividing cells there are two types of chromatin: euchromatin and heterochromatin.
- 4. Schematic figure ofchromatin and histone structure. Muhonen P , and Holthofer H Nephrol. Dial. Transplant. 2009;24:1088-1096 © The Author [2009].
- 5. Chromatin Fibers 30 nm chromatinfiber 11 nm (beads) Chromatin as seen in the electron microscope. (source: Alberts et al., Molecular Biology of The Cell, 3rd Edition)
- 6. The basicrepeating unit of chromatin. It is made up by five histone proteins: H2A, H2B, H3, H4 as core histones and H1 as a linker. It provides the lowest level of compaction of double- strand DNA into the cell nucleus. It often associates with transcription. Nucleosome H2A H2B H3 H4 1974: Roger Kornberg discovers nucleosome who won Nobel Prize in 2006.
- 7. Core Histones arehighly conserved proteins - share a structural motif called a histone fold including three α helices connected by two loops and an N-terminal tail
- 8. Histone Octamer Eachcore histone forms pairs as a dimer contains 3 regions of interaction with dsDNA; H3 and H4 further assemble tetramers. The histone octamer organizes 146 bp of DNA in 1.65 helical turn of DNA: 48 nm of DNA packaged in a disc of 6 x 11nm < 6 nm > <11nm>
- 9. The N-terminal tailsprotrude from the core
- 10. EPIGENETICS • Coined byConrad Waddington to describe changes in gene expression during development • Refers to a change in gene expression without a change in the sequence of the gene OR • Modifications in gene expression brought about by heritable, but potentially reversible, changes in DNA methylation and/or chromatin structure • Provide an “extra layer” of transcriptional control that regulates gene expression
- 11. Examples of epigeneticmodifications observed in genome • Position effects • Paramutation • Transvection • Protein conformation • Imprinting • X-inactivation • Histone modification • DNA methylation • RNA silencing
- 12. Structure & Epigeneticsof Euchromatin versus Heterochromatin Paula Vertino, Henry Stewart Talks Me
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- 14. Bhaumik, Smith, andShilatifard, 2007. Types of Histone Modifications
- 15. • Covalently attachedgroups (usually to histone tails) • Reversible and Dynamic – Enzymes that add/remove modification – Signals • Have diverse biological functions Cell, 111:285-91, Nov. 1, 2002 Methyl Acetyl Phospho Ubiquitin SUMO Features of Histone Modifications
- 16. • Small vs.Large groups • One or up to three groups per residue Jason L J M et al. J. Biol. Chem. 2005 Ub = ~8.5 kDa H4 = 14 kDa Contd..
- 17. • Writers: enzymesthat add a mark • Readers: proteins that bind to and “interpret” the mark • Erasers: enzymes that remove a mark Tarakhovsky, A., Nature Immunology, 2010. Histone Modifications and Modifiers
- 18. • Others: Sumoylation(Lysine), ADP Ribosylation (Glutamate) Residue Modification Modifying Enzyme Lysine Acetylation Deacetylation HAT HDAC Lysine Methylation Demethylation HMT HDM Lysine Ubiquitylation Deubiquitylation Ub ligase Ub protease Serine/Threonine Phosphorylation Dephosphorylation Kinase Phosphatase Arginine Methylation Demethylation PRMT Deiminase/Demethylase
- 19. • Do notbind to DNA themselves – Can be recruited by: • Histone modifications (through chromodomains, bromodomains, etc.) • Transcription factors • RNA (fission yeast, mammals, plants) • DNA damage • Act as transcriptional co-regulators • Enhance activities of transcriptional repressors or activators – Co-repressor: ex. HDACs – Co-activator: ex. HATs
- 20. General Roles ofHistone Modifications • Intrinsic – Single nucleosome changes – Example: histone variant specific modifications (H2AX) • Extrinsic – Chromatin organization: nucleosome/nucleosome interactions – Alter chromatin packaging, electrostatic charge – Example: H4 acetylation • Effector-mediated – Recruitment of other proteins to the chromatin via • Bromodomains bind acetylation • Chromo-like royal domains (chromo, tudor, MBT) and PHD bind methylation • 14-3-3 proteins bind phosphorylation Kouzarides, Cell 2007 - Prevent binding: H3S10P prevents Heterochromatin Protein 1 binding
- 21. Euchromatin A lightlypacked form of chromatin; Gene-rich; At chromosome arms; Associated with active transcription.
