Chromatin and Histone Code

Question 1

What is chromatin?

Answer 1

Nuclear complex of DNA and associated proteins that forms chromosomes within the nucleus of eukaryotic cells.

Question 2

When can chromatin be seen?

Answer 2

Chromatin can be seen during interphase when it is organised into euchromatin and heterochromatin

Question 3

How is chromatin eleasticity enabled?

Answer 3

By choice of histone variants, modifications of DNA bases, and reversible post-translational modifications (PTM) of histone tails.

Question 4

How is DNA equally partitioned into both daughter cells?

Answer 4

DNA becomes highly compacted into the classic metaphase chromosomes that can be seen with a light microscope.
2.Once a cell has divided, its chromosomes uncoil again.

Question 5

Describe euchromatin(5)

Answer 5

· Low compacity, 10 nm fibre- beads on a string
· Lightly staining areas of chromatin
· Rich in genes
· Euchromatin is accessible to the enzymes involved in DNA transcription, replication, or repair.
· Euchromatin contains regions of DNA that are transcriptionally active ·

Question 6

Describe heterochromatin(4)

Answer 6

· Highly condensed, 30 nm fibre
· Darkly staining areas of chromatin often associated with nuclear envelope
· Gene poor
· Constitutive and Facultative

Question 7

Describe the difference between facultative and constitutive heterochromatin?

Answer 7

Facultative: →contains genes that are not expressed in that cell type.

→ DNA is as tightly packaged as constitutive heterochromatin, but it may be packaged as heterochromatin in other cell types.
Constitutive:→ remains condensed throughout the cell cycle and development.

→contains highly repetitive sequences that are not transcribed and play a role in chromosome structure.

Question 8

Describe the 1st level of packaging

Answer 8

“st level packaging- DNA+histones = nucleosomes Increases DNA packaging 7-fold ≈10nm

Question 9

Describe the 2nd level of chromatin packaging

Answer 9

“Nucleosomes pack themselves in fibers. Increases DNA packaging 6-fold 30nm”

Question 10

Describe the 3rd level of packaging in chromatins

Answer 10

· Fibres pack themselves into loops and TADs and form chromatin. Increases DNA packaging 3-fold. 100-250 nm

Question 11

Describe the 4th level of packaging in chromatins

Answer 11

· Represented by the mitotic chromosome. 1000-fold packaging 700-1000nm. Visible during mitosis due to condensing of chromosomes.

Question 12

What are TADs?

Answer 12

· Topologically Associating Domains (TADs): Highly conserved chromatin domains that shape functional chromosomal organization.
Function not fully understood, their disruption lead to diseases.

Question 13

What are nucleosomes?(3)

Answer 13

· Fundamental structural unit of chromatin. Composed of a little DNA wrapped around proteins called histones.
· Exposed to post translational modification
· First level of chromatin packing is the nucleosome

Question 14

Describe the components of the nucleosome(3)

Answer 14

· 146bp of DNA wrapped around the histones core (1.7 turns of DNA).
· Linker DNA (146bps) helps communication between different nucleosomes
· The core of the octamere is composed of central H3 H4 tetramer and two flanking H2A and H2B dimers
N-terminal tails stay outside the octamer core

Question 15

What are the different types of histones found in the nucleosome

Answer 15

· H2A, H2B, H3 and H4 (2 molecules each)

Question 16

What is the role of H1 protein?(2)

Answer 16

1. H1 protein wraps another 20 base pairs of ““linker DNA”” resulting in two full turns around the octamer.
2. It helps to stabilize the zig-zagged 30 nm chromatin fiber

Question 17

How do chromatin loops occur?(2)

Answer 17

1· Occurs when stretches of genomic sequence that lie on the same chromosome are in close physical proximity to each other.
2. They are looped by cohesion rings: protein ring that binds to DNA

Question 18

What are cohesion rings and how do they form loops?(5)

Answer 18

1. The main protein is Sister Chromatid cohesion Protein
2. Cohesion rings bind to the DNA
3. The cohesion rings slide of the CTCF molecules whose binding sequence points away from the loop.
4. The loop continues to grow.
5. Loop formation when each of the rings has reached an inward directed CTCF sequence

Question 19

What are chromosomal territories?

