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BIOL200 > RR7: Chromatin, epigenetics, and the histone code > Flashcards

RR7: Chromatin, epigenetics, and the histone code Flashcards

1
Q

What is heterochromatin?

A

It’s a condensed form of chromatin that localizes at the nuclear envelope often near the nuclear pores.
Transcriptionally inactive.

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2
Q

Are the regions in the genome that are enveloped by heterochromatin transcribed?

A

No, because heterochromatin is transcriptionally inactive and the transcription of those regions could be detrimental to the cell/organism.
Those genes are silenced.

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3
Q

What is euchromatin?

A

It’s a delicate and thread-like chromatin.
Abundant in actively transcribing cells.
It’s DNA that is unwound to provide a transcriptional template.

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4
Q

Explain the structure of DNA from the double helix to the chromosome.

A

Double-stranded DNA associates with Histone proteins (H2, H3, H4) to make chromatin.
Histones together form a nucleosome.
Nucleosomes are packed together by chromatin.
Packed nucleosomes form condensed chromosomes.

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5
Q

What’s the function of a nucleosome?

A

It’s necessary to package the genomic DNA into the nucleus in the form of chromsomes.

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6
Q

Is DNA naked in the cells?

A

No, it’s associated with histone proteins, then, it’s surrounded by chromatin to form nucleosomes, and then they get packed together to from a chromosome.

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7
Q

What do transcription elements have to overcome to get the to DNA?

A

The chromatin. (euchromatin)

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8
Q

Why are the genes that form euchromatin transcribed?

A

Because they are more accessible to DNA binding, the transcription factors, RNA polymerase and the general factors.
The region is not tightly wound, like heterochromatin.

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9
Q

How do we give rise to a diploid organism?

A

When conditions are bad, a haploid yeast will mate with another haploid yeast to give rise to a diploid organism.

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10
Q

What are the mating types in Saccharomyces cerevisiae?

A

Mating type alpha
Mating type a

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11
Q

Can an alpha yeast mate with another alpha yeast?

A

No. The mating as to be a with alpha.

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12
Q

How is the mating type decided in Saccharomyces cerevisiae?

A

On one side of chromosome 3 (chromosome that controls the mating type), there’s HML alpha loci (the alpha mating type region), and on the other, there’s HMRa loci (the a mating type region).
The gene is only expressed when it’s in the middle position.
HMRa and HML alpha are silenced when they’re on their side of the chromosome.
One of them will need to be changed place to the middle to get expressed.

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13
Q

What makes that one of the 2 loci is not being moved, thus not being expressed?

A

Because of silencer sequences.

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14
Q

What regions can do transcriptional repression of a gene?

A

Silencer sequences, histones and telomeres. There’s something physically blocking the DNA, so it can’t be expressed.

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15
Q

What proteins are responsible for the silencing regions?

A

RAP1
SIR1
SIR2
SIR3
SIR4

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16
Q

Where are found the proteins responsible for silenced regions?

A

In the telomeres and in the silencer regions of the yeast (in the study)

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17
Q

What are the roles of RAP1?

A

Binds to DNA in the region of the silencer
Binds to repetitive sequence in the telomeres (recognizes the region)

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18
Q

What are the roles of SIR1?

A

Cooperates with RAP1
Important for binding the silencer region in the silent mating type loci

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19
Q

What are the roles of SIR2, SIR3 and SIR4?

A

Bind to hypoacetylated histone tails (H3 and H4)
Recruit SIR2
Form large complexes with telomeric DNA

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20
Q

What does SIR stand for?

A

Silent Information Regulator.

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21
Q

What are the steps happening for the silencing of a gene in relation to the proteins RAP1, SIR1, 2, 3 and 4?

A
  1. RAP 1 recognizes the sequence, and binds to the telomere.
  2. SIR 2, 3, and 4 come in trough protein-protein interaction, but they also recognize the hypoacetylated histone tails, and form a complex around the region where RAP 1 was.
  3. SIR 2 has enzymatic activity and it changes the histone tails to make sure they are hypoacetylated.
  4. The hypoacetylated histone tails can interact with DNA and the chromatin can get more compacted.
  5. The more hypoacetylated tails, the more the complex grows so we end up with a higher chromatin structure.
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22
Q

What’s the role of SIR2?

A

It’s a histone deacetylase.
SIR 2 recognizes the histone tails and make sure that they’re all hypoacetylated.
Once the tail is hypoacetylated, it can interact with the DNA and it tightens the winding, the chromatin gets more compacted.

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23
Q

Are the N-terminal of the histone tails structured?

A

No, they are unstructured. The residues in the tail can be modified to create specific outputs.

24
Q

How can we modify the histone tails?

A

By post-translational modifications:
- phosphorylation
- acetylation
- ubiquitination
- methylation

25
Q

What does adding acetylation of lysines on the histone tail do?

A

It opens up the chromatin.
It neutralizes the electrostatic interactions between the histone tail and the phosphate backbone of the DNA, which loosens the grip of the histone to the DNA.

26
Q

What does methylation of lysines do on the histone tail?

A

It affects transcriptional activation.
Others are associated with heterochromatin (which is transcriptionally inactive)
It’s the opposite, so methylations do not always serve the same purposes.

27
Q

What are the types of methylation events that can take place on the lysines in histone tails?

A

Methylation of H4
Methylation of H3K4
Methylation of H3K9
Methylation of H3K27
Methylation of H3K36
And it can be mono, di or trimethylations

28
Q

Which type of methylation is associated with heterochromatin?

A

H3K9.
H3K27.
It’s transcriptionally inactive.