- 22. Heterochromatin A tightlypacked form of chromatin; At centromeres and telomeres; Contains repetitious sequences; Gene-poor; Associated with repressed transcription.
- 23. General Roles ofHistone Modifications Wade P A Hum. Mol. Genet. 2001. Gene Regulation Moggs and Orphanides, Toxicological Sciences, 2004. DNA Damage
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- 25. Histone Acetylation • Twoenzyme types involved in histone acetylation – HAT: histone acetyltransferase – HDAC: histone deacetylase • Note that acetylation eliminates the positive charge from the amino acid. • It is thought that this changes the chromatin conformation to a form more open to transcription. • ⬆ acetylation = ⬆gene expression.
- 26. HAT/HDAC and Hydrophobic Hormones •It is thought that hydrophobic hormones like thyroid hormone and glucocorticoid influence gene expression by binding to either HDAC or HAT proteins. ⬆ acetylation = ⬆gene expression.
- 27. Role of histoneacetylation • Acetylated histones open up the chromatin and enable transcription. Histones are acetylated by HAT (histone acetylases) which are parts of many chromatin remodeling and transcription complexes.
- 28. Role of histonede-acetylation • Deacetylated histones are tightly packed and less accessible to transcription factors. • Histones are deacetylated by HDAC (histone de-acetylase) proteins.
- 29. Acetylation of Lysines Acetylationof the lysines at the N terminus of histones removes positive charges, thereby reducing the affinity between histones and DNA. This makes RNA polymerase and transcription factors easier to access the promoter region. Histone acetylation enhances transcription while histone deacetylation represses transcription.
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- 31. DNA methylation • Occursat 5’ position of cytosine within the CpG dinucleotide • Approx 70% CpGs are methylated • methylated CpGislands are associated with transcriptional silencing
- 32. Methylation of Argininesand Lysines Arginine can be methylated to form mono-methyl, symmetrical di-methyl and asymmetrical di- methylarginine. Lysine can be methylated to form mono-methyl, di-methyl and tri-methylarginine.
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- 34. Effects of DNAmethylation on gene expression. Ling C , and Groop L Diabetes 2009;58:2718-2725 Copyright © 2014 American Diabetes Association, Inc.
- 35. CpG dinculeotide • Dinucletiderepeat CpG accounts for approx 1% genome • CpGs generally under represented in human genome, but occur close to expected frequency in small regions (~1kb) called CpG islands • Approx 45,000 such islands – most reside within or near promoters or 1st exon of gene • CpG islands have “open” chromatin structure – Contains nucleosomes enriched in acetylated histone (decreased affinity for DNA and each other)
- 36. How does methylationoccur?? • De novo methylation occurs via group of enzymes known as de novo methyl transferases (DNMTs) – Include DNMT1, DNMT3a, DNMT3b – Highly expressed in embryonic cells – Highly specific programme of methylation to include repetitive DNA/parasitic DNA, etc – Changes with development of cell types – Accessory factors also required – ds RNA may direct methylation; ~ Xist known to trigger X inactivation prior to methylation • Considered that transcriptional repression may be trigger for methylation rather than effect
- 37. Repression of transcription Openchromatin; active transcription De novo methyl transferases Methylation of CpGs Repressor complex; methyl domain binding positions histone deactelyases Closed chromatin; transcriptional silence
- 38. DNA Methylation 5-methylcytosine S-adenosylmethionine DNA methyltransferase deoxycytosine N N O OHH -O O N N N O OH H -O O N CH3
- 39. Histone phosphorylation (H3) Histones are phosphorylated during mitosis. Histones are also phosphorylated by signal transduction pathways like the ERK pathway in response to external signals. It is not known how (and if) this phosphorylation contributes to gene expression.
- 40. Schematic figure ofepigenetic regulation mechanism. Muhonen P , and Holthofer H Nephrol. Dial. Transplant. 2009;24:1088-1096 © The Author [2009].