Answer 19

Non-overlapping domains/regions of the nucleus occupied by uncondensed chromosomes. The come are composed of TADs

Question 20

Define epigentics

Answer 20

“Heritable and reversible changes in gene expression which do not involve a change in the DNA sequence.”

Question 21

Summarise the different types of modifications to DNA(4)

Answer 21

· DNA methylation

· Covalent histone modifications

· ATP dependent chromatin remodelling

· Noncoding RNAs

Question 22

Where on the histones do modifications occur

Answer 22

on the N- terminus of the histones which protrudes from the nucleosome

Question 23

Describe histone methylation and demethylation

Answer 23

1. Histone tails are methylated by histone methyl transferases (HMTs) and demethylated by histone demethylases (HDMs).
   2· methylation of lysines and arginines.

Question 24

Describe the effect of histone methylation and demethylation(2)

Answer 24

1· Methylation of some lysine residues causes chromatin condensation (e.g. H3K9me3).

2· Methylation of other lysine residues causes chromatin decondensation (e.g. H3K9me1, read as Trimethylation of lysine number nine of histone code 3)

Question 25

Describe histon acetylation

Answer 25

· Histone tails are acetylated by histone acetyl transferases (HATs) and deacetylated by histone deacetylases (HDACs).

Question 26

Describe the effect of histone acetylation

Answer 26

· Acetylated histones are generally associated with relaxed chromatin and gene expression and deacetylated histones with closed chromatin, silencing of genes.

Question 27

What are aberrant acetylation associated with?

Answer 27

· several solid tumours and haematological malignancies.

Question 28

What drugs target aberrant HDACs?

Answer 28

· HDACi as emerging drugs in cancer treatment (e.g. vorinostat (SAHA) in T-cell lymphoma).

Question 29

Describe histone phosphorylation

Answer 29

Histone tails are phosphorylated by protein kinases and dephosphorylated by proteases
· Can occur on serine, threonines and tyrosine.

Question 30

What is the effect of histone phosphorylation?

Answer 30

· Phosphorylation of H3S10 and H3S28 is involved in chromatin condensation during mitosis and meiosis, as well as in chromatin relaxation linked to transcription activation

Question 31

Describe ubiquitylation(2)

Answer 31

· Ubiquitin ligases and deubiquitinating enzymes.

· Primarily on lysines of histones H2A and H2B.

Question 32

What is the effect of ubiquitylation of H2A and H2B?

Answer 32

· H2Aub is more frequently correlated with gene silencing, while H2Bub is mostly associated with transcription activation

Question 33

What is the histone code?

Answer 33

combinations of post-translational modifications on the same histone’s tail/s

Question 34

What are code readers?

Answer 34

protein complexes that read combinations of marks

Question 35

What is the difference between chromodomains and bromodomains?

Answer 35

chromodomains specifically recognize methylated residues

while bromodomains bind acetylated residues.This read leads to chromatin remodelling (open or close)

Question 36

what determines whether nucleosomes are euchromatin or heterochromatin?

Answer 36

→chemical modification of lysine residues on histone tails:
→ acetylation
→methylation

Question 37

How is chromatin structure studied?(7)

Answer 37

→DNAse digestion.
→DNAse I cuts double-stranded DNA.
→Histone binding protects the DNA from DNAse digestion.
→there are DNAse I sensitive sites (HSS):
→sequences of DNA without histones
→it may be naked DNA or binding transcription factors
→ cut by very brief digestion with DNAse I
→found in promoters and enhancers

Question 38

How do the transcription factors open up the chromatin structure?

Answer 38

The transcription factors recruit chromatin modifying enzymes via a nuclear coactivator (NCoA) or corepressor (NCoR)

Question 39

how does the thyroid hormone receptor work?

Answer 39

→ (TR) thyroid hormone receptor binds to the thyroid response element (TRE) on the DNA.

→triggers the recruitment of the histone modification enzymes HDAC and DM via the N-CoR (which causes demethylation).

→A T3 (thyroid hormone) bonds to the TR.

→the histone modification enzymes HAT and HMT are recruited via the N-CoA (which causes acetylation, ie. the unpacking of DNA)