Brainscape's Knowledge GenomeTM

Browse over 1 million classes created by top students, professors, publishers, and experts.


See full index

LF206: Molecular Cell Biology > Lecture 5. Transcriptional Regulation > Flashcards

Lecture 5. Transcriptional Regulation Flashcards

You may prefer our related Brainscape-certified flashcards:
AP® Biology flashcards Biology 101 flashcards
1
Q

Why is transcriptional regulation needed?

A

Allows development of different tissues
Transition from childhood to adult
Deregulation can result in uncontrolled growth cancers
Allows reaction to environmental cues
And so on

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
2
Q

What is an example of transcriptional regulation?

A

The transition from foetal to adult haemoglobins – changes in protein subunits expressed and therefore changes in functions, all co-ordinated with birth

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
3
Q

What determines how/when/why genes are transcribed?

A

Chromatin structure
RNA polymerase (and general TF) binding specificity
Additional binding and activation factors

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
4
Q

How are expressed genes found in the chromatin?

A

Expressed genes are found in the ‘open’ conformation

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
5
Q

What genes within chromatin are not usually expressed?

A

Genes within highly packed heterochromatin

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
6
Q

What does methylation at position 9 on histone 3 (H3K9me) result in?

A

Heterochromatin formation - Gene silencing

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
7
Q

What does methylation at position 27 on histone 3 (H3K27me) result in?

A

Inactivation of HOX genes (developmental genes that are arranged int he same order as the segments of the body). X chromosome inactivation
Two X chromosomes could cause gene overdose so one of them is inactivated randomly in all cells

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
8
Q

What does H3S10p H3K14ac and H3K4me H3K9ac result in (acetylation)?

A

Relaxes chromatin allowing gene expression
Both open up the chromatin (methylation at position 4 says to stay open in H3K4me H3K9ac)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
9
Q

Why is methylation at position 4 on histone 3 important?

A

Important for the structure of the centromere that needs an open confromation at all times

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
10
Q

What is the structure of an interphase chromosome?

A

30nm fibre is compacted into loops held together by scaffold proteins
Scaffold proteins are rich in topoisomerases that regulate torsional changes caused by packing/unpacking (implies a lot of unwinding and rewinding)
Loops can extend when we need to express them (loops are mobile)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
11
Q

How does an interphase chromosome turn on a gene?

A

The loop that bears the gene that needs to be turned on opens up and relaxes
The loop moves into an area where all the requirements for transcription are concentrated

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
12
Q

How does an interphase chromosome turn off a gene?

A

The gene is moved towards the nuclear periphery and allowing enzymes associeted with the nuclear lamin to turn the gene off

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
13
Q

What are the two major types of heterochromatin?

A

Facultative heterochromatin and Constitutive heterochromatin

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
14
Q

What is facultative heterochromatin?

A

Cell-type-specific, can switch into euchromatin following developmental cues (turns gene back on). Characterised by a specific histone code mark: H3K27me3 that binds ‘polycomb’ proteins (important in development)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
15
Q

What is constitutive heterochromatin?

A

Regions that are consistently silenced in all cell types of an organism - centromeres, telomeres, some transposons and some gene-poor regions of the genome. Characterised by H3K9me3, a modification carried out by the histone methyltransferases (HMT). These HMTs propagate heterochromatin by recognising H3K9me3 and methylating adjacent nucleosomes. HMTs copy signals and pass it on to neighbours

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
16
Q

What is centric chromatin?

A

Flanked on each edge by pericentirc chromatin (normal H3K4me2)
Long highly repetitive chromatin structures (centromere specific H3)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
17
Q

How do centromeres remain open?

A

The centromeric histone force an open structure
Double methylation at position 4 also forces open structure

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
18
Q

Because centromeres remain open, what does this allow access to?

A

Kinetochore proteins which then associate with the centromere and acts as the focal points for the microtubules that will eventually pull the centromere apart

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
19
Q

What is the structure of telomeres?

