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Cmmb 411 > Topic 8 Transcription > Flashcards

Topic 8 Transcription Flashcards

1
Q

Differences in replication and transcription

A

Replication:

copies the entire genome once and only once per cell cycle

Both dna strands are the template for new DNA sysnthesis

Transcription:

Selectively copies only certain parts of the genome from one to multiple times

Only one of the dna strands is a template on which rna strand is built

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2
Q

What is dna dependent dna sysnthesis and dna dependent rna sysnthesis

A

When dna is made using dna (replication)

When rna is made using dna (transcription using rna pol)

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3
Q

Whag is the template strand for rna transcription called

What is the non template one called

Why

A

Non coding strand, antisense, 3-5

Coding, sense, 5-3

Because the sense strand is the exact same as the RNA except it’s has no U

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4
Q

What is the purpose of the rna transcript dissociating from the dna a few ribonucleotide from the point of synthesis

A

Multiple transcriptions of the same gene can happen at the same time

Translation can happen rapidly as soon as mRNA is made (doesn’t have to be fully made)

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5
Q

What is different in rna polymerase and dna polaymerase

A

RNA pol makes rna without using primers, dna pol needs primers

RNA pol has less proofreading mechanisms than dna pol

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6
Q

Explain the parts that make up rna polymerase

A

Eukaryotes has 3 rna pol and prokaryotes has 1

Each pol has two alpha and 2 beta subunits

But the eukaryotic pol I,II,III have 7-11 extra subunits that are specific to each pol

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7
Q

What are the role of each rna pol in eukaryotes

What is the expection

A

Pol I: transcribes the larger precursor ribosomal rna

Pol 2: protien coding gene

Pol3: tRNA and 5S ribosomal RNA

Even though we say that the pol II makes protien coding genes, it can also make no coding siRNA and microRNA

So they still have overlapping finction

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8
Q

What are the steps to transcription

A

Initiation

Elongation

Termination

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9
Q

Explain general transcription initiation

A

The promoter determines what region of the dna undergoes transcription

Three steps:

Form a closed complex structure by binding the rna pol to the promoter (this is the pre initiation complex)

The closed complex transformed into an open complex (this is the transcription bubble)

Then the initial transcribing complex makes the first 10 ribonucleotides

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10
Q

Explain general transcription elongation and termination

A

Transcription happens 5-3

The rna synthesis continues by unwinding the dna in the front and reannealing it behind

The growing rna emerges from the template and weak proofreading happens

Transcription stops and rna is released from the complex

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11
Q

What is the transcription start site

A

The +1 site

The very first nucleotide that is transcribed

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12
Q

Prokaryote vs eukaryotes transcription

A

Prokaryotes:

Have only one RNA pol

Need only one initiation factor (the sigma factor)

Eukaryotes:

Have 3 RNA pol

Need several initiation factors (like general transcription factors) for promoter specific initiation

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13
Q

What is needed for transcription in vitro (in the test tube)

A

The core promoter

This includes:

BRE: TFIIB recognition element (binds TFIIB)

TATA: the TATA box element (binds TBP)

Inr: initiatior (binds TFIID)

DPE: downstream promoter element (binds TFIID

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14
Q

What is the core promoter

A

Minimal sequence needed for accurate transcription in vitro

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15
Q

Whag is special about Inr

A

Inside the Inr sequence is the +1 site where everything downstream of that is transcribed as rna

Anything upstream is non coding

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16
Q

What are the transcription factors that bind to the core promoter in in vitro transcription

A

TBP: bind to TATA box and recruits ~10 other TAFs (TBP Accosiated factors)

TFIIB: bind the PIC after the TBP binds, sets the directionality for transcription, also brigdes between the TBP and pol II

TFIIF: bind to the promoter with pol II, the polII-TFIIF stabilized the DNA-TBP-TFIIB complex and recruits TFIIE and TFIIH

TFIIE: recruits and regulates TFIIH

TFIIH: uses atp to transition the pre initiation complex to open form by melting,

Phosphorylates the CTD (c term domain) of pol II which triggers the pol activity for transcription

