At the end of this
lecture, students will be able to –

• Discuss the process of transcription in

– Prokaryotes

– Eukaryotes

• Explain post transcriptional modification of mRNA

• Explain the mechanism, functions, and various classes of
transcription factors


• Transcription in Prokaryotes

• Transcription in Eukaryotes

Transcription factors

• Sequence specific DNA binding factors

• Protein that binds to specific DNA sequence

• Control the flow of genetic information from DNA to mRNA

• They perform function alone or with other protein in

• Act either by promoting or blocking the recruitment of RNA
polymerase to specific gene


• Process in which cells build proteins from information in
a DNA gene in a two major steps:

I-Transcription and


• Transcription:
Synthesis of an RNA (mRNA) that is complementary to one of the strands of DNA

• Translation:
Ribosomes read a messenger RNA and make protein according to its instruction


• RNA polymerase copies both the exons and the introns.

• Stretch of DNA that is transcribed into an RNA molecule a
transcription unit

• A transcription unit contains coding sequence that is
translated into protein and sequences that direct and regulate protein

• Transcription proceeds in the 5′ → 3′ direction

of RNA polymerase

• The holoenzyme is a complete RNA polymerase consisting of
an core enzyme and a sigma factor

• The core enzyme consists of 5 polypeptide chains

• Two α subunits, one β and β1 subunit and ώ subunit, σ factor

of RNA polymerase

• Having helicase activity for unwinding.

• Not requires primer

• Lacks proof reading

• σ – Recognition of promoter with the help of transcription

• α – activator

• β – phosphodiester bond

• β – DNA template


RNA polymerase;
in prokaryotes only the single enzyme          

• RNA polymerase governs the synthesis of all cellular RNAs

• In eukaryotic nuclei contain three RNA polymerases   

• RNA polymerase I is found in the nucleolus     

• The other two polymerases are located in the nucleoplasm    

• The three nuclear RNA polymerase have different roles In

• Polymerase I makes a large precursor to the major rRNA
(5.8S, 18S and 28S rRNA in vertebrates)

• Polymerase II synthesizes hnRNAs, which are precursors to
mRNAs and small nuclear RNAs (snRNAs)

• Polymerase III synthesize the precursor to 5SrRNA, the
tRNAs and several other small cellular and viral RNAs

in prokaryotes

Promoter recognition:

• Sigma factor interacts with core enzyme at β subunit site
to check transcription of both the strands by core enzyme

• The holoenzyme transcribes only one of two strands.

• Sigma factor of holoenzyme recognizes the promoter region
of the DNA

in prokaryotes

Centered at -10 to -35 bp from the transcription start point

Promoters in
3 different promoters

-25                             -40                       -110

T A T A                     GC                     CAAT

Binding of RNA
Promoters have binding site for proteins rather than RNA

• Most common binding site – a complex of cyclic AMP
receptor protein

Unwinding of DNA
double helix:
Binding of ώ factor results in unwinding of a double helix

• Open complex allows tight binding of the RNA polymerase
with subsequent initiation of RNA synthesis

of first base of RNA chain

• The base of RNA synthesized is always in the form of
purine i.e. triphosphate guanine (ppp G) or adenine (ppp A)

• Initiation of mRNA synthesis does not require primer
Initiation ends after the formation of first inter nucleotide bond.


• Core enzyme moves along the template from 3-5 end untwisting the helix bit by bit and adding
one complementary nucleotide

• After 8-9 bp of RNA synthesis occurs, sigma factor is
released and recycled for other reaction

• RNA polymerase completes the transcription at 30-50 bp/sec

• Unwinding and rewinding of DNA occurs simultaneously



Two types of terminator sequence occur in prokaryotes

1. Type 1(ρ-

• RNA molecule terminated without the aid of the rho factor
contain GC rich sequence followed by U residues

• GC region makes RNA to spontaneously fold into hairpin
loop that tends to pull the RNA away from DNA

• The weaker bonds between the sequence of   U residues and DNA template broken releasing
the newly formed RNA molecule

2. Type 2 (rho-

• RNA molecule that do not form GC rich hairpin loop
requires rho factor for Termination

• It is a hexameric factor which binds to specific
termination sequence 50-90 bases located near 3
end of newly forming RNA molecule

• It acts as an ATP – dependent unwinding enzyme, unwinds RNA
from DNA template as it proceeds

Transcription in eukaryotes Initiation  

• RNA polymerase cannot recognize the promoters, requires
general transcription factor (GTFs)

• Promoters forms pre intiation complex with GTFs

• Assembly of the proteins to TATA box forms TATA binding
protein (TBP)

• TBP is present as a subunit of much larger protein complex
called TFІІD which specially binds to TATA box


• TFІІB provide a binding site for RNA polymerase

• TFІІF contains subunit homologous to the bacterialσ
factor, bounds to the entering polymerase

• TFІІH contains 10 subunits, 3 possess enzymatic activity
helps in unwinding the DNA (helicase activity)