- 41. Mark Transcriptionally relevantsites Biological Role Methylated cytosine (meC) CpG islands Transcriptional Repression Acetylated lysine (Kac) H3 (9,14,18,56), H4 (5,8,13,16), H2A, H2B Transcriptional Activation Phosphorylated serine/threonine (S/Tph) H3 (3,10,28), H2A, H2B Transcriptional Activation Methylated argine (Rme) H3 (17,23), H4 (3) Transcriptional Activation Methylated lysine (Kme) H3 (4,36,79) H3 (9,27), H4 (20) Transcriptional Activation Transcriptional Repression Ubiquitylated lysine (Kub) H2B (123/120) H2A (119) Transcriptional Activation Transcriptional Repression Sumoylated lysine (Ksu) H2B (6/7), H2A (126) Transcriptional Repression Chromatin modifications
- 42. Techniques to StudyHistone Modifications • Chromatin immunoprecipitation (ChIP): Strategy for localizing histone marks • ChIP-qPCR: if you know the target gene • ChIP-chip, ChIP-seq, or other genome-wide techniques: unbiased • Limitations: » Antibody specificity » Inherent biases of localization methods • Mass Spectrometry: • Requires digestion of histones
- 43.
- 44. Massively parallel sequencing Localamplification ChIP-seq
- 45. Contd… Genome “Depth”: the numberof mapped sequence tags “Coverage”: how much of the genome the reads can be mapped to
- 46. ChIP-seq vs. ChIP-chip Mikkelsenet al., 2007
- 47. General Roles ofHistone Modifications Chromatin Condensation Spermatogenesis Kruhlak M J et al. J. Biol. Chem. 2001.
- 49. Epigenetics and complexdiseases • Epigenetics is the reflect of – Environmental factors – Stochastic events – Aging • It is less stable than DNA variations but it is stable enough to explain chronic diseases. It may add to or reverse the effect of DNA variations explaining uncomplete penetrances. • It may explain – Altered sex ratio in complex diseases – Incomplete concordance in monozygotic twins – Cancers Ptak et al. 2008
- 50. Representative Clinical Conditions Suggestedto Have Epigenetic Origins • Type 2 DM and Metabolic Syndrome • Coronary artery disease • Autoimmune Diseases • Cancer • Allergic Disorders • Depression • Neurologic: Alzheimer’s, PD, ALS, Autism
- 51. Methylation in humandisease • ICF- rare recessive disease – Immunodeficiency – Facial anomalies – Mental retardation and developmental delay – Chromosomal instability (1, 9 & 16) • Due to mutations in DNMT3b (de novo methyl transferase) • Specific areas of hypomethylation – Instability sites on chromosomes 1, 9 & 16 – Satellite DNAs 2 & 3 – DNA repeat D4Z4 – X chromosome • Loss of methylation results in either relief from transcriptional silencing or an upregulation of normally silenced genes
- 52. Epigenetic Biology ofCancer Cells: DNA • The coexistence of gene-specific promoter hypermethylation and global genomic DNA hypomethylation is an epigenetic characteristic of cancer cells. • Genes regulated via methylation mechanisms include tumor suppressors, cell cycle regulators, DNA repair enzymes and regulators of apoptosis, among others. • Global genomic DNA hypomethylation is a feature of malignancy because there is a loss in the methylation of repetitive sequences of the genome. This has been linked to the chromosomal instability of cancer cells
- 53. Epigenetic Biology ofCancer Cells: Histones • Loss of the monoacetylated Lys16 and trimethylated Lys20 residues of histone H4 appears in the early phase of cell transformation and increases with tumor progression. • Histone modifications may or may not be associated with promoter hypermethylation. • The tumor suppressor gene CDKN2/p16 is silenced via aberrant methylation in breast cancer (33%), prostate cancer (60%), renal cancer (23%), and colon cancer (92%).