Recruited other proteins to do 5’ capping and stabilize the rna

Also used in NER

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17
Q

The formation of the PIC is

A

Sequential

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18
Q

Explain how the PIC is formed in vitro

A
  1. The TBP binds to TATA box, TFIID is recruited, 11 TAFs are also recruited
  2. TFIIA AND TFIIB bind
  3. The pol II with TFIIF binds

Until this point the dna is in closed form

  1. TFIIE AND TFIIH bind to complete the PIC, TFIIH melts DNA and makes dna open form
  2. The CTD of rna pol II is phoporylsted by TFIIH, promoter escape happens and transcription elongation starts with the first few nucleotides being transcribed
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19
Q

How does the TBP bind to TATA box in vitro trasncription

A

The TBP has a beta sheet which binds to the minor groove of the TATA BOX

The TBP binding changes the confirmation of the TATA minor groove to bend which widen the minor grooves to a flat structure

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20
Q

How does TFIIB bind to the core promoter in vitro transcription

A

Bind to the major grooves lf the promoter region

This sets the unidirectionality of the transcription since it bind only to on side of the promoter (assymetric)

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21
Q

What is PIC

A

Pre initiation complex

Protien complex containing pol and general transcription factors

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22
Q

What is special about the rna pol II CTD

A

It has repeats for phosphorylation and it isn’t found on pol I AND III

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23
Q

Why do we do. In vitro transcription

A

To understand the minimum players/requirement for transcription

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24
Q

What is included in in vivo transcription upstream of the core promoter

A

Has the regulatory sequences needed to efficient transcription in vivo

Can be very far from the promoter

Includes:

Proximal promoter elements

Upstream activator sequences (UASs)

Enhancers

Silencers

Boundary elements

Insulators

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25
Q

Why are addition regulatory protien upstream of the core promoter needed for transcription in vivo

A

Because the dna template in vivo is in the chromatin form

So we need additional protiens to help relax the chromatin structure to allow for transcription factors to recognize the promoter region

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26
Q

What are the activators in vivo transcription

A

The activators recognize activation site upstream of the promoter

Recruit pol and stabilize the pol : promoter interaction

Bind to chromatin remodeling complexes: this includes chromatin remodeler and HAT so they can modify the nucleosome structure and open up the dna for transcription

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27
Q

What is the mediator complex in vivo transcription

A

Bridges/connects the CTD of the pol and the upstream activators

Regulate the activity of the TFIIH to regulate gene expression

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28
Q

What is different about mediator protiens between yeast and humans in vivo Includes

A

In human They do not bind one at a time in a sequential manner they instead form subcomplex structures called modules

Have similar shape and a larger than RNA pol

The Med17 subunit of the module is needed to polII transcription in vivo

The function of most individual subunits is unknown

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29
Q

What techniques can we use to understand the subunits in modules

A

Use immunoprecipitation

Make an antibody to a specific protien X, bind the antibody to the beads, when x comes down all thing is binds to comes down and we can see y and z come down with it using SDS page

This shows what protiens were specific to that module

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30
Q

What needs to happen for elongation to occur

A

The nucelosomes in the front of the polymerase need to be removed for transcription to occur

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31
Q

What removes the nucleosomes for transcription elongation to occur

A

FACT (FAcilitates chromatin transcription) dimer

Has the Spt16 and SSRP1 subunits which disassemble the histones in front of the RNA pol

But also important to reform the histones behind the pol after transcription is done

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32
Q

After elgonation has becgun what happens to the initiation factors

A

The initiation factors dissociate from pol II

Elongation factors are recruited to the CTD tail of pol II, this recruitment depends on the phosphorylation state of the tail

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33
Q

What are elongation factors

Give examples

A

Factors that stimulate elongation

The ELL protien family and TFIIS increase the rate of elongation by limiting the time that pol pauses

TFIIS can also proofread the new transcript

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34
Q

What are RNA processing enzymes

When are they recruited

A

5’ capping enzymes (puts cap on mRNA)

3’ polyadenylation and cleavage factors (to cleave the mRNA from the pol)

