12 - Eukaryotic Transcription Flashcards

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1
Q

do bacterial transcription and translation occur in the same compartment
- what compartment

A

yes
- the cytoplasm

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2
Q

where eukaryotic transcripts are processed an transported to
- give an example

A

processed in the nucleus (nuclear envelope)
- transported via nuclear pore to the cytosol

e.g. mRNA precursors processed in nucleus and transported to the cytosol for translation

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3
Q

Three main RNA Pol in eukaryotes

A

RNA Pol 1
RNAPol 2
RNA Pol 3

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4
Q

general structure of eukaryotic RNA Polymerases

A

contain:
- B’ and B-like subunits
- alpha-like subunits
- common subunits
- enzyme-specific subunits

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5
Q

Conserved elements of eukaryotic RNA Pol II promoters

A
  • lnr - initiator element
  • TATA box promoter has a TATA box
  • TATA-less promoter has a DPE (downstream core promoter element)
  • enhancer - can be several kb away
  • maybe a CAAT box or a GC box between -40 and -150
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6
Q

differences between bacterial and eukaryotic transcriptional promoters

A
  • spacing is different, and CAAT box (or GC box) of eukaryotic promoters can be on either strand
  • the -10 and -35 sequences interact with a part of the holoenzyme (sigma subunit)
  • whereas the TATA, CAAT and GC motifs are recognised by other proteins, and not RNA Pol II
  • meaning these are fundamentally different RNA replication mechanisms
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7
Q

Eukaryotic transcription - initiation mechanism

A
  • TFIID (a transcription factor) guides RNA Pol II to its promoters
  • TFIID contains the 30kDA TATA-box binding protein (TBP)
  • TBP binds tightly to the TATA box (10^5 times more than nonconsensus sequences)
  • bending of DNA caused by binding widens the minor groove
  • complex forms called the basal transcription apparatus
    formation of this complex in order:
  • TFIIA binds TBP
  • TFIIB binds TBP
  • TFIIB recruits TFIIF, RNA Pol II, TFIIE and TFIIH
  • TFIIH is bi-functional
  • it is a helicase that opens the DNA double helix
  • and also is a kinase that phosphorylates the C’ terminal domain (CTD) of the RNA Pol II L’ subunit
  • Phosphorylation of CTD marks transition from initiation to elongation
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8
Q

general nomenclature of transcription factors

A

e.g. TFIID
- TF (Transcription Factor)
- II (Pol II)
- D (A, B, C, D)

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9
Q

TFIIH function in eukaryotic transcription

A
  • TFIIH is bi-functional
  • it is a helicase that opens the DNA double helix
  • and also is a kinase that phosphorylates the C’ terminal domain (CTD) of the RNA Pol II L’ subunit
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10
Q

What marks transition from initiation to elongation in eukaryotic transcription

A
  • phosphorylation of CTD (C’-Terminal Domain) by TFIIH
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11
Q

eukaryotic transcription - elongation mechanisms

A
  • TFIIB, TFIIE and TFIIH dissociate from basal transcription apparatus complex
  • RNA is synthesised
  • RNA Pol II progresses
  • it frees the promoter and TFIID/TFIIA complex for further recuitment
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12
Q

Inhibitor of RNA Pol II
- process of inhibition

A
  • alpha (a)-amanitin
  • found in death cap fungi/mushroom
  • it is a cyclic octapeptide
  • binds tightly to active site of RNA Pol II
  • reduces rate of RNA production to a few nucleotides per hour
  • constrains the flexibility RNA Pol II requires to translocate DNA through its active site
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13
Q

enhancers of eukaryotic transcription
- how enhancers affect DNA looping model

A
  • they increase the transcription level of genes
  • can be active in a tissue-specific manner
  • so can play developmental roles in the body
  • DNA looping model
  • proteins bound to a distant enhancer interact with components of transcription initiation complex
  • this loops out the intervening DNA
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14
Q

The DNA looping model overview

A
  • Mediator protein acts as bridge between activator proteins that bind enhancer control elements and non-phosphorylated CTD of RNA Pol II in its initiator state
  • enhancer control elements bind activator proteins
  • silencer control elements bind repressor proteins
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15
Q

enhancer and silencer gene function in eukaryotic transcription

A

enhancer - turn genes ‘on’ - allow transcription of genes
- activator proteins required to bind enhancer control elements
silencer - turn genes ‘off’ - repress transcription of genes

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16
Q

histone acetylation in eukaryotic transcription

A
  • histone tails protrude outwards from a nucleosome
  • acetylation of histone tails promotes a loose chromatin structure that permits transcription
  • allows regulation of transcription
17
Q

control of gene activation in enhancer proteins for eukaryotic transcription

A
  • since activator proteins are required to bind enhancer control elements in transcription
  • transcription can only be enhanced if appropriate/specific activator proteins are present
  • this allows cell-specific control of gene expression stopping
18
Q

Processing of eukaryotic pre-mRNA - simple

A

RNA Pol II transcripts (mRNAs) are modified in three ways:
- 5’ end capped by a nucleotide triphosphate through an unusual linkage - accompanied by methylation
- 3’ end of mRNA is trimmed and poly-A tail is added
- Introns are removed by splicing (mRNA splicing)

19
Q

Addition of a 5’ cap on mRNA in Pol II Eukaryotic trancription - process

A
20
Q

Addition of poly-A tail on eukaryotic mRNA

A
21
Q

How do capping and tailing enzymes find the mRNA transcripts in eukaryotes

A
22
Q

Major differences between bacterial and eukaryotic mRNA transcription

A

Bacterial:
- primary transcript is often a mature mRNA and is used directly for translation
- transcription and translation occur in the same compartment - the cytoplasm - so may be coupled
- bacterial mRNAs are often polycistronic; they encode more than one protein

Eukaryotes:
- primary transcript is not mature and has to be processed before translation
- transcription is nuclear, translation is cytosolic - no coupling of the two processes
- eukaryotic mRNAs are monocistronic; they encode just one protein