Epigenetics:”From Phenomenon to Field”

and Impact on Plastic Surgery Research

Basic Science Seminar SeriesUniversity of MichiganDepartment of Surgery

Section of Plastic Surgery

Brent M. Egeland, MDOctober 13, 2007

Outline

Importance History Definitions Chemistry Laboratory Techniques Implications in Plastic Surgery

Why?

Definition: Epigenetics

Waddington 1942 Originally a combination of the

words "genetics" and "epigenesis" Epigenesis: differentiation of cells

from a totipotent state in embryonic development (used in contrast to "preformationism").

Definition: Epigenetics

69th CSH Symposia (2004): Conference Dedicated to Epigenetics No definition could be agreed upon

”An epigenetic phenomenon is a switch-like change in phenotype that is heritable, but does not involve DNA mutation.” C. David Allis

Why Epigenetics? Post-Genomic Era

ENCODE Genomics and proteomics is not enough Contol of above beyond promoters and

regulators is of utmost importance in: Oncogenesis Stem Cell Biology Genetic therapy Aging

History Coalesced into field of study that

affects all areas of biology Collection of apparently disparate phenomena Over 1000 manuscripts on topic First observations dates to 1941

HJ Mueller ”eversporting displacement” McClintock noted that these positional effects were

the basis of ”mutible loci”

History X-Chromosome Inactivation

Pre-32 cell stage mosaic cell population

History

1980's: Molecular Biology and Genetics

1990's: Genomics 2003: Human Genome

Complete 2004: ENCODE 2005: NCI announce:

Human Epigenome Project

Definition: Epigenome

”Sum of genome-wide epigenetic patterns, distinguishes and defines one tissue from another, stem cells from somatic cells, and aged from young cells.”

National Cancer Institute (NCI), Division of Cancer Biology (DCB)

Workshop on Defining the Epigenome: Addressing the Value and Scope of a Human Epigenome Project

November 28–29, 2005Andrew P. Feinberg and Peter A. Jones

Chromatin

Euchromatin: Uncompacted Active

Heterochromatin: Compacted Untranscribed Changes with cell cycle

30nm Fiber

Beads on a String

H1: Linker histone: contacts the exit/entry of the DNA strand on the nucleosome.

Linker DNA between Histones

Nucleosome

Histone

DNA Packaging

Varys greatly with Cell Cycle DNA never completely ”naked”

Scaffold or Dynamic?

Is chromatin simply a scaffold for packaging DNA?

Dynamic

Histones are Modified DNA is Modified Heritable transcription states

Histone Modifications

K K

Acetylation = Transcriptional CompetenceMethylation = Transcriptionally Silenced

DNA Cellular Differentiation

All cells inherit the same DNA sequences Cellular differentiation processes rely strongly on

epigenetic rather than genetic inheritance.

DNA Modification

Chemical modification of DNA Can be inherited without sequence

change Part of the epigenetic code Methylation is most characterized

epigenetic mechanism Commonly occurs at ”junk” DNA

DNA Methylation

DNA Methylation

CpG Islands

Laboratory Techniques Chromatin Immunoprecipitation Inhibition of Deacetylases (HDAC-I) Site-specific analysis of histone

methylation Multigenerational assesment of above Methylation-specific PCR in-situ

Hybridization Q-PCR in Combination with ChIP

Consequences

Genome is static Genotype Germ Cell Inheritance

Epigenome is dynamic Phenotype Somatic epigenetic inheritance

Cellular "memory" Most terminally (irreversibly)

differentiate Morphogens activate/silence

genes – DNA is not altered

Consequences Mammals cannot respond to loss of some tissues

Inability to regenerate limbs Plant cells do not terminally differentiate

Evolution Short-term adaptation

allowing for reversible phenotype Allows multigenerational revisible

phenotypic switch to express and repress that particular gene

DNA message is unchanged Differential mutation rates associated

with epigenetic features to control the mutation rates of particular genes

Teratogens Exert effects on fetus by epigenetic means Epigenetic effects may preserve effect of

teratogen throughout life of child Can a teratogen exert influence offspring?

Diethylstilbestrol: Second and possibly Third generation effects

Research

2000 2001 2002 2003 2004 2005 2006 20070

20

40

60

80

100

120

140

160

Epigenetic Grants

Year

Gra

nts

Aw

arde

d

Why Do I Care?

?

Plastic Surgery Fundamentally involved in clinical epigenetics

Aging Oncology Wound Healing

Basic Science in Plastic Surgery Regenerative medicine Stem Cell biology Organogenesis Neural plasticity

Oncology

Clin Cancer Res 2007;13(13) July 1, 2007

Aging Chronological or Epigenetically

Defined? Free-Radicals Oxidants Telomeres methylation

Aging Change in phenotype with methylation of DNA.

Werner Syndrome

Breast Cancer

Stem Cells

Organogenesis

Neuroal Plasticity

Why does the CNS lack plasticity?