Both important for mRNA stability

Splicing factors

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35
Q

When are rna processing enzymes recruited

A

Recruitment of different processing enzymes depends on the phosphorylation state of the CTD tail of the pol II

if the CTD phosphorylated at diff sites, diff processing enzymes are recruited

the capping enzyme is recruited to the 5’ end of the mRNA to Add the 5’ cap and protect from 5-3 Exonucleases

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36
Q

When does rna intron splicing happen

A

As the rna is made, the introns are exposed

The splicing can happen before the rna sysnthesis is completed and while the 5’ cap is being added

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37
Q

Explain how the 5’ cap is formed

A
  1. RNA triphosphatase removes the gamma phosphate from the 5’ end of the rna transcript (order is gamma beta alpha)
  2. Guanalyltransferase adds GMP (guanosine monophosphate) to the beta phosphate of the rna transcript
  3. Mehtyltransferase adds a methyl group to the guanine base from the GMP

This makes the 7 methyl guanylate cap at the 5’ end of the mRNA

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38
Q

What is the purpose of the 5’ cap

A

Stabilized the transcript

And signals that the transcript is correctly processed

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39
Q

Explain how the poly A tail is added

A

When transcription almost finished

The rna pol II transcribes the poly A signal AAUAAA

After this, the phosphorylated CTD tail recruits poly adenylation enzyme CPSF to the poly a signal on the mRNA

The recruited CPSF also recruits CstF to the poly A signal

The RNA pol stil transcribes a few nucleotides but falls off shortly

These cleavage factors cleave the rna downstream of the poly a signal to remove the mRNA from the pol II

Poly A polymerase (PAP) adds ~200 Adenines to the 3’ end of the mRNA

Then the poly A binding protien (PBP) coats the poly a tail

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40
Q

What is the purpose of the poly A tail

A

Stabilized the mRNA (protects from 3-5 Exonucleases)

Signals correct 3’ end processing

41
Q

What are the models of transcription termination

A

Torpedo model

Allosteric model

42
Q

Torpedo model

A

A 5’-3’ exonuclease (torpedo) called Xrn (humans) or Rat1 (eukaryotes)

After the poly A signal is added and the second mRNA is being made (since rna pol still transcribing some nucleotides) the torpedo degrades the second rna from pol II

This second rna is unprotected since no 5’ cap

The degredation from the exonuclease is faster than the sysnthesis speed of the pol so the pol II is chased out and dissociates

43
Q

Allosteric model

A

When the second mRNA is being made, the pol II loses its processivity due to a conformational change

Pol II loses its affinity for the template and dissociates and stops transcription

44
Q

What is the purpose of rna pol I

Why do we need it

A

Transcribes precursor/ ribosomal rna (non protien coding)

Meaning it only transcribes one gene that serves as the precursor RNA, dedicated to only one gene

We need this because that one gene has many isoforms that need to be transcribed in a very high level whcih is why pol I is dedicated to making all of those isofoms

45
Q

How does pol I transcription happen

A

The precursor transcript has the UCE (upstream control element) and the core promoter

These are impotent for the regulation of pol I transcription

UBF protiens bind to the UCE, recruits SL1 to the promoter region

SL1 has TBP + 3 TAFS to bind to the TATA box in the promoter

Then pol I recruited for transcription of that transcript

46
Q

What does pol III do

A

Transcribes tRNA, 5s rRNA and other small RNA

47
Q

How does pol III transcription happen

A

The promoter is downstream of the start (+1) site, meaning the transcription starts upstream of the promoter and the promoter is transcribed

TBP recognize the TATA box, recruits TFIIIC to the promoter, leading to recruitment of TFIIIB at the start site (order is IIIB,TBP,IIIC)

Then pol III binds to the TFIIIB and displaces TFIIIC to start transcription

48
Q

What is the primary point of gene regulation (first point we can regulate gene expression

A

Transcription initiation

49
Q

What is transcription controlled by

A

Activators and repressors

50
Q

What can influence the gene expression

A

Nucleosomes and their modifiers

Regulation of RNA splicing

51
Q

What elements are upstream of the core promoter and what are the definitions

A

Promoter proximal elements:

