Transcription

11/24/2014

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TRANSCRIPTION

Department of Biochemistry, Kathmandu Medical College Monday, November

24, 2014

1 Rajesh Chaudhary | @bloggerjesh

Monday, November 24, 2014 2 Rajesh Chaudhary | @bloggerjesh

Monday, November 24, 2014 3 Rajesh Chaudhary | @bloggerjesh Monday, November 24, 2014 4

Prokaryotes vs Eukaryotes Prokaryotes

rRNA: 23S, 16S, 5S

Eukaryotes rRNA: 28S, 18S, 5.8S and 5S

Rajesh Chaudhary | @bloggerjesh

Introduction of RNA

How is genetic information is executed?

Types of RNA

mRNA

tRNA

rRNA

Does our body have just these RNA or some more?

What is the role of RNA? (Structural, Catalytic)

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Transcription


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Catalytic RNAs are also known as RIBOZYMES.

What is the functional difference between DNA and

RNA?

What are the differences and similarities between

replication and transcription?

Entire structure or part of structure?


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Transcription / RNA synthesis


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Similarities

Fundamental chemical mechanism

Its polarity (direction of synthesis)

Use of template

Similarities in phages (initiation, elongation and termination)

Differences

Use of primer

Segment

What is cellular transcriptome?


Transcription


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DNA template is copied in the 3’ 5’ direction

Synthesis of new RNA molecule = 5’ 3’

Process of transcription

Initiation Elongation Termination


RNA polymerase


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Core enzyme vs

Holoenzyme

Termination factor


Steps in RNA synthesis


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How long is a transcription unit? (I.e. Its extension)

What is the name of the product formed after

transcription?

Initiation

RNA binding at promoter


Initiation of transcription


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Pribnow box

-35 sequence

Mutation in either the Pribnow box or the -35 sequence can affect the transcription of the gene controlled by mutant promoter!


Elongation in transcription 12

Monday, November 24, 2014 Rajesh Chaudhary | @bloggerjesh

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Elongation in transcription


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Elongation in transcription


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Supercoils are disentangled by topoisomerase I and II.

RNA polymerase synthesizes short transcript (<11 nucleotides)

of DNA sequence.

Elongation begins when transcript is over 10 nucleotide in

length.

RNA polymerase has no known proofreading function.

1. How fast nucleotides are added during the elongation step?

2. What is the error rate in transcription process?


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1. Transcription is regulated at

several steps


Termination of transcription


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Termination

r-independent r-dependent


r-dependent termination


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r-factor: hexameric ATPase with helicase activity

Binds C-rich “rho” recognition site near 3’-end of the nascent

RNA using ATPase activity.

ATP-dependent RNA-DNA helicase activity of rho separates

the hybrid new RNA

What does ATPase do?

Rajesh Chaudhary | @bloggerjesh Monday, November 24, 2014 18

s cycle


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Action of antibiotics


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Antibiotics prevent bacterial growth by inhibiting RNA

synthesis.

E.g.: rifampin, Actinomycin D


Transcription in Eukaryotes


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Transcription in Prokaryotes Vs

Eukaryotes


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Differences

In terms of RNA polymerase

Eukaryotes: Pol I, Pol II and Pol III

a) Pol I: transcribes rRNA

b) Pol II: transcribes RNA that becomes mRNA

c) Pol III: transcribes tRNA & 5S RNA

Binding of polymerase is affected by enhancer sequence.

Similarities

Polymerase, dNTP, 3 steps of transcription


Chromatin structure and gene

expression


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DNA, histone, nucleosome

and chromatin – how are

they arranged?

Nucleosome Chromatin

Euchromatin Heterochromatin


Mechanism of chromatin remodeling


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Chromatin

Inactive chromatin Active chromatin

(Acetylation)

Enzyme: histone acetyl transferase

(Deacetylation)

Enzyme: histone deacetylase (restores +ve charge)


Eukaryotic gene promoter consensus

sequences


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Transcription factors recognize DNA through a variety of

DNA-binding motifs, such as zinc fingers and leucin zippers.


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Transcription factors


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Rajesh Chaudhary | @bloggerjesh Monday, November 24, 2014 26

Enhancer vs Silencer





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What is transcription factors? Why is it necessary?

Different transcription factor for different genes.

Coactivator: include the histone acetyltransferase

proteins involved in chromatin remodeling.


Role of enhancers in eukaryotic gene

regulation


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Special cis-acting element

Increases process of transcription by RNA polymerase II

Located on the same chromosome where gene is

located

Presence can be upstream or downstream to the gene


Possible locations of enhancer

sequence


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Inhibitors of RNA polymerase II

a-amanitin of Amanita phalloides


Post transcriptional modification


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Both prokaryotic and Eukaryotic tRNA and rRNA is

posttranscriptionally modified.

Prokaryotic mRNA is generally identical while eukaryotic

mRNA is extensively modified.

tRNA are further modified to give a unique identity to each

species.


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Post transcriptional modification of

rRNA


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ribonuclease


Post transcriptional modification of

tRNA


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nucleotidyltransferase


Post-transcriptional modification of

eukaryotic mRNA


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Primary transcripts are heterogeneous nuclear RNA (hnRNA).

Modification occurs in nucleus.

Modifications 1. 5’-capping

2. Addition of poly-A tail

3. Removal of introns Rajesh Chaudhary | @bloggerjesh

References


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Science

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