Introduction and Definition, Factor affecting epigenetics and Inheritance, Mechanism, Epigenetic phenomenon Cancer and epigenetics, Diagnosis , Therapies targeting epigenetic modification, Future

1. EPIGENETICS

3. OUTLINES
• Introduction and Definition
• Factor affecting epigenetics and Inheritance
• Mechanism
• Epigenetic phenomenon
• Cancer and epigenetics
• Diagnosis
• Therapies targeting epigenetic modification
• Future

4. Introduction
• Genetics explains phenotypic trait in
organisms through presence or absence of
specific nucleotide sequence of DNA.
• However, that does not adequately explain
the expression or lack of expression of all
genes.
• That indicates, there are other control
mechanism; such as DNA methylation and
chromatin modification

5. Definition
• C.H. Waddington coined the term epigenetics
to mean above or in addition to genetics to
explain differentiation
• Epigenetics refers to the study of changes in
the regulation of gene activity and expression
that are not dependent on gene DNA
sequence.
• While epigenetics often refers to the study of
single genes or sets of genes, epigenomics
refers to more global analyses of epigenetic
changes across the entire genome

6. • Disruption of such control mechanism is
associated with a variety of disease
– With behavioral/neurological manifestation
– Disorder of tissue growth
– Endocrine disorder
– Neoplasia

7. Inheritance
• parent’s experiences are
passed on to offspring
through epigenetic tags
• When the zygote is
formed many epigenetic
tags are removed from the
chromosomes of the
parents, but some remain

8. • Identical twins
– from same zygote
– same genetic information
(including epigenetic tags)
• While infants
– similar environments,
– so little variation in
epigenome

9. • Difference in the
twins’ epigenomes
makes them
become different
when they are older
• One can develop a
disease while the
other is fine

10. Methylation of Agouti Genes in Mice

11. Environment can Influence Epigenetic Changes
Emma Whitelaw, Henry Stewart Talks

12. Factors during pregnancy affecting the
transmission of epigenome
• nutrition of mother
• Stress, social interactions, physical activity
as Stress hormones can travel from the
mother to a fetus
• exposure to toxins

13. GRANDMOTHER’S CURSE!!!!!
Some of the effects of
smoking may be passed from grandmother to
grandchild

14. Epigenetics Mechanisms
Gene Expression
RNA Interference
Histone Modifications
DNA Methylation

16. • Epigenetic alteration are outside the primary
sequence, but nonetheless affect the ability of
gene to be expressed.
• DNA in most cells packaged with histone to
form nucleosome in ‘beads on a string’
structure.
• Chromatin regulation involves high-order
conformational changes
– Relaxation or tightening of this thread of DNA-
histone complex

17. • Further regulation is by assembling promoter-
enhancer complexes via long-range DNA
looping
– Can be blocked by specific DNA-sequence called
insulator
• The core histones are subject to diverse post-
translational modifications, including
methylation and acetylation
– That project from tightly structured nucleosome core

18. DNA methylation

19. Histone
Methylated
DNA
Effect of methylation
Methylation turns
off genes.
Acetylation turn
genes on.

20. 5-Methyl Cytosine is Found in
Heterochromatic Regions

21. Natural Roles of DNA Methylation in
Mammalian System
 Imprinting
 X chromosome inactivation
 Heterochromatin maintenance
 Developmental controls
 Tissue specific expression controls

22. DNA Methylation and
Other Human Diseases
• -- Imprinting Disorder:
• Beckwith-Wiedemann syndrom (BWS)
• Prader-Willi syndrome (PWS)
• Transient neonatal diabetes mellitus (TNDM)
• -- Repeat-instability diseases
• Fragile X syndrome (FRAXA)
• Facioscapulohumeral muscular dystroph
• -- Defects of the methylation machinery
• Systemic lupus erythemtosus (SLE)
• Immunodeficiency, centromeric instability and facial
anomalies (ICF) syndrome

23. Histone Modifications

24. Histone modification
the histone code
– Acetylation
– Methylation
– Phosphorylation
– Ubiquitylation
– sumoylation
• Enzymes catalyzing
– Histone
acetyltransferase
– Histone deacetylase
– Histone
methyltransferase
– Histone kinase

25. Histone Modification Status Correlates with
Transcriptional Activity
• Gene activation correlated with H3-K9 acetylation
• Gene silencing associated with H3-K9 methylation