Upstream activator sequences (UASs): it’s an enhancer in yeast only found upstream of a gene but not a far distance

Enhancers: tight cluster of regulatory binding sites that affect long distances at either upstream or downstream of the gene

Silencers

Boundary elements

Insulators: block promoter activation by binding to activators at the enhancer

52
Q

What are

regulatory binding sites

Regulatory sequences

Promoter

A

regulatory binding sites: binding stirs for different transcription factors

Regulatory sequences: the entire collection of regulatory binding sites for a gene

Promoter: region of DNA involved in the binding of the pre initiation complex

53
Q

What am happens to gene regulation in more complex genes

A

As the genes get more complex , bacteria to human, the regulatory sequences and genes expression is more complex

54
Q

What is a transcription activator in eukaryotes give an example

Explain the structure of it

A

GAL4 activates transcription of the gal1 gene (galactose) in S cerevisiae by binding to the UAS region

GAL4 is a dimer (made of two monomers that come together) and can recognize palindromic sequences

Has a modular structure: DNA binding domain and activation domain

55
Q

Explain the GAL1 gene UAS

A

The UAS has four regulatory sequences the each bind a dimer of GAL4

It’s 275 BP upstream of the start site

56
Q

What is special about activator binding to regulatory sequences

A

A different activator binds to reach regulatory sequences

but all of the regulatory sites dont need to be bound at the same time to regulate gene expression

57
Q

Explain the domain swap experiment

A

It’s demonstrates the modularity of transcription factors like GAL4

WT gal4 activates transcription of the lacZ gene but Gal4 without the activation domain, the transcription of lacZ doesn’t happen

Then make a hybrid of gal4 and lexA (activation domain from gal4 bind it to the LEXa dna binding domain) found that the transcription of lacZ gets activated in the hybrid but not in the lexA dna binding domain

Showed that both the activation domain and the DSB binding domain is required for initiation and activation of transcription

58
Q

Explain the yeast two hybrid assay

A

Detect protien protien interactions based on yeast transcription factors , and the target protiens function based on the function of the things it’s binding to

protien a (bait) has the DNA binding domain

By itself gives no transcription of the reporter gene because only has one of the essential domains

If mix protien a and a bunch of other protiens with activation domains them together and transcription occurs, this means protien a and b (has activation domain) interact and bind to each other to allow transcription

59
Q

What are the ways to screen for transcription in the yeast two hybrid assay

A

Have his- Yeast cells that can’t make histidine properly and die without his media

the reporter gene expresses the his precursor so If transcription happens the yeast survive (are self suffienct), if not they die

Also can use GFP gene to see if green colour shows after transcription

60
Q

What can be present in the dna binding domain of a transcription factor

A

Helix turn helix motif (in homeodomain protiens)

Zinc containing dna binding domains (zinc finger like TFIIIA or zinc cluster like Gal4)

Leucine zipper motif (like the GCN4 in yeast)

Helix loop helix

61
Q

What can be present in the activation domain of a transcription factor

A

Hard to tell apart because they don’t have defined/distict motifs

So they are instead grouped by their amino acid composition: example GAL4 has acidic activation domain, or some are glu or pro rich

Have sticky surfaces since involved in protien protien interactions (but non specific so binds to any protien)

62
Q

Explain the helix turn helix motif

A

Two aplha helices separated by a short turn

The recognition helix recognizes specific base pairs in the major groove

The other helix positions the recognition helix into the major groove

63
Q

What are homeodomain protiens

A

Found in all eukaryotes, example of helix turn helix structure

Have 3 alpha helix , helix turn helix motif

Helix 1 terminal end interacts with the minor groove through arg amino acid

The helix 3 is serving as the recognition helix which interacts with the major groove though ser arg asn amino acids

All together helix 1 and 2 stabilize the 3rd helix into the major groove

64
Q

Describe the zinc contains domain

A

Have the recognition helix connected to a beta sheet

In between is the zinc ion that coordinates with 2 his (from recognition helix) and 2 cys (from beta sheet)

This coordination makes the structure stable and function properly to act as a dna binding domains