What are the possibilitys of increasing this plasticity towards regenerative capacity?

Hernia

Male Pattern Baldness

The list goes on....

Are there differing methylation signatures of fibroblasts in normal, hypertrophic, keloid scarring.

Can fibrotic tissue (fibroblasts) be reversed to form skin, bone, muscle?

Is cleft lip epigenetically inhereited?

Can we Modify Epigenetics? Can methylation be reversed? Rate decreased?

Environment? Diet? Pharmacologically?

Diet

Agouti Gene: Yellow fur, obesity, diabetes, tumorogenesis

.

Methyl Donor Diet: (onions, garlic, beets, folic acid, B12, genestein)

No Supplements

HCC: Histone deacetylase inhibitors (HDAC-I) & Demethylating agents

Journal of Clinical Oncology, 2006 ASCO Annual Meeting Proceedings Part I. Vol 24, No. 18S (June 20 Supplement), 2006: 10066

Decitabine & Azacitidine Mechanism of action:

Phosphorylation and direct insertion into DNA

Blocks effects of DNA methyltransferase Causing hypomethylation fo DNA. May reverse inactivation of supressor

genes Can lead to cytotoxicity, bone marrow

suppression

Azacitidine

aza-

5-aza-2'-deoxycytidine

ReferencesAge-related DNA methylation changes in normal huma...[Clin Cancer Res. 2007] - PubMed ResultA method to study the expression of DNA methyltran...[Methods Mol Biol. 2007] - PubMed ResultClinical Trial: Epigenetics in the Aging ProcessDecreased androgen receptor gene methylation in pr...[J Clin Endocrinol Metab. 2006] - PubMed ResultEMF Award: Epigenetic Control of Senescence and AgingEntirely new approach to the treatment of agingEpidermal stem cells are defined by global histone...[PLoS ONE. 2007] - PubMed ResultEpigenetic inactivation of the premature aging Werner syndrome gene in human cancer -- Agrelo et al. 103 (23): 8822 -- Proceedings of the National Academy of SciencesFrom The Cover: Epigenetic differences arise during the lifetime of monozygotic twins -- Fraga et al. 102 (30): 10604 -- Proceedings of the National Academy of SciencesGoogle Image Result for http://edoc.hu-berlin.de/dissertationen/seitz-stefanie-2004-10-20/HTML/seitz_html_21996ee1.pngGoogle Image Result for http://www.biophysik.uni-bremen.de/radmacher/gallery/osteoblast_b.gifGoogle Image Result for http://www.med.ufl.edu/biochem/keithr/fig2pt2_flowchart-sm.gifGoogle Image Result for http://www.mun.ca/biology/desmid/brian/BIOL2250/Week_Two/bead2sol.jpgHuman hair genealogies and stem cell latency. [BMC Biol. 2006] - PubMed ResultIngentaConnect Techniques Used in Studies of Age-Related DNA Methylation ChangesIt Is Time for a Human Epigenome Project -- Rauscher 65 (24): 11229 -- Cancer ResearchLeukemia - Abstract of article: Age-related differences in leukemia biology and prognosis: the paradigm of MLL-AF4-positive acute lymphoblastic leukemiaMitochondria, oxidants, and aging. [Cell. 2005] - PubMed ResultModern Pathology - Abstract of article: DNA methylation in breast and colorectal cancersNOVA | Ghost in Your Genes | Epigenetic Therapy | PBSNOVA: Epigenetic Therapyhttp://www.bio.sci.osaka-u.ac.jp/bio_web/lab_page/ogura/photo-neuron.jpgPLoS Biology - DNA Methylation Profiling of the Human Major Histocompatibility Complex: A Pilot Study for the Human Epigenome ProjectProgram | The First Annual Iberian Congress on Anti-Aging Medicine and Biomedical TechnologiesProteome analysis of human hair shaft: from protei...[Mol Cell Proteomics. 2006] - PubMed ResultReversal of Maternal Programming of Stress Responses ... Altering Epigenetic Marking Later in Life -- Weaver et al. 25 (47): 11045 -- Journal of NeuroscienceTitle: Epigenetics and aging: the targets and the marks. Source: Trends in genetics [0168-9525] Fraga yr:2007 vol:23 iss:8 pg:413 -8The DNA methyltransferases of mammals. [Hum Mol Genet. 2000] - PubMed ResultThe emerging role of epigenetics in cellular and o...[Exp Gerontol. 2003 Nov-Dec] - PubMed ResultTransposable Elements: Targets for Early Nutritional Effects on Epigenetic Gene Regulation -- Waterland and Jirtle 23 (15): 5293 -- Molecular and Cellular BiologyTransposable Elements: Targets for Early Nutritional Effects on Epigenetic Gene Regulation -- Waterland and Jirtle 23 (15): 5293 -- Molecular and Cellular BiologyNCI Epigenome Project