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Genes and Genomics- Em > Transcription 1 > Flashcards

Transcription 1 Flashcards

1
Q

When the open complex forms between RNA polymerase and DNA how many base pairs are unwound?

A

17bp of DNA are unwound at the start site.

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2
Q

How large is the transcription bubble?

A

Remains as 17bp of DNA unwound as the RNA polymerse moves along the DNA.

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3
Q

In RNA polymerase where is the active site?

A

In the cleft between the beta and beta’ subunits.

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4
Q

What happens to the DNA strands once they have been unwound?

A

The two strands go through separate channels in the RNA polymerase.

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5
Q

What are the two mechanisms of termination in E.coli?

A

ρ dependent and ρ independent.

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6
Q

What is the ρ dependent mechanism of termination?

A

Requires an additional protein factor, ρ, which moves through RNAP and binds CA-rich sequences in the RNA chain- rut sequences. It then uses ATP driven helicase to unwind the RNA/DNA helix within the transcription bubble.

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7
Q

What is the ρ independent mechanism of termination?

A

Hairpin structures followed by UUU in the RNA chain causes RNAP to pause and release the transcript due to weaker A-U bonds. This pauses transcription as the hairpin clogs the RNA exit channel.

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8
Q

Describe RNA polymerases in eukaryotes.

A

Large proteins (>500kDa) and multisubunit (>8)

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9
Q

What is RNA polymerase I used for and where is it found?

A

Makes a single pre-rRNA transcript that is processed to give 18S, 5.8S and 28S rRNAs. Found in the nucleolus.

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10
Q

What is RNA polymerase II used for?

A

Makes some small RNAs and mRNAs.

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11
Q

What is RNA polymerase III used for?

A

Makes tRNAs, 5S rRNA and other small RNAs such as those associated with the spliceosome.

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12
Q

Which of the eukaryotic RNA polymerases is most susceptible to inhibition by α-aminitin?

A

RNA polymerase II.

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13
Q

What are the main differences between eukaryotic and prokaryotic promoter sequences?

A

Eukaryotic promotes are larger and more complex. Prokaryotic promoters are all upstream, whereas RNAPIII in eukaryotes has downstream promoter sequences, in the region being transcribed. Eukaryotic promoters rely on the binding of basal transcription factors for initiation.

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14
Q

What are the features of RNAPII promoters?

A
  • TATA box at -30 - Inr sequence near +1- Upstream and downstream elements- Enhancer sequences
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15
Q

Name the transcription factors that must bind to an RNAPII promoter for initiation.

A

TBP to TATA box, TFIID to TBP. Followed by TFIIA, TFIIB, TFIIF, TFIIE and TFIIH.

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16
Q

What are the two activities of TFIIH?

A
  • Helicase; unwinds DNA- Kinase; phosphorylates Ser rich region at C-terminal of RNAPII
17
Q

What does phosphoylation of the C-terminal of RNAPII allow?

A

Allows the RNA polymerase to leave the promoter, once all the transcription factors have bound.

18
Q

Why can RNAPII not initiate transcription itself?

A

There is no σ factor to allow polymerase binding to promoters.

19
Q

What is the two metal mechanism of RNAP?

A

Uses 2 magnesium ions. One is permanently bound in the catalytic site. One is brought in by NTP (where it chelates oxygen)

20
Q

What is the bridge helix?

A

alpha helix that spans the ‘claw’ of RNAP. Helix kinks when the complementary rNTP binds to the DNA/RNA duplex- moves the hybrid along making the next base on the DNA template available.

21
Q

What determines the direction of transcription?

A

Orientation of the promoter- prevents RNAP clashing during simultaneous transcription of genes on opposite strands.

22
Q

What does TFIID consist of?

A

TBF and TBP associated factors.

23
Q

What is the structure of TBP?

A

Single polypeptide. Contains an internal repeat so consists of two identical substructures.

24
Q

What does TBP do?

A

Binds to the TATA box. Distorts DNA around 90 degrees. Influences what DNA can bind to.

25
Q

What does TFIIA do and where does it bind?

A

Stabilises TFIID binding. Binds upstream of +1 start site.

26
Q

Where does TFIIB bind and what does it do?

A

Upstream of +1 at the TFIIB recognition element (BRE) via sequence specific interactions. Stabilises TFIID binding and recruits TFIIF.

27
Q

What is the role of TFIIF?

A

Carries RNAP to the promoter.

28
Q

What is the role of TFIIE?

A

Binds to hold DNA and RNAP in place. Completes formation of the pre-initiation complex. Contains a zinc finger motif than can bind ssDNA in the transcription bubble.

29
Q

What are Inr and DPE?

A

Other promoter elements that are recognised by TAFs and recruit TFIID when there is no TATA box.

30
Q

What controls the transcription rate?

A

Promoter melting- has a half life of 45 secs, transcription can only occur when the bubble is present.

31
Q

How does activator binding affect gene regulation?

A

Activators bind to specific sites upstream/downstream of the promoter. Allows mediator binding- modulates TFIIH activity. Transcription starts.

32
Q

How do gene specific transcription factors form sequence specific interactions?

A

Read the DNA sequence from the outside of the helix, using base pair geometry- pattern of hydrogen acceptors/donors.

33
Q

What is the role of activation domains in gene specific transcription factors?

A

Stimulate basal transcription machinery. Create open chromatin domains.

34
Q

Why do the activation domains of gene specific transcription factors have no stable fold?

A

They are excessively acidic, Q-rich or I-rich.

35
Q

What is a helix-turn-helix domain?

A

C-terminal recognition helix- fits in major groove. N-terminal helix stabilises structure. Can act as dimers to recognise palindromic DNA sequences.

36
Q

What is a helix-loop-helix domain?

A

Must dimerise to bind DNA- clamps DNA between the monomers. Recognition helices line up with adjacent major grooves. Basic N termini interact with -vely charged backbone.

37
Q

Where are helix-loop-helix domains found?

A

In transcription factors involved in cell proliferation, e.g. c-jun/c-fos dimer.

38
Q

What is a zinc finger domain?

A

beta turn and alpha helix held together by a zinc atom (coordinated by Cys/His). Recognises 3nts. can be used in tandem to recognise a more specific sequence (more nts). Only found in eukaryotes

39
Q

What is different about p53 binding?

A

Its central binding domain has no similarity to other DNA binding domains.

Genes and Genomics- Em (19 decks)

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