Epigenetics and cancer

Question 1

What are epigenetic traits?

Answer 1

Stable, heritable phenotypes resulting from changes to chromatin structure without alterations to the DNA sequence

Question 2

What alters the chromatin?

Answer 2

Chromatin remodelling proteins-

they compress or open up the chromatin

Question 3

How do epigenetics modifications result in a different phenotype?

Answer 3

Alterations of chromatin structure results in a change in gene expression
this results in an altered phenotype

Question 4

How are chromatin remodelling proteins regulated?

Answer 4

In a signal regulated way

Question 5

What are the 2 main impacts of epigenetic modifications?

Answer 5

1. They control the accessibility of target genes to incoming transcription machinery e.g RNA polymerase 2
2. They directly control the biochemical activity of transcriptional machinery itself

Question 6

What are the 2 main histone modifications- what do they cause?

Answer 6

Methylation and acetylation
either:
conversion of genes into active domains of transcribable chromatin
or
inactive genes of transcriptionally silent chromatin

Question 7

What 3 things must occur for a proliferating progenitor cell to become a differentiated cell?

Answer 7

1. Suppression of cell renewal
2. Lineage commitment
3. One programme of differentiation pursued

Question 8

Where are modifications made on histones?

Answer 8

The N terminal tails of each of the 4 substituent histone components

Question 9

What are the 4 components of a histone?

Answer 9

H2A
H2B
H3
H4

Question 10

What is a nucleosome?

Answer 10

DNA wound around 8 histone protein cores-

building block of chromatin

Question 11

What are the covalent modifications made to core histones?

Answer 11

Acetylation- of lysine tails
Methylation- of lysine and arginine tails
Phosphorylation- of serine 10 in histone H3

Question 12

What is the relationship between acetylation and methylation?

Answer 12

They are competing reactions

both can occur on lysine tails

Question 13

What are HATs and HMTs? What is their main difference?

Answer 13

HATs= histone acetyltransferases
HMTs= histone methyltransferases
HATs can modify many different lysine residues whereas HMTs are much more site specific

Question 14

Which histone methylation modifications are associated with activation and repression?

Answer 14

Dependent on the modification and where it was made
Histone lysine 4 methylation= activity
Histone lysine 27/9= inactivity- repression

Question 15

What is acetylation predominantly associated with?

Answer 15

Transcriptionally active chromatin

Question 16

What is the significance of histone modifications?

Answer 16

They act as recruitment signals- they recruit parts of the transcription machinery

Question 17

What does acetylation recruit?

Answer 17

Proteins that contain an acetyl-lysine binding domain called a bromodomain

Question 18

What does methylation recruit?

Answer 18

Proteins that contain either a chromodomain (REPRESSORS) or a PHD zinc finger domain (ACTIVATION)

Question 19

What happens when genes in the ‘polycomb group gene enhancer of zeste’ are mutated?

Answer 19

Extra sex combs develop on the drosophila

normally just the forelegs

Question 20

What does enhancer of zeste code for?

Answer 20

A HMT of lysine 27- involved in converting a homeotic complex to inactive- transcriptionally repressed
when its mutated= lose the inhibition and thus develop more sex combs

Question 21

The polycomb group of proteins code for various proteins- what are the main ones?

Answer 21

Code writers- components of a complex in which EZH2 methyltransferase lies
Code readers- proteins of a complex that a chromodomain containing protein binds
The group codes for a chromatin regulatory system- which marks chromatin for silencing and makes it transcriptionally inert

Question 22

What happened when they isolated the polycomb protein genes in vertebrates? What is the relevance with prostate cancer?

Answer 22

The polycomb family of proteins are frequently and highly expressed in cancers
In normal prostate and advanced prostate cancer-
immunostaining showed that there were increased levels of EZH2 in the advanced prostate biopsies

Question 23

How can you study metastasis in cells?

Answer 23

Use a matragel membrane- punch small holes into it

invasive cells will try and squeeze through the holes

Question 24

What happens in you KO EZH2 in cells and study their ability to invade? What can be concluded?

Answer 24

Much less invasion

EZH2 is required for the metastatic phenotype

Question 25

What is EZH2 also required for?

Answer 25

Cell proliferation

Question 26

What is the mutant EZH2 protein like? Where are mutations in EZH2 common?

Answer 26

Hyper-active protein which makes it constitutively active- hyper-methylating through the genome non-Hodgkin's lymphoma- EZH2 is the most frequently modified chromatin modifying gene

Question 27

What else is characteristic of mutant EZH2 proteins? How is this being as a result?

Answer 27

The proteins are hypersensitive to a class of small molecule can make specific inhibitors which target mutant EZH2 and have relatively no effect on WT EZH2 kills mutant lymphoma cells

Question 28

How does EPZ affect mutant EZH2?

Answer 28

It prevents EZH2 from methylating histone lysine 27 (in a concentration dependent manner) Inhibits proliferation of EZH2 mutated cells- cancer specific inhibitor (conc dependent)

Question 29

What are the 2 point mutations associated with mutant EZH2?

Answer 29

Tyrosine to a phenylalanine | Alanine to a glycine

Question 30

What can aberrant crossovers due to chromosomal translocations (between non homologous chromosomes) cause?

Answer 30

The creation of hybrid genes which code for hybrid proteins- which are characteristic of many lyphomas and leukaemias

Question 31

What are the 2 cross overs associated with acute myeloid leukaemia and acute lymphoblastic leukaemia?

Answer 31

Cross over between chromosomes 8 and 16- between 2 HATs- AML Cross over between chromosomes 11 and 16- between a HMT and HAT- ALL Both of these translocations create fusion proteins that make a population of immortalised haemopoietic stem cells

Question 32

What are the 3 main domains of the MLL protein?

Answer 32

Catalytic domain- allows it to make methylation lysine 4 marks Interaction domain- can bind to HATs PHD finger domain- can bind to histone lysine 4 when its methylated

Question 33

Whats the main role of MLL?

Answer 33

To be able to recognise methylated histone lysine 4 and propagate new methylation marks on adjacent nucleosomes =make a transcriptionally active DNA domain

Question 34

Following a leukemogenic translocation- what is lost (and retained) in the MLL protein?

Answer 34

Lost- domains involved in histone lysine 4 methylation and recognition Retains its transcriptional repressor domain

Question 35

What can be suggested about the fusion MLL proteins new function?

Answer 35

MLL exerts its leukemogenic function by acting as a dominant negative inhibitor of its WT counterpart It compromises the ability of target genes to be methylated- and thus activated