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 2007

0

20

40

60

80

100

120

140

160

Epigenetic Grants

Year

Gra

nts

Aw

ard

ed

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

References

Age-related DNA methylation changes in normal huma...[Clin Cancer Res. 2007] - PubMed Result

A method to study the expression of DNA methyltran...[Methods Mol Biol. 2007] - PubMed Result

Clinical Trial: Epigenetics in the Aging Process

Decreased androgen receptor gene methylation in pr...[J Clin Endocrinol Metab. 2006] - PubMed Result

EMF Award: Epigenetic Control of Senescence and Aging

Entirely new approach to the treatment of aging

Epidermal stem cells are defined by global histone...[PLoS ONE. 2007] - PubMed Result

Epigenetic inactivation of the premature aging Werner syndrome gene in human cancer -- Agrelo et al. 103 (23): 8822 -- Proceedings of the National Academy of Sciences

From The Cover: Epigenetic differences arise during the lifetime of monozygotic twins -- Fraga et al. 102 (30): 10604 -- Proceedings of the National Academy of Sciences

Google Image Result for http://edoc.hu-berlin.de/dissertationen/seitz-stefanie-2004-10-20/HTML/seitz_html_21996ee1.png

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Human hair genealogies and stem cell latency. [BMC Biol. 2006] - PubMed Result

IngentaConnect Techniques Used in Studies of Age-Related DNA Methylation Changes

It Is Time for a Human Epigenome Project -- Rauscher 65 (24): 11229 -- Cancer Research

Leukemia - Abstract of article: Age-related differences in leukemia biology and prognosis: the paradigm of MLL-AF4-positive acute lymphoblastic leukemia

Mitochondria, oxidants, and aging. [Cell. 2005] - PubMed Result

Modern Pathology - Abstract of article: DNA methylation in breast and colorectal cancers

NOVA | Ghost in Your Genes | Epigenetic Therapy | PBS

NOVA: Epigenetic Therapy

http://www.bio.sci.osaka-u.ac.jp/bio_web/lab_page/ogura/photo-neuron.jpg

PLoS Biology - DNA Methylation Profiling of the Human Major Histocompatibility Complex: A Pilot Study for the Human Epigenome Project

Program | The First Annual Iberian Congress on Anti-Aging Medicine and Biomedical Technologies

Proteome analysis of human hair shaft: from protei...[Mol Cell Proteomics. 2006] - PubMed Result

Reversal of Maternal Programming of Stress Responses ... Altering Epigenetic Marking Later in Life -- Weaver et al. 25 (47): 11045 -- Journal of Neuroscience

Title: Epigenetics and aging: the targets and the marks. Source: Trends in genetics [0168-9525] Fraga yr:2007 vol:23 iss:8 pg:413 -8

The DNA methyltransferases of mammals. [Hum Mol Genet. 2000] - PubMed Result

The emerging role of epigenetics in cellular and o...[Exp Gerontol. 2003 Nov-Dec] - PubMed Result

Transposable Elements: Targets for Early Nutritional Effects on Epigenetic Gene Regulation -- Waterland and Jirtle 23 (15): 5293 -- Molecular and Cellular Biology

Transposable Elements: Targets for Early Nutritional Effects on Epigenetic Gene Regulation -- Waterland and Jirtle 23 (15): 5293 -- Molecular and Cellular Biology

NCI Epigenome Project