65
Q

Describe the leucine zipper motif

A

A dimer of two leucine zipper alpha helices

They flank and recognize both sides of the dna major groove

The top of the dimer is a coiled coil domain that has leucine residues to hold he two monomers together

66
Q

Describe the helix loop helix (HLH) protien

A

Similar to helix turn helix but more loop than turn

Monomer of 1 short alpha helix and 1 long alpha helix comes together with another to form a dimer

Flank on both sides of the major groove for dna binding

67
Q

How can we tell which transcription factor is interaction with what region on the dna

A

Through chromatin immunoprecipitation (ChIP)

68
Q

Explain ChIP

A

Used to determine protien DNA interactions , called chromatin because any dna protien complex is called a chromatin structure

Want to find a specific transcription factor (B) that’s interacting with the promoter region (so separate from all another TF binding to their promoter)

They cross link/fix the Transcription factors to the dna promoter using formaldehyde

They shear/fragment the dna, do immunoprecipitation with it through anti-B-antibody beads

Then collect the dna protien b was interacting with and do two things:

Microarray: can tell the sequence of the dna

PCR: to amplify the dna sequence

All to see what DNA protien b was interacting with

69
Q

What are the major functions of transcription activators

A

Protien recruitment:

Recruiting other TF

Recruiting nucleosome modifiers (to change chromatin structure)

Indirectly recruiting RNA pol

Reciruitng factors needed for initiation and elongation

They work cooperatively and synergistically (together and at the same time) to enhance transcription activation

70
Q

How do activators recruit protiens

A

Bound to the enhancer region of the DNA upstream of start site

They recruit TFIID and the mediatior directly

Recruit RNA pol indirectly

The effects of its are additive meaning the more interactions with activators, the stronger the stimulation of transcription

71
Q

How do activators recruit nucelosome modifiers

A

DNA is in heterochromatin (tight structure)

They recruit histone modifiers such as histone acetyl transferase that opens up the chromatin

Recruit nucleosome modifiers like the chromatin remodeling complex that use ATP to physically move nucleosomes and expose the dna (also like the SWI/SNF family of modification complexes)

72
Q

What are enhancers and insulators

A

DNA sequences:

Enhancers are bound by activators to enhance transcription, far away upstream or downstream from the promoter region

Insulators are between the enhancer and promoter regions, protiens binding to the insulator blocks the signal of the enhancer and turn gene expression off

But if sequence is promoter enhancer insulator promoter , then the transcription can be selectively activated for the left side genes

Also if enhancer insulator promoter enhancer, the downstream enhancer can still activate the promoter upstream

73
Q

Explain the looped out model for enhancer function

A

Because the enhancer region can fold and contact the pre initiation complex both downstream and upstream by folding

This is why the enhancer is distance and orientation independent

74
Q

What is the LCR (locus control region and what does it do

A

A gene sequence which regulates the expression of a cluster of genes

In human it’s regulating the 5 globin genes in adult bone marrow which are temporally and spatially regulated in their expression

The expression order is eta gamma g gamma a delta beta

This sequential expression is done by the LCR region that is upstream of these genes

75
Q

In the LCR in human how do we switch. On the globin genes in the correct order

A

We don’t know yet but the LCR in mouse gives an idea

76
Q

Explain the LCR in mice

A

They have the GCR (global control region) which works like the LCR

The GCR controls HoxD gene expression which is invoked in body segmentation and the patterning of limbs

Don’t know exactly how it works but now we know that there is a regulatory element other than enhancer and insulator that regulates the gene expression (the GCR/LCR)

77
Q

Explain the synergistic direct effects of activators

A
  1. Two activators can bind to their sites and directly interact with each other
  2. The two activators are spaced further apart and interact with a common protein to bridge the two activators

Together they have stronger transcription activation than alone

78
Q

What is synergistic

A

Different activators can recruit a single protien by touching different parts of it

The effect of the activators working together is greater than the sum of individual alone

Synergy serves as a checkpoint to ensure proper signals are received

79
Q

Explain the indirect effects of activators

A

Have the coiled nucleosome structure and Activetor B cant bind

  1. Activator A recruits a nucleosome remodeler which reveals a binding site for activator B
  2. Binding of Activator A unwinds the nucleosome , this reveals the binding site for activator B