26. Role of histone modification
• DNA transcription
• DNA repair
• DNA replication

27. Histone Modifications and
Human Diseases
Coffin-Lowry syndrome is a rare genetic disorder
characterized by mental retardation and
abnormalities of the head and facial and other
areas. It is caused by mutations in the RSK2 gene
(histone phosphorylation) and is inherited as an X-
linked dominant genetic trait. Males are usually
more severely affected than females.
Rubinstein-Taybi syndrome is characterized by short
stature, moderate to severe intellectual disability,
distinctive facial features, and broad thumbs and
first toes. It is caused by mutations in CREB-binding
protein (histone acetylation)

29. RNA interference

30. siRNA Mediated Heterochromatin
Maintenance

31. X-inactivation
• One X-chromosome in female cells is
condensed and located on nuclear
periphery
• Complete silencing of one entire
chromosome is certainly one of the
most dramatic epigenetic
phenomenon
• Key-transcript, X-inactivation specific
transport lack protein-coding function

32. • Production of Xist rapidly attract histone-
modifying enzymes, followed in turn by DNA-
methylation
• Inactive x-chromosome is evidently ‘locked
in’ by chromatin modification-
– Methylation of Histone H3 at lysine K27
– Hypoacetylation of histone at H3 ande H4
– Dense CpG methylation of CpG islands

33. Genomic imprinting
• Most autosomal genes in mammalian have
biallelic expression
• But a small number of genes are expressed
from only one parental allele – Imprinting –
become essential for viability
• Maternally derived genes are growth-limiting
• Paternally derived genes are growth-
enhancing

34. Androgenetic and gynogenetic tumor
• Benign overian teratoma (bimaternal
origin)
• Hydatidiform mole (biparenatal
origin)
• Usually benign
• Diagnosis: histopathology +
immunostaining of protein product of
imprinted gene
– Neoplastic trophoblast of complete
hydatidiform mole fails to express
imprinted gene (CDKN1C and
PHLDA2)- those normally imprinted
from maternal allele

35. Prader-willi and Angelman syndrome
• Chromosomal band 15q11-q13 is a
megabase scale chromosomal domain
including multiple imprinted gene
– One portion of gene is maternally
expressed/paternally silenced
– Adjacent portion oppositely imprinted
– Deletion of paternal homologue – PWS
– Deletion of maternal homologue - AS

36. • Whether these diseases
are due to large deletion,
microdeletion, epimutation
majority can be diagnosed
by simple Southern blot or
PCR-based assays for
DNA mutation
• Gain of methylation in
PWS
• Loss of mutation kin AS

37. Cancer epigenetics
• A common paradigm of cancer epigenetics is
hypermethylation of CpG island of tumor
suppressor gene promoter
• Hypermethylated promoter DNA is
associated with virtually every type of human
tumor
– With each type of tumor having own signature of
methylated genes

38. Cancer Methylated genes
Prostate GSTP1
Renal VHL
Colon and endometrial MLH1 (mismatch repair gene)
Esophageal APC

39. • Global hypomethylation: overall in 5-
methylcytosine content in the genome
– Found in premalignant and early stages of some
neoplasm
– Important in tumor progression
• Gene-specific hypomethylation:
– Often affect promoter region of proto-oncogene
and oncogene which are normally highly
methylated

40. DNA Methylation and Cancer

41. Technologies for Studying
Epigenetics/Epigenomics
DNA Methylation
Irizarry et. Al. (2008) Genome Research 18(5):780
Microarray or deep sequencing

42. Bisulfite Sequencing

43. 5’
3’
Expression
array
Exon array
Splicing array
Promoter
array
Tiling array
exon intron
Global Interrogation of DNA
Methylation using Microarrays
promoter

44. • Earliest method, still the most direct and least
subject to artifact is southern blotting of
genomic DNA with methylation-sensitive
restriction enzyme.
• Restriction enzymes ( e.g. HpaІІ, SmaІ, NotІ)
with CpG as part of their recognition
sequence do not cut that sequence when the
C is methylated

45. Technologies for Interrogating
Epigenetics/Epigenomics
http://www.nature.com/jid/journal/v125/n2/extref/5603467x1.jpg
ChIP-chip
Antibody specific
to one type of
histone
modification
Histone Modifications
ChIP-seq
Deep sequencing

46. Demethylation in cancer therapy

49. HDAC inhibitors

51. Interchromosome regulation of genes

52. Scaffold protein
SATB1
• More higher
structure than
chromatin
• Play important role
in T-cell
development