A

Telomeres are repetitive structures
Vary in length and repeated DNA sequence (constant repeating of same sequence)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
20
Q

What are examples of telomere repeats?

A

GGGTTG in the ciliate Tetrahymena (unicellular organism)
GGGTTA in vertebrates
G1–3A in Saccharomyces cerevisiae

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
21
Q

What is the telomere problem/how do telomeres shorten?

A
  1. Because DNA synthesis can only proceed 5’ to 3’ there is continuous synthesis on the leading strand and discontinuous synthesis on the lagging strand: synthesis requires RNA primers
  2. The lagging strand template can be primed near the telomere (and then extended)
  3. The DNA polymerase complex disengages (two Okazaki fragments, each with RNA primer)
  4. The RNA primers are erased leaving two gaps
  5. The gap on the leading strand is filled by a DNA polymerase and repaired by a DNA ligase. The gap on the lagging strand cannot be filled by a DNA polymerase as there is no primer, so there is no 3’-OH for extension (telomere shortened)
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
22
Q

What is the consequence of both the 5’ to 3’ synthesis of DNA and the erasure of RNA primers?

A

Telomeres shrink

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
23
Q

What is the Hayflick limit?

A

The number of times a normal somatic, differentiated human cell population will divide before cell division stops (between 40 and 60 divisions): the cell becomes senescent (cell will die). It is an inbuilt ageing mechanism

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
24
Q

How does the compensatory mechanism for telomere shortening function?

A

Telomerase binds the single-stranded G-overhang: a ribonucleoprotein (RNP) enzyme made of the telomerase RNA (TER) and telomerase reverse transcriptase protein (TERT)
It extends the 3’ end of the parental strand using its own RNA subunit as a template
And so on resulting in RNA-templated DNA synthesis that extends the telomeres

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
25
Q

In summary how does the compensatory mechanism for telomere shortening function?

A
  1. DNA primase lays down an RNA primer
  2. 3’-OH (hydroxyl) of primer extended with DNA nucleotides by DNA poylmerase
  3. Nick is joined by ligase
26
Q

How are telomeres partially lengthened by RNA-templated DNA synthesis?

A

RNA-templated DNA synthesis by telomerase extends the G-overhang 5’-3’ DNA primase lays down an RNA primer on the extended G-overhang

27
Q

How are telomeres partially lengthened by DNA-templated DNA synthesis?

A

DNA-templated DNA synthesis by DNA Polymerase extends this primer 5’-3’ DNA ligase ligates the new Okazaki fragment to the old lagging strand 5’ end

28
Q

Despite having a free 3’ unpaired end, why do telomeres not fuse?

A

3’ end must shelter somewhere from repair mechanisms

29
Q

How does the free 3’ end of the telomere shelter from repair mechanisms?

A

By base pairing
Hides in a complex called shelterin
Shelterin is made up telomeric repeat-binding factors 1 and 2 (TRF1, TRF2) and repressor/activator protein (RAP1)
These stimulates t-loop formation and displaces a d-loop that results in the base pairing at the 3’ end
The 3’ end shelters from repair mechanisms in a telosome

30
Q

What is Werner syndrome?

A

Accelerated ageing
The WRN helicase protein is important for DNA repair and telomeric DNA replication
Telomeres that are normally replicated by lagging strand synthesis are not replicated efficiently in Werner cells
Over-expression of telomerase in vitro counteracts WRN mutation

31
Q

How is Werner syndrome inherited?

A

Autosomal recessive

32
Q

What is the incidence rate of Werner syndrome?

A

1 in 1,000,000 but in Japan and Sicily it is 1 in 30,000

33
Q

What genes does RNA polymerase II (Pol II) transcribe?

A

All protein-coding genes
snoRNA genes (small nucleolar)
miRNA genes (micro)
siRNA genes (small interfering)
Most snRNA genes (small nuclear)

34
Q

How are all RNA polymerases related?

