Transcript Transcription 111 - Tehran University of Medical Sciences

‫دانشگاه علوم پزشكي وخدمات بهداشتي‬
‫درماني تهران‬
Dr. Parvin Pasalar
Tehran University of Medical Sciences
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Objectives:
To know and explain:
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What are needed for transcription
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Template
Building blocks
Enzymes
Energy
Protein Factors
Different Stages of transcription
Initiation (promoter recognition)
Elongation
Termination
Differences between transcription in eu and prokaryotes
Toxins and antibiotics that inhibit transcription
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Prokaryotic Transcription
Cell
DNA
Transcription
Translation
mRNA
Ribosome
Polypeptide
(protein)
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What are needed for transcription
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Permission
Template (DNA, Gene)
Building blocks (4 types of NTPs)
Enzymes
Energy (ATP and the other NTPs)
Protein factors (σ & ρ)
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A
“Simple”
Prokaryotic
Gene
Transcription
Start Site
5’
3’
3’
5’
Protein Coding Region
Terminator
Sequence
Promoter/
Control Region
5’
AUG
UAA
RNA Transcript
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E. coli RNA polymerase
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Holoenzyme: 2 ’ 
Core enzyme: 2 ’
for initiation
for elongation
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Different stages of Transcription
 Recognition
& Initiation
 Elongation
 Termination
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Initiation: close and open complex
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The elongation stage
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RNA chain termination
Inverted repeats
Termination occurs at
terminator DNA sequences.
1. Rho-independent: the most
ρ- independent
common stop signal by
RNA hairpin formation(selfcomplement structure)
commonly GC-rich to favor
the structure stability
2. Rho-dependent Termination:
RNA wraps around Rho
hexamer and stops
transcription.
ρ- dependent
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Termination
Rho Independent
RNA
Pol.
RNA
Terminator
5’
RNA
Pol.
RNA
5’
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Termination
Rho Independent
RNA
Pol.
RNA
5’
5’
RNA
Pol.
RNA
Terminator
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Termination
Rho Dependent
Terminator
RNA
Pol.
RNA
5’
r
RNA
Pol.
RNA
5’
r
The terminator
sequence slows
RNA polymerase15
Termination
Rho Dependent
Terminator
RNA
Pol.
RNA
5’
r
Help, rho
hit me!
r
RNA
5’
RNA
Pol.
Rho catches up
with RNA
polymerase 16
Termination
Rho Dependent
Terminator
RNA
Pol.
RNA
5’
5’
r
The elongation
complex disintegrates
RNA
Pol.
RNA
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A “Simple” Eukaryotic Gene
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Eukaryotic gene
Cytoplasm
DNA
Transcription
RNA
RNA
Processing
mRNA G
G
AAAAAA
Nucleus
AAAAAA
Export
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Different Eukaryotic RNA polymerases
Basis of classification:
1- Elution at different salt concentrations during ion-exchange
chromatography
2- Different sensitivity to α amanitin
Pol I: very insensitive to α amanitin, transcribes the
gene for pre-rRNA (28S, 5.8S, 18S)
Pol II: very sensitive to α amanitin, transcribes all
protein coding genes and the gene for snRNA
Pol III: Intermediate sensitivity to α amanitin,
transcribes the gene for tRNA, 5S rRNA, U6, RNA
component of SRP
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Transcription factors
Definition:
Proteins other than purified RNA polymerase subunits, involved in
Initiation, Elongation or Termination of transcription
Classification:
A: General transcription factors (GTFs):
1- are needed for basal gene expression
2- are required for RNA polymerase to bind avidly and specifically to
the promoters.
3-GTFs for RNA polymerase II are called TFIIA , TFIIB , TFIID , …
4- can have multiple subunits
B: Specific transcription factors:
They are used for fine tune control of gene expression of specific
genes
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RNAP II termination and poly adenylation
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mRNA modification
The modifications signal the mRNA is ready to move out of
the nucleus and may control its life span in the cytoplasm
 1) 5’ cap: modified guanine; protection; recognition site for
ribosomes
 2) 3’ tail: poly(A) tail (adenine); protection; recognition;
transport
 3) RNA splicing: exons (expressed sequences) kept, introns
(intervening sequences) spliced out; spliceosome
 4) Alternative Splicing
 5) RNA editing
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Mechanism of RNA Splicing
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Alternative splicing
Calcitonin
gene-related
peptide
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Toxins and antibiotics that inhibit
transcription
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Actinomycine D: makes cross link between the
two strands of DNA, preventing it from being
template for transcription
Rifamycine ( Rifampin): inhibits beta subunit
of RNAP of prokaryotes
Ricin: A toxin from plant that inhibits RNAP
of eukaryotes
Amanitin: A toxin from mashroom that
inhibits RNAP of eukaryotes
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