Epigenetic diseases + diagnostics therapies

Question 1

What are Methyl-CpG binding proteins and what do they share?

Answer 1

They are ‘readers’ of modified cytosines and interact with modified, CpG-dense regulatory regions. They share a MBD domain.

Question 2

Genetic syndromes - mutations in ‘epigenetic’ genes

Question 3

What causes Rett Syndrome?

Answer 3

When the MECP2 gene is mutated and this leads to a non-functional MECP2 (Methyl CpG binding protein). This causes the expression of the mutated allele and the normal allele, usually expressed, to be repressed.

Question 4

ICF syndrome is caused by?

Answer 4

Mutations in DNMT3B leading to hypomethylation of DNA on the inactive-X chromosome (CpG islands)

Question 5

What causes ATR-X syndrome?

Answer 5

Mutated ATRX gene which encodes DNA helicase. It is an X-linked recessive gene and involved in chromatin remodelling.

Question 6

What is Uniparental Disomy (UPD)?

Answer 6

Where an individual inherits two copies of a chromosome from their parent or mother, and none from the other parent.

Question 7

Which chromsome and region is Prader-Willi Syndrome?

Answer 7

15q11-13
q = chromosome
11-13 = region

Question 8

What is the most common defect?

Answer 8

65-75% have a deletion at the PWS region on their paternal chromosome 15.

Question 9

25% have maternal UPD. 5% have a mutation in the genomic region that controls imprinting.

Question 10

What are some symptoms of PWS?

Answer 10

Insatiable appetite/morbid obesity in later childhood. Small hands and feet, short stature, hypogonadism. 1/10,000 - 1/25,000 live births.

Question 11

What is the association between ART and inmprinting disorders which cause PWS?

Answer 11

Imprinting disorders may take place just after fertilisation at a time where the epigenome is most vulnerable. Differentially Methylated Regions (DMRs) are abnormally methylated.

Question 12

What is similar with Angelman Syndrome to PWS?

Answer 12

It is also 15q11-q13, but on the maternal chromosome

Question 13

What is the most common defect?

Answer 13

65-75% have a deletion at the AS region on their maternal chromosome 15.

Question 14

5% have paternal UPD. 5% have a mutation in the genomic region that controls imprinting.

Question 15

10% have a mutation in the UBE3A gene, why is this difficult to correct?

Answer 15

Because the paternal UBE3A gene (allele) is normally silenced by imprinting.

Question 16

Ubiquitin-protein ligase - UBE3A is an important gene in mice (and humans) for controlling what?

Answer 16

It affects the circadian activity and metabolism

Question 17

How can it be treated?

Answer 17

Pharmacological reversal of the silencing rescues functional circadian physiology clock properties.

Question 18

How can sleep disorders associated with Angelman Syndrome be treated?

Answer 18

Chronotherapy

Question 19

Shows the same genomic region between PWS and AS, but different phenotypes.

Question 20

q = long chromosomal arm
p = short chromosomal arm

Question 21

Beckwith-Wiedemann Syndrome is caused by?

Answer 21

Both IGF-2 genes are expressed leading to increased levels of IGF-2 (growth factor). Normally, one of the alleles is inactive.

Question 22

What does IGF2 promote?

Answer 22

Embryonic growth. If in excess, it can lead to a 1000-fold higher risk of tumours

Question 23

What is the other gene involved in Beckwith-Wiedemann Syndrome which when repressed, can cause the syndrome?

Answer 23

CDKN1 gene which suppresses embryonic growth.

Question 24

So excessive IGF2 gene expression and no CDKN1 expression causes Beckwith-Wiedemann Syndrome, what is the opposite?

Answer 24

Both CDKN1 alleles are expressed and no IGF2 exprssed leading to Silver Russell Syndrome.

Question 25

Double allele expression in Silver Russel syndrome can cause a specific mutation, what is it?