29
Q

Which type of methylation is associated with active transcription?

A

H3K4.
H3K36
It’s associated with transcriptional activation.

30
Q

What does the methylation of H3K27 do?

A

It’s mediated by the Polycom group.
Associated with the formation of heterochromatin.

31
Q

How can we identify regions of the genome that have been affected by the histone modifications?

A

We have antibodies that can recognize the specific modifications. For example, there’s an antibody that recognizes H3K4 trimethylations. That way, we can better see where the modifications are happening.

32
Q

How can we see how the modifications are affecting the gene expression of those modified regions (histone marks)?

A

With ChIP-seq using the antibodies associated with the modified regions.
1. crosslinking agent proteins bound to chromatin
2. Isolate the agent proteins with antibodies
3. Immunoprecipitate
4. Determine the sequence with PCR (use known primers if we want to know if a specific gene is affected)
5. Or use NGS (we can analyze the entire genome to see which regions are affected)

33
Q

Why can we use ChIP-seq to figure out which regions are affected?

A

Because most methylation are associated with enhancers or transcriptional factors, so by identifying where the modifications are with antibodies, we can also identify where those factors are.
It’s used for individual genes but also the wide genome.

34
Q

What does a histone deacetylase do?

A

It deacetylases the histone tails.
Acetylation neutralizes electrostatic interactions between DNA backbone and histone tail, to loosen up the grip.
Deacetylae is used to tighten the grip and make sure the genes are not transcribed. They’re transcriptional repressors.
SIR2 is one.

35
Q

What is the role of Histone Acetyl Transferases? (HATs)

A

It neutralizes the electrostatic interactions between the DNA phosphate backbone and the histone tail.
It allows the chromatin to loosen up and helps with transcriptional activation.

36
Q

What are examples of Histone Acetyl Transferases (HATs)?

A

Gcn4p
Gcn5p
CBP
p300

36
Q

What are examples of Histone Deacetylation Complexes?

A

Rpd3p
SIR2

37
Q

What are co-activators?

A

They’re complexes that have enzymatic activity so they can add an acetyl group. They interact with DNA binding factors to enhance and activate transcription.

38
Q

What are co-repressors?

A

They interact with DNA binding transcriptional repressors.
They recruit other proteins to create a co-repressor complex.
They change the degree of acetylation to increase compaction of the chromatin.

39
Q

When you introduce a lac repressor in a cell, what happens to the chroamtin?

A

It will condense.

40
Q

What happens to the chromatin when you put a Lac repressor and a very strong transcriptional activation?

A

The chromatin opens up and is more accessible to the transcriptional elements.

41
Q

What are epigenetic traits?

A

Traits transmitted independently of the DNA sequence itself.
There’s a modification, but the sequence is not changed.

42
Q

What are examples of epigenetic marks?

A

Inactive X (Xist, histone methylation and heterochromatin spreading)
Developmental restrictions (Polycomb)
Imprints (DNA methylation)

43
Q

Do epigenetic marks have to be changed?

A

Yes. They have to be recognized by proteins, and those traits are rewritten in the next generation.
We need an epigenetic reader and an epigenetic writer.

44
Q

What’s an epigenetic reader?

A

It’s a protein complex that recognizes those epigenetic marks.
They ensure that every daughter cell acquires the appropriate fate after division. They make sure that the epigenetic marks are inherited and acted upon accordingly.

45
Q

What’s an epigenetic writer?

A

It’s a protein complex that puts down and rewrite those epigenetic marks.

46
Q

Can a protein be a reader and a writer?

A

Yes.

47
Q

What are histone methyltranferase?

A

They’re an enzyme that represses gene activity across an entire genetic region.
Histone marks can nucleate them (form them in the nucleus).
They’re epigenetic writers.

48
Q

What are DNA marks?

A

They’re DNA segment marked by methylation.
They’re read by specific proteins, then they,re used to modify histones close to them by recruiting mSin3

49
Q

What does mSin3 do?

A

It’s involved in transcriptional repression.
Associated with deacetylation (repressor, condense the chromatin).
Recruits Histone Deacetylation Complexes (HDACs)

50
Q

What are Histone Deacetylation Complexes (HDACs)?

A

They’re transcriptional repressor complexes. The complexes recruit proteins that will take out acetyl groups from histone proteins.
It leads to more compact chromatin.

51
Q

How can we make sure that DNA that was heterochromatized stay heterochromatized even after cell divison?

A

H3K9me3.
After replication, the methylated histones will be separated between the 2 daughter cells. But we can’t have that, they all need to be methylated.
H3K9me3 recognizes the methylation marks, it will associate with the histone and if they’re not methylated, it will methylate them.
H3K9me3 is both a reader and a writer.

52
Q

What are pioneers transcription factors?

A

They are DNA-binding transcriptional factors.
They interact with DNA with sequences exposed on the outside of the nucleosome.
They’re the first transcription factors that will lead to a cascade of transcription factors that will interact with specific regions.

53
Q

What are pioneers transcription factors used for?

A

They recruit enzymes that modify the configuration of the histone tails next to them, which makes the chromatin open for transcription.
They distinguish cell types during embryogenesis.
Start of the early transcription events when the chromatin is very compacted.

54
Q

The histone tail is on which terminal?

A

N-terminal tails. That’s where the methylations are happening.

55
Q

Are histone codes random?

A

no. They’re highly conserved.

56
Q

Methylation marks need to be conserved. True or false.

A

True. They can tell if a gene has to be silenced or not. Very important to keep them and give them to the daughter cell.

BIOL200 (16 decks)

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