A

RNA polymerases in bacteria, archaea, and eukaryotes (Pol III) are closely related: the basic features of the enzyme were in place before the divergence of the three major branches of life

35
Q

From upstream to downstream, what does a (human) Pol II promoter (typically) look like

A

Distal control elements (same function as proximal but considered effectors)
Proximal control elements
Upstream B recognition element (BRE) - binds to TFIIB, determines where the DNA binds
TATA box - binds TATA-binding protein (TBP)
Downstream B recognition element (BRE) - binds to TFIIB
Initiator element (INR) - binds TFIID
Exon-intron-exon-intron-exon attached to poly-A signal
Termination signal

36
Q

What do all RNA Pol II promotor regions contain and what is important to remember about the structures of the Pol II promoters?

A

Multiple cis-acting elements that bind proteins
Individual elements may not always be present
Pol II promotors are very variable

37
Q

What happens when the TATA binding protein (TBP, part of TFIID) binds to the TATA box?

A

TFIID complex binds TATA box via TBP, aided complex by TFIIA
TBP recruits TFIIB which recognises upstream BRE and downstream BRE and accurately positions RNA Pol II at the start site of transcription (+1)
TFIIE, RNA Pol II/TFIIF are recruited. TFIIF stabilises RNA Pol II interactions with TFIIE and TFIIH
TFIIH is recruited by TFIIE (TFIIH consists of a helicase and a kinase)
The complex formed is known as the basal transcription apparatus/complex

38
Q

How does elongation occur in the basal transcription complex?

A

The TFIIH helicase opens the DNA double helix, allowing the polymerase to access the template strand
The TFIIH kinase phosphorylates the C’-terminal domain (CTD) of the RNA Pol II L’ subunit
Phosphorylation of the CTD marks the transition from initiation to elongation
RNA Pol II now disengages from the cluster of general transcription factors, undergoing conformational change that tightens its interaction with DNA

39
Q

How does the TATA-binding protein (TBP) locate the TATA box?

A

The TATA box is a consensus sequence: individual TATA boxes have different affinities for TBP - and so some are more efficient at stimulating transcription than others

40
Q

How do we know that individual TATA boxes have different affinities?

A

G-less cassette transcription assay
A promoter is cloned upstream of a G-less cassette (made from A, C, T bases only)
Add purified TFs and RNA Pol II, ATP, CTP and [α³²P]-UTP
Because no GTP is supplied, the RNA is truncated at the point at which ‘G’ should insert
A radioactive RNA transcript of defined size is formed. It can be electrophoresed through polyacrylamide gels and quantified following autoradiography
Human TFIID binds stronger to TATAAAA than TATAAAG (shown by electropheroses)

41
Q

How can the sequence in the TATA box change?

A

Depending on what time the gene is expressed (temporal control)
Genes in Epstein-Barr virus (EBV) have 33 and 35 length TATA boxes for early and late genes respectively

42
Q

In EBV, how complex is the IE (immediate early) gene?

A

Complexity: high
TATA box with proximal positive cis-acting elements that enhance transcription and both proximal and distal negative cis-acting elements (red) that inhibit transcription

43
Q

In EBV, how complex is the E (early) gene?

A

Complexity: intermediate
TATA box with both proximal and distal positive cis-acting elements that enhance transcription

44
Q

In EBV, how complex is the L (late) gene?

A

Complexity: minimal
Essentially a TATT version of a TATA box

45
Q

What are general transcription factors?

A

Assemble at the promoter and form a complex with RNA PolII

46
Q

What are specific transcription factors?

A

Specific transcription factors regulate the rate of transcription:
Activators increase transcription
Repressors decrease transcription

47
Q

What do all binding factors have?