Answer 25

Excessive production of brown adipose tissue which generates too much heat.

Question 26

For Wilms’ tumour (type of kidney cancer) is caused by cells losing their epigenetic memory. What does this lead to?

Answer 26

LOI - Loss Of Imprinting
Both alleles for IGF2 are expressed, leading to excessive growth = cancer tumours.

Question 27

What are two dietry essentials which when in lack of abundance, leads to epigenetic diseases?

Answer 27

Folic acid & Methionine

Question 28

Explain each

Answer 28

Folic acid = critical for DNA synthesis and DNA methylation. Folate deficiency leads to neural tube defects in babies, and depression in adults.

Methionine = Precursor of methyl groups needed for DNMTs. Levels affect the availability of SAM as a donor of methyl groups.

Question 29

Explain how deamination of meCpG can lead to cancer?

Answer 29

Deamination of a methylated cytosine will turn it into a thymine. When it is a thymine, the DNA repair machinery is unable to excise the thymine out of the sequence. Whereas if it the cytosine was unmethylated, then it forms uracil, and the uracil bases are excised out of the sequence.

Question 30

What is another possible cause of mutation to a methylated cytosine?

Answer 30

UV light

Question 31

Why would you screen for DNA methylation of saliva, blood, urine etc?

Answer 31

A normal epithelium can hyperproliferate to an adenoma (non-cancerous tumour) to class 1-3, then carcinoma, and then metastasis (cancer spread from primary site) if DNA methylation is increasing. So, screening of these samples can determine an altered epigenome.

Question 32

Why is understanding epigenetic changes in early tumourigenesis more hopeful than identifying mutational drivers of metastasis?

Answer 32

Because epigenetic changes are reversible - can reverse the state of a cell back to a normal state.

Question 33

What is one of the first detectable epigenetic modifications in cancer?

Answer 33

An increased DNA methylation of tumour suppressor genes.

Question 34

Where are the two locations where CpG islands being methylated is normal and needed?

Answer 34

1. Imprinted genes
2. X-linked genes silencing

Question 35

Explain the paradox in tumours for global hypomethylation & hypermethylation of CpG islands

Answer 35

• DNMT1 (maintenance) expression is low
• DNMT-3b (de novo methylation of CpG islands) is increased

Question 36

MGMT & GSTP1 are both genes used as predictors of cancer. Explain both of their roles and why it is useful in understanding when they are methylated.

Answer 36

MGMT - Needed for DNA repair of damaged DNA. It is needed for dealing with the entry of toxins. Brain biopsy to determine hypermethylation

GSTP1 - Leads to tumours within the urine system. Biopsy can help understand an effective treatment for hypomethylation tailored to the patient.

Question 37

What is the fundamental aim of epigenetic drugs and two examples?

Answer 37

To reactivate silenced genes or prevent repression of active genes. ZEB & SAHA

Question 38

Simply, how does bisulfite treatment work and why?

Answer 38

Treat DNA with bisulfite where unmethylated cytosines are converted to uracil. Methylated cytosines remain as cytosine. DNA sequencing and uracil will form a thymine residue, showing that there is methylation of cytosines.

Question 39

Can compare DNA methylation arrays % data anyalysis between different individuals which shows different methylation levels.

The DNA methylation maps can highlight aberrant methylation, which act as biomarkers for cancer. CpG sites may be hyper/hypomethylated for diseases.

Question 40

What is meant by the Epigenetic clocks?

Answer 40

Methylation levels of CpG sites within the genome correlate with chronological age. If these sits are methylated earlier than expected, it shows an accelerated biological age, even though their chronological age is younger (and vice-versa).

Question 41

If this epigenetic clock is ahead of the chronological clock, pharmacological/dietry interventions can prevent early onset of diseases by decelerating the epigenetic processes as they’re reversible. There can be environments which slow/speed up biological age compared to chronological age. Microarrays can determine the methylation sites as mentioned previously above.