Epigenetics

Question 1

Which core histone proteins are in the nucleosome?

Answer 1

H2A, H2B, H3, H4 (2 of each)

Question 2

Define epigenetics.

Answer 2

Meiotically and mitotically heritable changes in gene expression that are not coded in the sequence itself.

Question 3

What percentage of mutational events in cancer are thought to be epigenetic?

Answer 3

50%

Question 4

In which two ways does chromatin provide opportunities to regulate gene expression?

Answer 4

1. The folding of the chromatin can be regulated, controlling the access of proteins to the DNA eg. nucleosome remodelling enzymes.
2. Post-translational modifications of the chromatin control DNA accessibility and transcription factor and RNA polymerase recruitment.

Question 5

What are the three main methods of post-translational modification of core histone tails?

Answer 5

Acetylation
Phosphorylation
Methylation

Question 6

What effect does acetylation have on the lysine residue?

Answer 6

Neturalises the positive lysine residue.

Therefore the histone tail does not interact with linker DNA so well and contributes to chromatin unfolding.

Question 7

What recruits histone acetyl transferases and histone deacetylases?

Answer 7

Transcription factors.

Question 8

What type of protein complexes do acetylated lysines recruit?

Answer 8

Protein complexes with bromodomains, e.g.:
Nucleosome remodelling complexes - SWI/SNF
Co-activators - CBP/P300

Question 9

What does assembly of the pre-initiation complex allow to happen?

Answer 9

Phosphorylation of the polymerase C-terminal domain. (CTD)

Question 10

What does CTD phosphorylation allow to happen?

Answer 10

Allows the polymerase holoenzyme to clear the promoter and initiate transcription.

Question 11

Which type of proteins carry out lysine methylation?

Answer 11

SET domain proteins.

Question 12

Which molecules are important for transcriptional silencing and heterochromatin formation?

Answer 12

HP1

Polycomb complexes.

Question 13

What do HP1 and polycomb complexes bind?

Answer 13

They bind H3Kme marks using their chromodomains.

Question 14

What do remodelling complexes do?

Answer 14

Slide or loosen nucleosomes to allow access to the underlying DNA.

Question 15

Give one way of studying histone modifications in vivo?

Answer 15

ChIP

Question 16

How does ChIP work?

Answer 16

Treat cells with formaldeyhyde to crosslink chromatin.
Sonicate to break chromatin into short fragments.
Mix with antibody.
Immunoprecipitate with protein A beads.
Dissociate with strong detergent.
Purify DNA.
Detect sequence of interest by Southern blotting, PCR, Q-PCR, or NGS.

Question 17

Name a transcription factor essential for haemopoeisis.

Answer 17

TAL1 (SCL)

Question 18

What percentage of CpG dinucleotides in human somatic cells are subject to cytosine methylation?

Answer 18

Approximately 80%

Question 19

Give 2 ways DNA methylation leads to transcriptional silencing.

Answer 19

1. Methylation of a transcription factor binding site can prevent binding of the factor to the DNA.
2. DNA methylation combines with certain histone modifications and repressor proteins to form a repressive chromatin structure.

Question 20

What is Me-CpG recognised by?

Answer 20

Me-CpG binding domain (MBD) proteins.

Question 21

What can MBD proteins recruit?

Answer 21

HDACs and HMTs (e.g. Suv39h)

Question 22

What does Suv39h recruit?

Answer 22

DNA methyl transferases (DNMTs) that methylate DNA.

Question 23

What does H3K9me recruit?

Answer 23

HP1, leading to transcriptional silencing.

Question 24

What percentage of human gene promoters are CpG islands?

Answer 24

60%

Question 25

Which gene is hypermethylated and silenced in retinoblastomas?

Answer 25

Rb

Question 26

Which gene is hypermethylated and silenced in Breast cancer?

Answer 26

BRCA1