Transcription-Gene Expression

BSC205-Introduction to Molecular Biology

Dr. Saadia Naseem
BIO240
COMSATS University Islamabad
 Transcription

The synthesis of RNA molecules using

DNA strands as the templates so that
the genetic information can be
transferred from DNA to RNA is called
TRANSCRIPTION.
 Section 1
Operon: Fine control of
Bacterial Transcription
 Recognition of Origins
• Each transcriptable region is called
operon.
• One operon includes several structural
genes and upstream regulatory
sequences (or regulatory regions).
• The promoter is the DNA sequence that
RNA-pol can bind. It is the key point
for the transcription control.
 Promoter

regulatory
structural gene
sequences
5' 3'
promotor
RNA-pol
3' 5'
The lac Operon
• The lac operon was the first operon discovered
• Contains 3 genes coding for E. coli proteins that permit
the bacteria to use the sugar lactose
• Galactoside permease which
transports lactose into the cells
- b-galactosidase cuts the lactose into
galactose and glucose
• Galactoside transacetylase whose
function is unclear
Genes of the lac Operon
• Genes are grouped:
• lacZ = b-galactosidase
• lacY = galactoside permease
• lacA = galactoside transacetylase

• All 3 genes are transcribed together

producing 1 mRNA, a polycistronic message
that starts from a single promoter

• Each cistron, or gene, has its own ribosome binding site

• Each cistron can be transcribed by separate ribosomes that

bind independently of each other
 Section 2

Transcription Process
 General concepts
• Three phases: initiation, elongation, and termination.
• The prokaryotic RNA-pol can bind to the DNA
template directly in the transcription process.
• The eukaryotic RNA-pol requires co-factors to bind to
the DNA template together in the transcription
process.
§2.1 Transcription of Prokaryotes

• Initiation phase: RNA-pol recognizes the promoter and

starts the transcription.
• Elongation phase: the RNA strand is continuously
growing.
• Termination phase: the RNA-pol stops synthesis and the
nascent RNA is separated from the DNA template.
Transcription bubble
Transcription of Eukaryotes
Three RNA Polymerases in Eukaryotic cells
Transcription of Eukaryotes
a. Initiation
• Transcription initiation needs promoter and upstream
regulatory regions.
• The cis-acting elements are the specific sequences on
the DNA template that regulate the transcription of
one or more genes.
 Cis-acting element

cis-acting element
structural gene
GCGC CAAT TATA
exon intron exon

start
TATA box (Hogness box)

enhancer CAAT box

GC box
 Transcription factors
• RNA-pol does not bind the promoter directly.
• RNA-pol II associates with six transcription factors,
TFII A - TFII H.
• The trans-acting factors are the proteins that
recognize and bind directly or indirectly cis-acting
elements and regulate its activity.
 RNA
Polymerase II
requires a set
of
Transcription
factors
TF for eukaryotic transcription
 Pre-initiation complex (PIC)
• TBP of TFII D binds TATA
• TFII A and TFII B bind TFII D
• TFII F-RNA-pol complex binds TFII B
• TFII F and TFII E open the dsDNA (helicase and
ATPase)
• TFII H: completion of PIC
Pre-initiation complex (PIC)