Shows how they still work together synergistically

80
Q

Explain the real life example of synergy with activators for S cerevisiae

A

S. Cerevisiae yeast has a and alpha haploid mating type, to express, the mat locus has to be digested by the endonuclease HO

This HO gene then needs to be expressed , but it is only expressed in the mother cells not daughter cells (daughter cells have HO expression suppressor)

This expression of HO is only in the G1-S transition of mitosis

To make the HO gene, first SWI5 binds to the activator region upstream and recruits chromium remodeler and histone acetyl transferase to open the chromatin

The region between the first HO nucleotide and the activator region is now open for SBF to bind there

This SBF turns on the transcription of HO

Shows how the activators work synergistically/cooperatevely to activate expression

81
Q

Explain the real life example of synergy with activators for the human beta interferon gene

A

Beta interferon gets expressed upon infection

Upstream of the interferon genes is the enhancer region but its is bent and inaccesible

Architertural protiens HMGA1 bind to the enahncer region which recruit the histone modifiers to straighten out the enhancer region

This allows other activator: Jun/ATF, IRF, NF- kappa B, to bind to the enhancer region

form a complex called the enhanceososome to turn on Beta interferon expression by HMGA1 keeping the enhancer straight

82
Q

What are the ways that transcription repressors can work

A

Only in prokaryotes: They block RNA pol binding sites by binding to the promoter

Interact with the RNA pol at the promoter to inhibit initiation of transcription

Competition: overlapping dna binding sites of the repressor and activator makes it so that when the repressor binds the activator is blocked

Inhibition: binding of repressor allows the repressor to interact with the activator to stop its function

Direct repression: Interfere with the activity of activators, mediators, or rna pols at the promoter to inhibit initiation of transcription

Indirect repression: Recruit histone modifiers like deactylase to further compact the chromatin structure and stop expression

83
Q

Explain the combinatorial control of activators and repressors

A

Mutiple combinations of activators/ repressors on a single gene come together to allow regulation and activation of gene expression

A common regulator is found in the promoters of different genes (gene a and b) to regulate their expression and turn in multiple genes at the same time

They also each have their cell type specific regulators

84
Q

Explain real life case of combinatorial control

A

S cerevisiae expressing its specific mating type

Has three types of cells: alpha haploid, a haploid, and a/alpha diploid

Each has a mat locus and each express their own cell type specific regulators

For a: a1 regulator

For alpha: alpha1 and alpha 2

For diploid: a1 and alpha2 (repressor)

In each of these 3 cell types they also have a common regulator called Mcm1

These regulators work in combination to express either the a, aplha, or haploid specific genes (aSG, alphaSG, hSG)

85
Q

Explain each case for the mating type genes in S cervisae

A

These regulators work in combination to express either the a, aplha, or haploid specific genes (aSG, alphaSG, hSG) in each cell

a cell: hSG no regulators bound, hSG is expresss and cell stays in halpoid stage,

aSG has mcm1 bound to express aSG

Alpha cell: hSG no regulators bound, hSG is expresss and cell stays in halpoid stage,

aSG has mcm1 bound but also alpha 2 as a dimer (repressor) to stop aSG expression

The alphaSG promoter region has weaker binding to mcm1, so has alpha1 and mcm1 bound to allow tighter binding, turns on alphaSG

A/alpha diploid: hSG has a1 and alpha 2 bound, hSG is repressed by alpha 2 and cell stays in diploid stage

aSG has mcm1 and the alpha 2 dimer repressor so aSG off

There is no alpha1 in the diploid cell so alphaSG not expressed

So all three genes off for cells to stay in diploid stage

All of these combinatorial regulation of activators and repressors can help change cell type

86
Q

How else can gene/transcription expression be regulated

What are they called

A

By signal transduction pathways

JAK/STAT pathway

MAPK pathway (mitogen activated protien kinase)