- 54. DNA methylation incancer • Hypermethylation of promoter regions is most well catergorised epigenetic change to occur in tumours – Seen in virtually every tumour type – Associated with inappropriate silencing – Is at least as common as disruption of TSGs eg BRCA1 – associated with familial breast cancer- evidence that 10-15% non familial cases have tumours with hypermethylation of BRCA1
- 55. DNA methylation incancer • Also hypomethylation – may result in enhanced genomic instability; activation of oncogenes • Expression profile of DNMTs also observed in tumours – DNMT1, 3a, 3b all increase with progression of cancer – Expression profile varies depending on cancer type • Methylated CpG may also undergo spontaneous deamination; C>T and result in germline mutation – ~50% inactivating point mutations in TP53 are C>T mutations
- 56. DNA Methylation andGene Silencing in Cancer Cells 1 32 4 1 2 3 4 X CGCG CG CG CG MCG MCG Normal Cancer CG CG CGMCGMCGMCG MCG C: cytosine mC: methylcytosine CpG island
- 57. Normal Cancer Region-Specific Hypermethylation Global Hypomethylation + Progressive Alterationsin DNA Methylation in Cancer
- 58. CpG Island Methylation:A Stable, Heritable and Positively Detectable Signal Normal Epithelia Dysplasia Carcinoma in situ Carcinoma Metastasis 1 2 3 4 5
- 59. Normal Epithelia Dysplasia Carcinoma in situ Carcinoma Metastasis 1 2 3 4 5 CpGIsland Methylation: A Stable, Heritable and Positively Detectable Signal
- 60. Normal Epithelia Dysplasia Carcinoma in situ Carcinoma Metastasis 1 2 3 4 5 CpGIsland Methylation: A Stable, Heritable and Positively Detectable Signal
- 61. Normal Epithelia Dysplasia Carcinoma in situ Carcinoma Metastasis 1 2 3 4 5 CpGIsland Methylation: A Stable, Heritable and Positively Detectable Signal
- 62. DNA Methylation InfluencesCancer Processes DNA Repair Hormonal Regulation Carcinogen Metabolism Apoptosis Differentiation Cell Cycle DNA Methylation
- 63. Hormones, Growth Factorsand Epigenetics Epigenetic drugs can restore the expression of estrogen, progesterone, androgen and retinoic acid receptors.
- 64. Rett syndrome • AffectsX chromosome • Usually presents in females only • Normal development until 6-18months then • Gradual loss of speech, purposeful hand use • Develop microcephaly, ataxia, autism • Due to mutations in the MeCP2 gene (methyl binding domain protein) • Again hypomethylation occurs at specific sites • Target genes not yet identified
- 65. Fragile X syndrome is most commonly caused by a CGG trinucleotide repeat expansion in the 5’ region of the FMR1 gene. Unaffected individuals have 6-50 CGG repeats. >200 CGG repeats is seen in individuals with fragile X. >200 CGG repeats is correlated with hypermethylation at CpG dinucleotides and silencing of the FMR1 gene.
- 66. Epigenetics and inflammation •Th1/Th2 ratio: – Methylation between IL-4 and IL-13 (5q21) reduces expression of Th2 – Th2 polarization increases methylation and decreases histone deacetylation of g-IFN promoter • IL-4/IL-13: – Demethylation + histone modification – Allowing GABA and STAT6 fixations – Lead to IL-4 synthesis that induces IL-13 and IL-15 • FOXP3: – CpG motifs in promoter : methylted in naif and activated LT4 but demethylated in Treg. Lee. Immunity 2002 Jones EMBO J 2006
- 67. Epigenetics in lupus ThabetJ Autoimmunol 12
- 68. Epigenetics in diabetes KeatingJ Cardiovasc Transl res 2012
- 69. Model proposing arole for epigenetic mechanisms in the pathogenesis of type 2 diabetes. Ling C , and Groop L Diabetes 2009;58:2718-2725 Copyright © 2014 American Diabetes Association, Inc.
- 71. A conceptual modellinking epigenomics to cardiovascular disease and cardiovascular risk factors. Baccarelli A et al. Circ Cardiovasc Genet. 2010;3:567-573 Copyright © American Heart Association, Inc. All rights reserved.
- 72. Epigenetic mechanisms canlead to the inhibition of protective genes and activation of pathologic genes associated with renal disease. Reddy M A , and Natarajan R JASN 2011;22:2182-2185 ©2011 by American Society of Nephrology
- 75. Different carbohydrates produceunique genomic responses! High Glycemic Carbs Low Glycemic Carbs 62 genes regulating Same genes turned off; Inflammation, stress, Genes regulating same insulin sensitivity production turned off. Immune responses Kalle et al. Am J Clin Nutr;2007:851:1417-27
- 77. Therapies Targeting EpigeneticErrors • In contrast to mutations, epigenetic changes can be reversed. • Are there therapies that influence epigenetic patterns? – Yes Vorinostat (trade name Zolinza) approved by FDA for cutaneous T cell lymphoma in 2006. Vorinostat is a histone deacetylase inhibitor. ⬆ acetylation = ⬆gene expression. X
- 78. Combination Therapy • PhaseIII Clinical Trial – Vorinostat plus cytarabine and idarubicin. – 85% remission rate after initial treatment.
