Boyes Epigenetics

Question 1

What do epigenetic factors do?

Answer 1

Bring about an inheritable change in expression of a gene without introducing mutation

Question 2

What is the structure of a nucleosome?

Answer 2

H3/4 tetramer, 2xH2A/B dimers, H1 at entry and exit points.
147bp wrapped around with dyad 80p in
50 nucleotide linker

Question 3

How many times does DNA wrap around the histone?

Answer 3

1.65 turns

Question 4

What is the structure of a histone protein?

Answer 4

High R/K proportion, 2alpha helices conected by loops

Question 5

How does the nucleosome remodelling complex work?

Answer 5

DBD binds entry point to push, forming loop
translocation domain binds at the dyad to pull 10bp via hinge
uses ATP

Question 6

What increase in DNase1 hypersensitivity is found at promoters?

Answer 6

100x

Question 7

How can promoters be mapped?

Answer 7

DNase digestion at promoters then restriction digests at known points, to find promoters based on size of fragment. Known as indirect end labelling

Question 8

What is indirect end labelling?

Answer 8

Method of finding promoter sites

Question 9

What additional functions do nucleosome remodelling complexes have?

Answer 9

spacing following replication
exposure of DNA
exchange of H2A/B dimerss

Question 10

How are nucleosome remodelling complexes classified?

Answer 10

By ATPase subunit

Question 11

What are the classes of remodelling complex?

Answer 11

SNI/SWF, ISWI, CHD, INO80

Question 12

What is the structure of nucleosome remodelling complexes?

Answer 12

2-15 subunits, up to 2MDa
ATPase and helicase domains
other targetting domains

Question 13

How can nucleosome remodelling complexes be recruited to a promoter?

Answer 13

Via activators or direct to chromatin

Question 14

Where is the ovalbumin gene activated?

Answer 14

Oviduct

Question 15

Where is the vitellogenin gene stimulated?

Answer 15

By oestrogen in the liver

Question 16

How are X chromosomes inactivated?

Answer 16

DNA methylation of CpG islands in promoters

Question 17

Does epigenetic memory exist in Drosphilia flies?

Answer 17

Yes, even though they do not have DNA methylation

Question 18

What timescale can epigenetics occur on?

Answer 18

Accumulate over a lifetime

Question 19

What methylation state does newly replicated DNA exist in?

Answer 19

Hemi-methylated

Question 20

Which Dnmt is responsible for methylation after replication?

Answer 20

Dnmt1

Question 21

Which enzyme is responsible for DNA methylation de novo?

Answer 21

Dnmt3a/b

Question 22

When does Dnmt3 act?

Answer 22

During embryonic development

Question 23

What is the structure of Dnmt?

Answer 23

Highly conserved catalytic C terminal domain

N terminal binds to other proteins

Question 24

When is Dnmt1 recruited?

Answer 24

PCNA clamp association at replication fork

Question 25

When is Dnmt3 not recruited?

Answer 25

When 3L regulatory unit dissociates

Question 26

How are CpG islands protected from methylation?

Answer 26

Transcription by Pol 2 Ser5 recruits Set1 and Cfp1 to promoter H3K4me3 opens chromatin structure no association of 3L to dnmt3

Question 27

Which novel proteins can be isolated from a promoter?

Answer 27

Set1 and cfp1

Question 28

How can Set1 and Cfp1 be isolated from promoters?

Answer 28

HinP1 methylation sensitive digestion of GCGC sites

Question 29

What is the structure of CpG islands?

Answer 29

1kb streches of G rich

Question 30

Where are CpG islands found?

Answer 30

All housekeeping genes and 40% of tissue specific genes

Question 31

How many CpG islands are the hapliod human genome?

Answer 31

25,500

Question 32

What frequency are CpG islands?

Answer 32

1/10

Question 33

What type of CpG islands have differential expression?

Answer 33

Weak promoters 1/20

Question 34

What CpG frequency has no effect on transcription?

Answer 34

1/100

Question 35

How does methylation repress transcription?

Answer 35

Directly by blocking major groove binding sites or indirectly by binding MBPs. MECP1/2 has HDAC/Sin3a activity

Question 36

When does CpG island methylation occur?

Answer 36

X chromosome inactivation and cancerous mutations

Question 37

How can CpG islands be detected?

Answer 37

Bisulphite sequencing deaminates C to U, so only C left for sequencing are mC Digestion with Msp1 (non specific) and Hpa11 (unmeth)

Question 38

What is the histone code hypothesis?

Answer 38

Predicts that distinct combinations of modifications act to control chromatin structure and gene expression

Question 39

Where do histone modifications act?

Answer 39

Exposed Lysine tails

Question 40

Where does acetylation occur?

Answer 40

H3K14 binds bromodomains | H3K16 prevents compaction to a 30nm fibre

Question 41

What type of proteins have bromodomains?

Answer 41

Coactivator/HATs associated with Pol2 elongator form | Remodelling complexes SWI/SNF and RSC which displaces H2A/B dimers

Question 42

How is acetylation removed?

Answer 42

HDACs recruited after the Pol2 has passed

Question 43

How can acetylation be observed?

Answer 43

Removal of nucleosomes alters sedimentation in centrifugation Increased DNas1 sensitivity

Question 44

How does acetylation activate chromatin?

Answer 44

Neutralises DNAphos - K/R interactions

Question 45

Is acetylation epigenetic?

Answer 45

NO

Question 46

How is H3K4me3 produced?

Answer 46

Set1 methyltransferase associated with Pol2 CTD phospho-ser5 at promoters

Question 47

What type of domain does H3K4me3 recruit?

Answer 47

plant homeodomain (PHD) in remodelling complexes and HATs

Question 48

Which histone methylations are activating?

Answer 48

K4, K36

Question 49

Where is H3K4me1 found?

Answer 49

Active enhancers

Question 50

Where is H3K36me3?

Answer 50

In gene body, associated with Set2 (Ser2)

Question 51

How does H3K36 act?

Answer 51

Recruits HDACs to reset chromatin after transcription

Question 52

What type of methylation does HP1 oproteins bind?

Answer 52

Chromodomains bind H3K9

Question 53

How is H3K9 maintained?

Answer 53

Chromodomains of HP1 bind and recruit Suv39h set domain to act on adjacent nucleosomes to spread heterochromatin

Question 54

Where can H3K9 be observed?

Answer 54

Position Effect Varigation in Droshphilia eyes by silencing in some cells producing patches of colours

Question 55

What type of protein is Set1?

Answer 55

A trithorax protein

Question 56

Which methylation do Polycomb group proteins act on?

Answer 56

H3K27me3

Question 57

How do polycomb group proteins act?

Answer 57

PRC1 chromodomain binds H3K27me3 producing compaction | Recruits PRC2 Ezh SET to act on adjacent nucleosomes

Question 58

What does H3K27me3 do?

Answer 58

Repression by compaction into heterochromatin. Hox genes silencing and X chromosome silencing

Question 59

How is histone methylation maintained epigenetically?

Answer 59

Asf1 chaperones move Old H3/4 tetramers during replication, modifying enzymes stay attached to act on new tetramers brought by Caf1

Question 60

How much heterochromatin is maintained epigenetically?

Answer 60

75% has HP1 or PRC proteins

Question 61

Is Set a conserved sequence?

Answer 61

No