A

A modular design
A DNA-binding domain that binds specific DNA sequences and an activating/repressing domain (protein interaction domain) that stimulates/inhibits transcription by interacting with mediator proteins, general transcription factors or RNA PolII

48
Q

What are examples of DNA-binding domains?

A

Homeodomains, Zinc finger motifs, Leucine zippers

49
Q

How different are the homodomain DNA-binding sites of different species and why?

A

The shape and DNA binding is conserved between species (only difference is the amino acids present in the DNA)
Helix 3 binds in the major groove of DNA making specific interactions between amino acids and nucleotides

50
Q

What is a zinc finger motif?

A

A small structural motif with key Cys and His residues that coordinates a zinc ion (Zn²⁺), stabilising the fold

51
Q

How are zinc finger motifs arranged?

A

Often there is cluster, arranged one after the other so that the α-helix of each binds the major groove of the DNA. A strong and specific DNA-protein interaction is built up through a repeating basic structural unit (binds to GC rich environments)

52
Q

How does a leucine zipper bind as a homodimer (such as yeast Gcn4 protein)?

A

Each DNA-binding domain binds one-half of a symmetrical, palindromic DNA motif

53
Q

How does a leucine zipper bind as a heterodimer?

A

Three distinct DNA-binding specificities could, in principle, be generated from two types of leucine zipper monomer, while six could be created from three types of monomer, and so on
Heterodimeric binding is common (e.g c-Jun/c-Fos heterodimers)

54
Q

What do leucine zippers allow?

A

Very tight, very regulated expression of key genes in a tunable fashion

55
Q

What does combinatorial regulation allow?

A

A powerful mechanism that enables tight control of gene expression, via integration of multiple signalling pathways that induce different transcription factors required for enhanceosome assembly

56
Q

What is the enhanceosome?

A

In special cases, in genes that require tight control, activators bind cooperatively along an enhancer sequence forming an enhanceosome
Each enhanceosome is unique to its specific enhancer

57
Q

What does the enhanceosome recruit?

A

It recruits co-activators and general transcription factors to the promoter region of the target gene to begin transcription
It also recruits non histone architectural transcription factors, called high-mobility group (HMG) proteins, which regulate chromatin structure – they ensure that the target gene can be accessed by transcription factors
Co-operative binding of different proteins. Once all proteins bound you get a fully activated effector

58
Q

What is the best known example of an enhanceosome?

A

The enhanceosome recruits HMG-1 and the transcriptional machinery to the promoter and initiates high level gene expression of IFN-β

59
Q

What does the balance between expression or repression depend on?

A

Depends upon coactivators
Proteins can chanage their orientation and ability to bind to coactivators and corepressors which increases the repertoire of responses
You can reuse the proteins in different orientations to bind either effectors or repressors

60
Q

What does the expression of an RNA PolII - transcribed gene depends on the integrated output of?

A

DNA methylation status
Chromatin structure and histone modification status
General transcription factors and RNA Pol II
Regulatory complexes bound both upstream and downstream of the gene
And mediator proteins all act together

61
Q

What happens in the Philadelphia (Ph) chromosome?

A

ABL1 on chromosome 9 translocates onto break-point cluster region (BCR) on chromosome 22
Brings the BCR promoter and enhancer next to a gene fusion of BCR and ABL1
This deletes the first exon of ABL, the exon that encodes for a protein that turns ABL off
Inappropriate expression of a fusion protein that is permanently locked on causing constant gene division that results in cancer

62
Q

What happens in Burkitt’s lymphoma

A

Translocation between chromosomes 8 and 14 brining the MYC gene (regulates cell division) now has control of the IgH promotor resulting in high levels of unregulated expression of MYC causing cell proliferation and genome instability
Inappropriate regulation of an oncogene by a very strong enhancer

LF206: Molecular Cell Biology (18 decks)

Brainscape helps you reach your goals faster, through stronger study habits.
© 2024 Bold Learning Solutions. Terms and Conditions