- 79.
- 80. Position effects • Changein the level of gene expression brought about by a change in the position of the gene relative to its normal environment – Chromosomal rearrangement may separate promoter and transcription unit from an essential distant regulatory element » Leading to reduction or absence of expression – Rearrangement may dissociate transcription unit from an element that serves to silence expression » Leading to inappropriate activation of the gene – Rearrangement may place gene with an enhancer from a second gene » Leading to inappropriate expression of the gene
- 81. Examples of disorderscaused by position effects mechanisms • Burkitts lymphoma - translocation event that places the cMYC gene under the control of an immunoglobulin enhancer • Holoprosencephaly – caused by mutations in/deletions in the sonic hedgehog (SHH) gene. Can also be due to chromosomal rearrangements that involve translocations 15-25kb upstream of gene • Aniridia – caused by mutations in/deletions of the PAX6 gene. Can also be due to chromosomal rearrangements that involve breakpoints downstream of gene – suggests that the rearrangement separates the PAX6 gene from an essential regulatory element
- 82. Position effects • Proximityof genes to centromeres, telomeres or heterochromatic regions can suppress expression – Facioscapularhumeral muscular dystrophy (FSHD) • Maps to 4q35 (close to telomere) • Shown to be associated with deletion of copies of a repeat unit (D4Z4) from the subtelomeric region • 95% patients with FSHD have contraction of repeat units (1-10 vs control up to 100)
- 83. FSHD 4qA 4qB Allelic variants 4qA/B– equal in population But FSHD allele is always of 4qA type D4Z4 repeat blocks 6.2kb beta satellite repeat Actual disease mechanism is unclear – no specific gene identified as causative but considered that the D4Z4 units form boundary between the heterochromatic and euchromatic region •upon contraction the local chromatin relaxes and allows transcription of genes normally under repression, or •upon contraction, the boundary between regions is abolished allowing heterochromatinisation and subsequent downregulation of genes in region upstream genes
- 84. Paramutation • A meioticallyheritable change in expression of one allele invoked by another allele • Initially observed and extensively studied in maize and other plants (pigment genes) but recent evidence suggests it may be more widespread • Usually results in a decrease in expression • Paramutagenic alleles – have ability to induce change in allelic partner • Paramutable alleles – can be induced to alter by paramutagenic allele • Neutral alleles – can neither change or invoke change • Possible human example – the insulin minisatellite which is associated with protection against diabetes – Paternally inherited class I alleles do not predispose to disease when have been allelic to class III allele in father.
- 85. Transvection • Another transsensing mechanism ie involves the interaction of homologous alleles • Observed in Drosophila eg spreading of DNA methylation through successive generations • Modifications can occur across homologs • No known examples in human genome
- 86. Protein conformation • Observedin prions – infectious agents implicated in scrapie, BSE and Creuzfeldt-Jakob disease – have little or no associated DNA – consist primarily of single protein (PrP) which is host encoded and is present at low levels in disease free individuals – Exists as two stable conformers (wild type and abnormal infectious type) – Infectious nature of disease believed to derive from capacity of abnormal protein (PrPsc) to interact with wild type protein and convert it to abnormal type • Observed in yeast – Similar scenario where protein conformer alters a second – Results in inheritance of altered form in daughter cells to produce new strain with different metabolic phenotype
- 87. Imprinting • Most autosomalalleles expressed from both alleles – maternal and paternal origin • Subset of genes are “imprinted” ie expressed from only the mother’s or father’s allele • Alleles identical in each case but regulated differently • Involves several different mechanisms – Differential DNA methylation – Allele specific RNA transcription – Anti sense transcripts – Histone modification – Differences in replication timing • Example of disorders involving imprinting – Prader Willi syndrome – Angelman syndrome – Beckwith Wiedemann • Discussed later