RNA pol II

TF II F TF II E
TF II TBP TAF
TF II
A TATA B
TF II H DNA
 Phosphorylation of RNA-pol
• TF II H is of protein kinase activity to phosphorylate
CTD of RNA-pol. (CTD is the C-terminal domain of
RNA-pol)
• Only the p-RNA-pol can move toward the
downstream, starting the elongation phase.
• Most of the TFs fall off from PIC during the elongation
phase.
 Transcription initiation by
RNA polymerase II in a eukaryotic cell
 Comparison of the
steps leading from gene to protein in
eukaryotes and bacteria
A comparison of the structures of prokaryotic and
eukaryotic mRNA molecules.
 Section 3
Post-Transcriptional
Modification
• The nascent RNA, also known as primary transcript,
needs to be modified to become functional tRNAs,
rRNAs, and mRNAs.
• The modification is critical to eukaryotic systems.
BIO240-Introductory Mol. Biology
§3.1 Modification of hnRNA
• Primary transcripts of mRNA are called as
heteronuclear RNA (hnRNA).
• hnRNA are larger than matured mRNA by
many folds.
• Modification includes
• Capping at the 5- end
• Tailing at the 3- end
• mRNA splicing
• RNA edition
 a. Capping at the 5- end
• After the mRNA transcript is about 25-30
nucleotide long a 7-methylguoanosine is
added to the 5’ end.
• The capping enzyme associates with the
phosphorylated CTD of RNA Polymerase
II.
• Protect the 5’ from enzymatic
degradation in the nucleus and assist in
export to the cytosol
 a. Capping at the 5- end
OH OH
O
N
NH
O O O
O 5'
H2N N N H2C O P O P O P O CH 2 N NH 2
N
5' O
O O O
HN
N
O
CH 3
O OH
Pi
O P O AAAAA-OH 3'
O

m7GpppGp----
 ppp5'NpNp
removing
Pi phosphate group
pp5'NpNp
GTP forming 5'-5'
triphosphate group
PPi
G5'ppp5'NpNp

methylating at G7

7
m GpppNpNp
methylating at C2' of the
first and second
nucleotides after G
7
m Gpppm2'Npm2'Np
• The 5- cap structure is found on hnRNA too.  The
capping process occurs in nuclei.
• The cap structure of mRNA will be recognized by the
cap-binding protein required for translation.
• The capping occurs prior to the splicing.
b. Poly-A tailing at 3 - end
• Primary transcript is cleaved and a poly A tail is added
to the free 3’ OH
• Up to 250 A residues may be added
• Carried out by poly(A) polymerase
• Protects the mRNA from degradation.
Poly-A tailing at 3 - end
• There is no poly(dT) sequence on the DNA template. 
The tailing process dose not depend on the template.
• The tailing process occurs prior to the splicing.
• The tailing process takes place in the nuclei.

BIO240-Introductory Mol. Biology

c. mRNA splicing

mRNA

DNA

The matured mRNAs are much shorter than

the DNA templates.
 mRNA splicing
Introns are removed and the exons are spliced
togather
Splice sites occur at both the 5’ and 3’ ends
of introns
The intron is cut at the splice sites
Only 30-40 nucleotides at each end of an intron
are required for splicing.
The splicing reaction is catalyzed by a large
protein complex called the spliceosome
assembled from protein and small nuclear RNA
molecules.
 Split gene

The structural genes are composed of coding and non-

coding regions that are alternatively separated.

7 700 bp
L 1 2 3 4 5 6 7
A B C D E F G

A~G no-coding region 1~7 coding region

 Exon and intron

Exons are the coding sequences that

appear on split genes and primary
transcripts, and will be expressed to
matured mRNA.

Introns are the non-coding sequences

that are transcripted into primary
mRNAs, and will be cleaved out in the
later splicing process.
mRNA splicing outline
d. mRNA editing

• Taking place at the transcription level

• One gene responsible for more than one proteins
• Significance: gene sequences, after post-
transcriptional modification, can be multiple purpose
differentiation.
§3.2 Modification of tRNA

BIO240-Introductory Mol. Biology

 1. Precursor transcription

DNA

TGGCNNAGTGC GGTTCGANNCC

RNA-pol III

tRNA precursor
2. Cleavage

RNAase P

BIO240-Introductory Mol. Biology

endonuclease

ligase
3. Addition of CCA-OH

tRNA nucleotidyl

BIO240-Introductory Mol. Biology

transferase

ATP ADP

4. Base modification
§3.3 Modification of rRNA

• 45S transcript in nucleus is the precursor of 3 kinds

of rRNAs.
• The matured rRNA will be assembled with ribosomal
proteins to form ribosomes that are exported to
cytosolic space.
 rRNA

18S 5.8S 28S 45S-rRNA

transcription

splicing
18S-rRNA
5.8S and 28S-rRNA