Chromatin structure and histone code

Question 1

what is chromatin?

Answer 1

chromosomes + associated proteins

Question 2

what are the four levels of packaging?

Answer 2

1st : DNA + histones = nucleosomes
(beads on a string)

2nd : nucleosomes pack themselves into fibres of 30nm (solenoid)

3rd : 30nm fibres pack themselves into 80-1–nm fibres

4th : represented by the mitotic chromosomes

Question 3

what do chromosomes consist of?

Answer 3

• DNA
• histone proteins
• non-histones proteins
• non-coding RNA

Question 4

nucleosomes

Answer 4

• histones assemble to form an OCTAMER
• this is 2 molecules of each histones (H2A, H2B, H3 and H4) with the DNA molecule wrapped around the structure
• the N-terminal tails of the histone proteins are +ve charged and stick out of the octamer structure

Question 5

compaction of nucleosomes

Answer 5

compaction of nucleosomes occurs to form higher order structures which involves:

• linker histones (H1) link together octamer cores (nucleosomes) to form a solenoid (30nm fibre)
• packaging proteins bind to histones tails to bring them together

Question 6

what are histone remodelling factors?

Answer 6

• enzymes that remove and replace nucleosomes

Question 7

euchromatin

Answer 7

• lightly staining areas of chromatin
• less DNA
• rich in genes
• made up of nucleosomes, but not dense higher order packaging (low order of packaging)

Question 8

heterochromatin

Answer 8

• darkly stained areas
• more DNA
• few genes (most of the DNA is “useless” to the specific cell)
• dense higher order of packaging (has a lot of DNA to pack) of nucleosomes

Question 9

facultative heterochromatin

Answer 9

• contains genes that are not expressed in that cell type
• DNA tightly packaged as heterochromatin
• BUT it may be packaged as euchromatin in other cell types as other cell types may need to express those certain genes

Question 10

what determines whether nucleosomes are packed as HETEROCHROMATIN or EUCHROMATIN?

Answer 10

the type of cell

Question 11

constitutive heterochromatin

Answer 11

• there is a section of heterochromatin that will NEVER become euchromatin as the cell will NEVER need those genes to be expressed

Question 12

EXAMPLE: the production of a beard

Answer 12

• pre-puberty, skin cells on the face will not express the genes for hair follicles, this gene will be packaged under the category of “FACULTATIVE heterochromatin”
• post-puberty, chemicals (testosterone) will activate the genes for hair follicles and will therefore become euchromatin
• this will change the heterochromatin to allow the expression of the hair follicles for a beard to grow
• mashallah boys

Question 13

what chemical modifications take place to convert heterochromatin to euchromatin and vice versa?

Answer 13

• methylation —-> coverts euchromatin to heterochromatin

- acetylation ——> converts heterochromatin to euchromatin

Question 14

what are the enzymes that allow chemical modification of histone tails to take place?

Answer 14

• HAT –> histone acetyl transferase

- HMT –> histone methyl transferase

Question 15

how does chemical modification occur?

-acetylation/methylation

Answer 15

1. TF bind to DNA
2. recruitment of histone modification enzymes (HAT/HMT) via N-CoA (nuclear coactivator)
3. this leads to unpackaging of DNA

Question 16

what is acetylation?

Answer 16

• the loss of +ve charge from the histone tails

Question 17

what is methylation?

Answer 17

• addition of a methyl group to DNA

Question 18

HATs

Answer 18

• histone acetyl transferase
• acetylate lysine residues (AA) on histones
• leads to unpacking of chromatin

Question 19

HDACs

Answer 19

• histone deacetylases
• De-acetylate histones
• lead to compaction of chromatin

Question 20

EXAMPLE : thyroid hormone receptors

Answer 20

1. thyroid hormone receptor (TR) binds to the thyroid response element (TRE) on the DNA strand
2. this recruits (HDAC/DM (de-methylase)) via the N-CoR (nuclear corepressor) which causes De-methylation
3. Then we, recruit HAT and HMT via the N-CoA (nuclear coactivator), which then causes acetylation, so the DNA unwinds even more (making euchromatin).
4. RNA Pol II binds and transcribes thyroid genes.

Question 21

nuclear co-repressor (N-CoR)

Answer 21

-aids in the recruitment of HDAC and DM to remove chemical groups from DNA

Question 22

nuclear co-activator (N-CoA)

Answer 22

• aids in the recruitment of HMT and HAT to add chemical groups to DNA

Question 23

Histone modification via methylation

Answer 23

• histone tails are methylated by histone methyl transferase (HMTs)
• De-methylated by histone demethylase (HDMs)
• a lysine residue can be mono, di or tri methylated

Question 24

Loops and chromatin domains

Answer 24

There is evidence that suggests
- Each loop may have a different degree of chromatin compaction
- The scaffold isolates the chromatin in one loop from the next loop
- So one loop may have open chromatin and active genes
The neighbouring loop could be tightly packed as heterochromatin

Question 25

Methods to investigate chromatin structure- DNAse digestion

Answer 25

DNAse I cuts double stranded DNA - HYPERSENSITIVE SITES (HSS). They are:

• Sequences of DNA without histones
• May be naked DNA, or binding transcription factors
• Cut by very brief digestion with DNAse I
• Found in promoters and enhancers

-Histone binding protects DNA from DNase digestion

Question 26

Marks for promoters and enhancers

Answer 26

• Promoters strongly enriched for H3K4me3

- Active enhancers enriched for H3K4me1

Question 27

Histone code

Answer 27

• Histone “marks” are read by binding proteins
• Related domains are found in multiple code reading proteins e.g.
• Bromodomain proteins bind to acetylated lysines
• Tudor domain and chromodomain proteins read lysine methylation

Question 28

Examples of how methylation has varying effects.

Answer 28

• Trimethylation of histone H3 lysine at position 9 (H3K9me3) associated with heterochromatin
• Monomethylation of histone H3 lysine at position 9 (H3K9me1) usually associated with active chromatin