Eukaryotic RNA polymerases and their promoters

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Chapter 10

Eukaryotic RNA polymerases and their promoters
Page 276-295
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Polymerase II components can be categorized into
three groups

• Core subunits (Rpb1, 2, 3 similar to bacterial
  RNA polymerase subunits ?, ? and ?,
  respectively)
• Rpb1 -- ?, Rpb2 -- ?, Rpb3 -- ?
• Common subunits (Rpb5, 6, 8, 10, 12)
• Not much is known about these subunits
• 3) Nonessential subunits (Rpb4 and Rpb9)

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/?
/?

• Rpb3 may be a functional homologue of E. coli ?
• Both have two monomers per holoenzymes
• Both have similar sizes
• Both are involved in assemblying holoenzymes

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Common subunits (Rpb5, 6, 8, 10, 12)

• Functions unknown
• Could be important for
• Localization in the nucleus
• Processivity of the holoenzyme
• Fidelity of transcription
• Nonessential subunits (Rpb4 and Rpb9)
• Yeast cells with a deletion in either of these
  two genes are viable in normal temperature but
  cannot survive in high or low temperatures.
• Pol II purified from a yeast mutant deleting Rpb4
  also lack Rpb7, suggesting that Rpb4 plays a role
  in anchoring Rpb7 to Pol II.

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Pol II ?4/7
Rpb4 may play a role similar to ? and has limited
sequence homology with ?
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• What of the followings are the most critical
  components for doing an in vitro transcription
  assay?
• 1) DNA template, 2) RNA polymerase, 3)
  Nucleotides
• 4) DNA polymerase, 5) radiolabeled UTP, 6) RNA
  primers
• 1, 4, 5, 6
• 1, 2, 3, 5
• 1, 4, 3, 6
• 1, 2, 5, 6
• 1, 3, 4, 5
• B

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Heterogeneity of the Rpb1 subunit The Largest
subunit of RNA polymerase II
RPB1 governs the sensitivity of Pol II to
?-amanitin
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Figure 10.10
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Three RNA Polymerases have different subunit
compositions and transcribe different class of
genes
They may recognize different promoters
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Class II Promoters
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There are also TATA-less promoters

• Promoters for house-keeping genes (GC boxes)
• Promoters of genes that are regulated
  developmentally

Specialized genes (cell-type specific) do have a
TATA-box in their promoters.
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• TATA box locates the start of transcription 30
  bp downstream
• TATA box sometime is important for the efficiency
  of transcription
• TATA-binding protein (TBP) binds to TATA-box and
  initiates the assembly of transcription factors
  and RNA polymerase

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Initiator --PyPyANT/APyPy -- is sufficient by
itself to direct transcription Downstream
promoter elements (DPE) -- quite common in
fruitfly genes. Found 30 bp downstream of the
transcription initiation site Have the consensus
sequence -- G(A/T)CG -- behave as a TATA-box BRE
-- (G/C)(G/C)(G/A)CGCC -- TFIIB-binding site
Upstream element
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Class I promoter (Pol I promoter)
Relatively simple -- one template (the rRNA
precursor gene)
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The spacing between the two elements is important
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Class III promoter (Pol III promoter)
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A promoter must have

• a TATA box
• a GC box
• an element that direct where RNA polymerase
  should start
• a CAT box
• a site where primer can bind
• C

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Class III promoter (Pol III promoter)
Classical class III promoter
Unlike class I and II promoters, classical class
III promoters locate within the genes.
U6 snRNA, 7SL, and 7SK RNA gene still need
elements 5 of the genes. Their promoters
resemble class II promoters (nonclassical class
III promoter).
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Enhancers and Silencers
Elements that are not a part of the promoter but
can either enhance (enhancer) or inhibit
(silencer) transcription at a manner that is
position- and orientation-independent.
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How do enhancers work?

• An enhancers binds transcription factors
  (activators) to activate transcription
• An enhancer can localize upstream, within, or
  downstream of a gene.