- 88. X- inactivation • Methodof ensuring dosage compensation • In mammals dosage is equalised between sexes by inactivating 1 X chromosome in female cells – Requirement for counting the X chromosomes – Choosing which to inactivate – Inactivation process • Occurs under control of X inactivation centre (XIC) – Region of ~1Mb which contains several control elements and at least 4 genes – Two of these genes are • Xist – encodes a large non coding RNA • Tsix – encodes an RNA which is synthesised antisense to Xist; regulates activity of Xist
- 89. X- inactivation – BothXs are expressing Xist and Tsix at low levels prior to X inactivation – Prior to inactivation the two X chromosomes “communicate” at the XIC - decision is made as to which chromosome will be inactivated – Selected inactive chromosome adopts a heterochromatic form which upregulates the expression of Xist - coats the inactive X chromosome – Process also involves methylation of the inactive X chromosome and modification of the histones • Modification allows silencing of most of the X chromosome genes and thereby dosage compensation
- 90. environment environment CH3 CH3 CH3 CH3 CH3 CH3 methylase demethylase active chromatin inactivechromatin Hypothesis: The steady state methylation pattern is a dynamic equilibrium between methylase and demethylase activities nutrients toxins social behavioral pathological physiological
- 91. Methylated DNA isactively demethylated in a replication independent manner upon induction of histone acetylation by TSA CH3 Human HEK cellsHuman HEK cells CH3 X TSA demethylase Human HEK cells No origin of replication-no replication- no passive demethylation
- 92. HDAC inhibitors inducereplication-independent active demethylation Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e.Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e.Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e.Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e.Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e.Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e.Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e.Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e.Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e.Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. CH3 CH3 CH3 demethylase X TSA HAT binding Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. Q uickTim e™ and a decom pr essor ar e needed t o see t his pict ur e. CH3 Ac Ac Ac Ac Ac Ac Ac demethylase CH3 CH3 ((++)) TTSSAA - M H- M H - M H- M H EcoRIEcoRI controlcontrol MM UU (529bp)(529bp) - R1 D1 X- R1 D1 X Cervoni et al., J. Biol. Chem 276, 40788 (2001) Cervoni et al., J. Biol. Chem. 277, 25026 (2002) Detich et al.,J Biol. Chem. 278, 27586 (2003).
- 93. N CH CN O HNH CH3 C DNA C N CH CHN HNH O DNA dMTase DNMT Is there invivo evidence for a dynamic methylation pattern in postmitotic tissue?
- 94. LOW LG HIGHLG Glucocorticoid receptor gene GR GENE GR RECEPTORS GR GENE GR RECEPTORS Environmental programming of gene activity STRESS RESPONSE STRESS RESPONSE PUP (Day 1-6) ADULT (Day 90) QuickTime™ and a YUV420 codec decompressor are needed to see this picture.
- 95. MECHANISM How are thelong-term effects of maternal care on gene expression in the offspring maintained into adulthood? How are these differences transmitted across generations?
- 96. HIPPOCAMPAL GR(17) REGION16 (5’ NGFI-A RE) METHYLATION TIMELINE LOW HIGH MeanC-Methylation 0 0.2 0.4 0.6 0.8 1.0 1.2 GR (17) PROMOTER NGFI-A EMBRYO DAY 20 BIRTH DAY 1 ADULT DAY 90 WEANING DAY 21 PUP DAY 6 Age * * *
- 97. In the adult(day 90) rat hippocampal GR gene expression of Low LG-ABN offspring is reversed by TSA DNMT NGFI-A X High LG Low LG NGFI-A CH3CH3 Ac Ac Demethylase HDAC TSA
- 98. demethylase DNMT SAM X SAM NGFIA binding NGFIAbinding X Detich et al. J Biol. Chem. 278, 20812-20820 (2003). SAM inhibits replication independent demethylation NGFI-A CH3CH3 Ac Ac Is it possible to reverse the effects of maternal care on DNA methylation and behavior in the adult rat?
- 99. 0 10 20 30 40 50 60 70 80 90 100 110 Low LG-ABN HighLG-ABN Maternal care Timespentininnerfield(min) Vehicle TSA Methionine (a) (b) 0 50 100 150 200 250 300 350 400 Low LG-ABN High LG-ABN Maternal care Totalnumberofsquarescrossed Vehicle TSA Methionine * ** Methionine treatment reverses Open Field Behavior of Adult (Day-90) Male Offspring of High LG-ABN maternal care
- 100.
- 101. What are theimplications of a life-long dynamic epigenome? chemical social Signaling pathways Epigenome phenotype Non-genotoxic agents might have a profound effect on our genome and our health icluding obesity, diabetes, cnacer autoimmune disease