Epigenetics

Question 1

What is epigenetic?

Answer 1

Heritable changes (between cell lines, not generations) that are not caused by changes in DNA sequence

Question 2

What are the normal functions of epigenetics?

Answer 2

Cell differentiation
X chromosome inactivation
Embryonic development
Gene imprinting

Question 3

What are the two epigenetic modifications?

Answer 3

DNA methylation

Histone Modification

Question 4

How does DNA methylation work?

Answer 4

Addition of a methyl group on to the 5 position of a cytosine in a CpG site - forms 5-MC
Catalysed by DNMT1/3a/3b
DNMT 1 - inheritance - affinity for semi methylated DNA
DNMT 3a/3b - de novo - equal affinity for hemi and non methylated DNA - supported by DNMT 3L
Doesn’t occur in CpG islands - errors can lead to CANCER

Question 5

How does DNA methylation affect gene expression?

Answer 5

1) Methylation of CpG sites in TF affect binding to the DNA (not all TFs have CpG sites)
2) Methylation of DNA causes recruitment of 2 methyl binding protein complexes - MBD2 and MeCP2 - contain HDACs which inactivate DNA (conversion to heterochromatin)

Question 6

What other sites can DNA methylation occur at?

Answer 6

CpA, CpT (embryonic stem cells), CpC

Question 7

What are the different forms of histone modification?

Answer 7

Acetylation - Lysine - HAT/HDAC - affect charge - use Acetyl CoA as cofactor

Methylation - Lysine - HKMT with SAM (cofactor) - K27/K9 (H3) inactive - K4 (H3) - active - Arginine - RME Type 1 and Type 2 - use SAM as cofactor

Phosphorylation - Serine/Threonine/Tyrosine - Kinase/phosphotase - transfer ATP to OH group of AA - introduce negative charge which affects binding of chromatin

Question 8

What is epigenetic drift?

Answer 8

With age, our epigenetic change:

• Increase in methylation within CpG islands
• Reduction of methylation in global genome
• Increase in mutations within CpG sites (5-MC more prone to mutation)

Question 9

How can DNA methylation be linked to cancer?

Answer 9

High methylation within promoter sequences of Tumour suppressor genes (CpG islands) leads to suppression of expression
Reduced global methylation - reduced genome stability - cell transformation
Increased mutations within CpG sites

Question 10

What techniques have we used to identify DNA methylation links with cancer?

Answer 10

Bisulfite modification - allows for study of methylation across multiple genes - indentified CIMP (CpG Island methyl Phenotypes) and Type A/C genes

Question 11

What drug can we use to target DNA methylation with the aim of treating cancer?

Answer 11

Azacitidine - DNA methyltransferase inhibitor - Myelodysplastic syndrome

Question 12

What are the two classes of HDAC?

Answer 12

Zn dependent - Class 1,2,4

NAD dependent - SIRTS

Question 13

What is an example of an HDACi?

Answer 13

Zolinza - Cutaenous T Cell Lymphoma - targets active site of HDAC and chelates Zn ions - targets Class 1, 2, 4

Question 14

What environmental effects can influence epigenetic?

Answer 14

Diet (Folate, HFHS, med diet)
Smoking
Endocrine disruptors (Bisphenol A, vinclozinin, phthalates)
Behaviour (PA, maternal grooming, early life trauma)

Question 15

How can diet influence epigenetics?

Answer 15

Folate - SAM - retro transposal element - agouti mice experiment
HFHS diet - autonomic NS impact (CVD) - reduction in ACE methylation - increase in ACE 2 gene expression - increase in autonomic NS dysfunction - CVD
Med diet - study giving mothers omega tablets - using bisulphite sequencing found that children had increased IGF2 hypermethylation - IGF2 increase in seen in diabetic patients

Question 16

How can smoking influence epigenetics?

Answer 16

Smoking can affect epigenetics in both adult life (over 2500 CpG sites - CVD, obesity etc) and during pregnancy (over 500 CpG sites) - can lead to problems such as LBW, SIDS and addiction later in life

Thought to cause effect from cigarette smoke - damages DNA (double strand breaks) - leads to altered DNA methylation patterns. Also thought to impact on DMT1 - affecting inheritance of methylation patterns. Also increase in smoking smoke increases hypoxia and this leads to recruitment of more SAM (cofactor for DNA methylation) so results in increased methylation

Question 17

How can endocrine disruptors influence epigenetics?

Answer 17

Bisphenol A (plastic manufacture) - exposure during sensitive periods (in uterus, infancy, adolescence) can lead to cancer, obesity though impact on DNA methylation

Phthalates - plastic manufacture - 300 children cohort study looking at parental exposure to this before pregnancy and risk of hypospadias - found that paternal occupation influenced risk of hypospadias, with those exposed to high Phthalates having higher hypospadias risk.

Vinclozinin (fungicides) - shows heritability of epigenetic changes - pregnant mice - passed on to foetus and germline cells - affects sperm motility

Question 18

What are the classes of HDAC?

Answer 18

18 in human - HDAC 1-11 (Zn dependent - Class 1, 2, 4) - SIRT (1-7 - NAD dependent)

Question 19

What studies have shown an associated between ageing/chronic disease and HDAC?

Answer 19

C.elegens - increase of SIR2 (homologue of SIRT1) increased lifespan
Drosophila - resveratrol (increases activity of SIRT2) increases lifespan
Drosophila - caloric restriction (increases SIRT2) increases lifespan
Mice - caloric restriction associated with health benefits (heart disease, obesity, liver disease, mood, brain injury recovery, diabetes)
Mice - caloric restriction alleviated symptoms of machado-joseph syndrome (CNS disease with muscular degeneration and loss of control)

Question 20

What is the impact if we give someone a HDACi for treatment of cancer (such as Zolinza)?

Answer 20

Reduces cancer - increased ageing - reduced lifespan - alleviates other chronic diseases such as liver disease, heart disease and diabetes.