• Same as prokaryotes

• Involves sequential addition of nucleotide units


• Transcription by RNA polymerase І is terminated by a
protein factor that recognizes an 18-nucleotide termination signal

• Termination signals for RNA polymerase III include short
run of Us (as in prokaryotic signal). No proteins factors are needed for their

transcription processing of mRNA

• Post-transcriptional modification is a process by which,
in eukaryotic cells, primary transcript RNA is converted into mature RNA

• Conversion of precursor messenger RNA into mature
messenger RNA (mRNA), which includes splicing and occurs prior to protein

• The pre-mRNA molecule undergoes three main modifications

• 5′ capping

• 3′ polyadenylation

• RNA splicing – occur in the cell nucleus before the RNA is

• The 5′ capping:
5 end chemically
modified by the addition of 7 methylguanosine

• Replacement of triphosphate group at the 5′ end of the RNA
chain with a special nucleotide GMP nucleotide

3’ adenylation

• Addition of poly A tail to 3’ end

• Added before it leaves the nucleus

• AAUAA sequence recognized by a specific endonuclease that
cleaves the RNA around 20 nucleotide down stream

• Poly A tail associated with protein, retard action of 3’-


• RNA splicing – introns are removed from the pre-mRNA

• Remaining exons connected to re-form a single continuous

• Catalyzed by a large protein complex called the spliceosome

• Allows production of a large variety of proteins from a
limited amount of DNA

of transcription factors

• Stabilize or block the binding of RNA polymerase to DNA

• Catalyse acetylation or deacetylation of histone

• Histone acetyl transferase activity – acetylates histone

• Histone deacetylases activity – decetylates histone

• Recruit coactivators or corepressor proteins to the
transcription factor-DNA complex

of transcription factors 

• Reads and interprets the genetic “blue print” in the DNA

• Bind to DNA and initiate program of increase or decrease gene

• Basal transcription regulation

– General transcription factors necessary for transcription
to occur

– They interact with polymerase directly

of transcription factors 

enhancement of transcription

• Regulate the expression of various gene by binding to
enhancer region of DNA adjacent to regulated gene

• Ensure that genes are expressed in right cell at the right

Response to
intracellular cells

• Transcription factors are involved in the downstream of
signalling cascade

Response to

• Involved in downstream of signaling cascade in
environmental stimuli

• Heat shock factors, hypoxia inducible factors

Cell cycle control

• Proto-oncogenes or tumor suppressor gene regulate cell

• Myc oncogene – in cell growth & apoptosis

of transcription factors 


• Transcription factors are transcribed from gene on a
chromosome into RNA, then RNA to proteins

• Any of these steps can be regulated to affect the
production of transcription factors

Nuclear localization

• Transcribe in nucleus but not translated in cytoplasm

• They have nuclear localization signals that direct them to


• Transcription factors can be activated or inactivated by
through signal sensing domain

• Ligand binding –
influence factors present in cell; decided if factors are in active state or
capable to bind DNA

Phosphorylation –
STAT protein must be phosphorylated before they can bind to DNA

• Interaction with other transcription factors

Accessibility of DNA
binding site

• DNA in nucleosome is inaccessible to many transcription

• Nucleosome should be actively removed by molecular motors

Availability of other
cofactors/ transcription factors

• Most transcription factors do not work alone

• For transcription many factors must bind to DNA regulatory

• Recruitment of intermediary proteins

Classes of
transcription factors 

Classified based on –

• Mechanism of action

• Regulatory function

• Sequence homology in their DNA binding domain

class of transcription factor 

• General
transcription factors

– Involved in the formation of pre initiation complex


– Ubiquitous, interact with core promoter region

• Upstream
transcription factors

– Binds upstream to initiation site

– Stimulate or repress transcription

class of transcription factors

Constitutively active

• Present in all cells at all time – general transcription
factors Sp1, NF1, and CCAAT

Conditionally active
– requires activation

• Developmental – cell specific

• Signal – dependent – requires external signal for

class of transcription factor 

• Based on sequence similarity and tertiary structure of
DNA- binding domain

• 1- superclass – Basic domains

• 2 – Superclass – Zinc co-ordinating DNA binding domain

• 3 Superclass – Helix-turn-helix

• 4 superclass- beta-scaffold factors with minor groove

• 5 superclass – other transcription factors


• Protein synthesis in prokaryotes and eukaryotes occurs
through two steps – Transcription and Translation

• Process of transcription involves 3 steps – initiation,
elongation and termination

• Process of transciption in similar in prokaryotes and
eukaryotes but differs in post transcriptional modification

• Post transcriptional modifications include addition of 5’
cap, 3’ adenylation and RNA splicing

• Transcription factors are sequence specific DNA binding factors
and protein that binds to specific DNA sequence

• Stabilize or block the binding of RNA polymerase to DNA

• They reads and interprets the genetic “blue print” in the

• Also bind to DNA and initiate program of increase or
decrease gene transcription

• Transcription factor is classified based on mechanism of
action, regulatory function and sequence homology