87
Q

Explain the JAK STAT pathway

A

Have RTK, the cytokine ligand comes in

Receptor dimerizes, intracellular domain auto phosphorylates tyrosines, receptor activated

STAT with has SH2 domain which recognizes the phosphotyrosines, then STAT dimerizes

Once dimer, STAT goes into nucleus as a transcription activator to activate gene expression

STAT: signal transducer and activator of transcription

88
Q

Explain the MAPK pathway

A

Have RTK, growth factor ligand, dimerizes, autopohsphoylates

This recruits adaptor with sh2 domain like SOS (which is a gef: guanine nucleotide exchange factor) and grb2

These bind gdp RAS (a small gtpase), whcih becomes active in GTP form (due to SOS)

The active RAS actives mapKKK to activate mapKK then mapK

When mapK active, it phosphorylates transcription activators like JUN to send them to nucleus and activate beta interferon expression

89
Q

What is RAS

Why is the called MAPK mapkk mapkkk

A

Earliest discoevered oncogene

Named in order of discovery

90
Q

How can gene silencing regulation happen

A

By histone modifiers and nucleosome remodelers forming heterochromatin

91
Q

Explain the transcriptional silencing by heterochromatin formation

A

Most heterochromatin is at Telomeric regions because more packed and less transcription but below that is euchromatin

This happens because when telomere is being formed RAP1 dna binding protien binds then recruits sir2 histone deacetlyase (closes chromatin structure)

Also recruit sir3,4 which extends/spread the heterochromatin to a larger region to make larger telomeric closed regions

92
Q

Where is heterochromatin mostly found

A

In telomeres and centromeric regions

Centromere has rRNA repeats which are very redundant and a lot of it isn’t needed, this is why more compact so less expression of that ribosomal rna

93
Q

What are sir protiens

A

Yeast telomeric SILENCE INFORMATION REGUALTOR protiens

94
Q

What can stop the spread of heterochromatin silenced region

A

Insulator elements can block the spread of histone modifications

Histone methyl transferases (close down the dna structure) repress the spreading of SIR2 mediated silencing (since now silencing using diff method)

Methylation of the histone H3 tail: can either increase or silence transcription (h3K4 methylated increase, h3k9 decrease

95
Q

How can dna be modified to silence gene expression

A

DNA has cpG islands (CG repeats in the promoter region)

The cytosines get methylated on the 5’ end of this ring to become 5’ methyl cytosine

This causes leakage of gene suppression, meaning not fully suppressed

To fully suppress, protien MeCP2 need to bind to the 5 Me cytosine and recruit histone modifiers/nucleosome remodelers which fully close chromatin and silence

Sometimes want full silence not just a little

96
Q

Explain how imprinting can determine gene expression in the offspring

A

Have maternal and paternal chromosome

The maternal chromosome: igF2, ICR (insulator), H19, enhancer

has enhancer downstream of h19 and igf2. H19 (non conding rna that suppresses cell division) is transcribed. The ICR insulator blocks the enhancer from also expressing igF2

Paternal: the H19 and the ICR insulator region is methylated. Since no insulator or h19, enhancer expresses igF2

Igf2 (insulin growth factor 2) allow cell division

This is the balance between igf2 expression and h19 expression between maternal and paternal balances cell division

97
Q

Explain the disorder that happens when the imprinting is wrong

Explain symptoms

A

Beckwith-wiedermann syndrome

Caused by overexpresion of paternal genes (more igf2), or mutation in maternal genes (less H19) on chromosme 11

This result in over activity of igf2 so overgrowth of the cells

Overgrowth, enlarged tongue, midline abdominal wall defects, severe hypoglycaemia (low blood sugar after birth), increase risk of childhood cancers

98
Q

Explain how the dna methylation can go from one generation to the next and what happens

A

Modification Passed on through cell division

the CpG sequence is palindromic meaning the opposite strand also has the CpG modification. After replication this needs to methylating needs to be added again to the daughter strand

Upon replication the hemimethylated CpG are methylated by a maintence methylase

The maintence methylase only recognizes the hemimethylates cytosine not the unmethylated one so that only the ones that are supposed to be methylated are methylated

Cmmb 411 (8 decks)

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