Transcription
C483 Spring 2013
1. RNA Polymerase
A.
B.
C.
D.
Polymerizes RNA in a 3’5’ direction.
Has 3’5’ exonuclease activity
Has 5’3’ exonuclease activity
Catalyzes a reaction driven by pyrophosphate hydrolysis
2. Which of the following statements is true?
A.
B.
C.
D.
E.
The template strand of DNA is copied by RNA polymerase 3’5’.
The “sense” strand is DNA; the “antisense” strand is RNA.
The coding strand of DNA is identical to the transcribed RNA.
The template strand of DNA is identical to the transcribed RNA.
A gene is read in the 3’5’ direction.
3. The rate limiting step in transcription is
A.
B.
C.
D.
Promoter binding
Initiation
Elongation
Termination
4. Weak promoters contain
A. no consensus sequences.
B. sequences that match consensus sequences poorly.
C. no TATA box.
D. a TATA box but no -35 region.
5. Which of the following is not a difference between bacterial and
eukaryotic transcription?
A. Bacterial transcription has less types of RNA polymerase.
B. Eukaryotic transcription makes use of more general transcription factors.
C. Bacterial mRNA undergo extensive post-translational modifications.
D. Chromatin plays a large role in regulation of transcription in eukaryotic
cells.
Transcription
• Gene—a DNA sequence that is transcribed
• Housekeeping vs. regulated genes
• Many types of RNA
Information Flow
• Central Dogma:
information in protein
cannot be incorporated
into information in DNA
• RNADNA in
retroviruses
• We will focus on
messenger RNA (mRNA)
RNA Synthesis
• Definition of transcription: DNA-directed RNA
synthesis
• RNA Polymerase
• Three stages
– Initiation
– Elongation
– Termination
Bacterial RNA Polymerase
• Multisubunit
• Smaller than
replisome
• Core enzyme
• Plus s-subunit for
initiation
Chain Elongation Reaction
• Very similar to DNA
polymerization
• Interacts with the DNA
template over a short
distance, then DNA double
helix reforms
• No 3’5’ exonuclease
Genes
• 5’ 3’ (downstream and upstream)
• Coding strand vs. template strand
• Sense strand vs. antisense strand
Initiation
• Promoter—region of DNA that serves as
site for transcription initiation
• Consensus region
– TATA box
– -35 region
• Strong promoter matches consensus region
better—efficient transcription of rRNA
• Weak promoters—infrequent transcription
Initiation
• Promoter binding is relatively fast and
selective
• role of s subunit
– Decreases selectivity
for nonspecific sequences
• Initiation is slow step
– DNA unwinding
– Synthesis of primer
• Elongation is fast
Nonspecific binding
One-dimensional promoter search
Holoenzyme and promoter form closed complex
Open complex allows initiation
Sigma subunit released; promoter is
cleared and elongation starts
Termination
• Termination sequences
– Pause site
– Rho dependent
Differences in Eukaryotic Transcription
• Multiple RNA polymerases based on
compartment and job
Differences in Eukaryotic Transcription
• General Transcription Factors—complex
promoter binding
Differences in Eukaryotic Transcription
• Role of Chromatin
– 35% of DNA is transcribed
– Most of it is quiescent in any given cell
– Buried in chromatin
– Nontranscribed genes are relatively inaccessible
– “Remodelling” enzymes make genes accessible
(for example, HATS)
Differences in Eukaryotic Transcription
• mRNA processing
• Makes primary transcript into “mature”
transcript
• PolyA tail makes RNA less susceptible to
exonucleases
• “Splicing” of exons together, with removal of
introns
– Mature RNA much shorter than primary transcript
Answers
1.
2.
3.
4.
5.
D
A
B
B
C