Topic 10:

Transcription
Dr. E. B. Chaponda
Introduction
• Transcription is the synthesis of RNA using information from
DNA.
• This process ensures that synthesis of proteins using
information from DNA is properly regulated.
• Because of this intermediary process, proteins are
synthesised in the right place in the right amounts at the
right time.
Mechanism of transcription
• Transcription involves the following stages:
i. Initiation
ii. Elongation
iii. Termination
Initiation

• Before transcription can take place the following must occur;

i. the DNA double helix of the gene to be transcribed has to
unwind.
ii. the two DNA polynucleotide chains have to separate
(unzip) in order to expose the nucleotide bases.
Initiation

• To begin transcribing a gene, RNA polymerase binds to the DNA of the

gene at a region called the promoter.
• Basically, the promoter tells the polymerase where to attach on the
DNA and begin transcribing.
Initiation
• Each gene (or, in bacteria, each group of genes transcribed together)
has its own promoter.
• A promoter contains DNA sequences that let RNA polymerase or its
helper proteins attach to the DNA.
• Once the transcription bubble has formed, the polymerase can start
transcribing.
Initiation

• Promoters in bacteria
A typical bacterial promoter contains two important DNA sequences,
the 10 and 35 elements.
RNA polymerase recognizes and binds directly to these sequences.
Once the RNA polymerase has bound, it can open up the DNA and
get to work.
Figure 1: Promoter site in bacteria. The -10 and the -35 elements. These get their
names because they come 35 and 10 nucleotides before the initiation site (+1 in the
DNA). The minus signs just mean that they are before, not after, the initiation site.
Elongation
• Once RNA polymerase is in position at the promoter,
elongation—can begin.
• During elongation, RNA polymerase moves along one strand
of DNA, the template (sense) strand, in the 3' to 5' direction.
• For each nucleotide in the template, RNA polymerase adds a
complementary RNA nucleotide to the 3' end of the RNA
strand.
Transcription
• RNA polymerase catalyses the formation of phosphodiester
bonds between adjacent ribonucleotides using
complementary base pairing (A to U, T to A, C to G, and G to
C).
• The other strand which is not transcribed is called the
coding (anti-sense) s t ra n d

• T h e c o d i n g s t ra n d has the same code as the RNA

transcript except for T in place of U.
Figure 2: Transcription – Elongation: Addition of nucleotides to the three prime end of the
mRNA
Termination
• There are two types of transcription termination
mechanisms.
i. rho (ρ)-independent termination
ii. ρ-dependent termination
Termination
• rho (ρ)-independent termination of transcription, occurs at specific
base sequences called palindromic sequences in the RNA molecule
(AAAGGCUCC-UUUU-GGAGCCUUU).

A strand is said to have a palindromic sequence if the sequence of

nucleotides is the same as the reverse of its complement

• When the palindromic sequence is transcribed, it forms a hair-pin

loop which disturbs the RNA polymerase and results in termination of
transcription
Termination

Figure 3: Example of a palindromic sequence

Termination
• Rho-independent termination depends on specific sequences in the
DNA template strand.
• As the RNA polymerase approaches the end of the gene being
transcribed, it hits a region rich in C and G nucleotides.
• The RNA transcribed from this region folds back on itself, and the
complementary nucleotides bind together.
• The result is a stable hairpin/stem loop that causes the polymerase to
stall.
Termination

Figure 4a : Rho-independent termination of transcription

Termination

Figure 4b: Rho-independent termination of transcription

Termination
• In a terminator, GC rich stem loop/hairpin structure is formed.
• There is weak pairing of A(DNA template): U(RNA transcript)
• Stem loop displaces A:U and forms more stable A(template):T(non-
template) base pairs.
• The DNA double helix “zips up” behind the RNA polymerase
• Dissociation of RNA transcript from DNA.
Termination
• The second mechanism is the ρ-dependent termination which occurs
in the following steps;
i. RNA polymerase reaches a termination signal on the DNA sequence
ii. the rho factor attaches to the ρ factor binding site on RNA and
moves up the transcript towards RNA polymerase at the
transcription bubble.
iii. the ρ factor unwinds the DNA-RNA hybrid helix using it’s ATP
dependent helicase activity.
iv. this releases the RNA molecule and ends transcription.
Figure 5: Rho-dependent termination
Posttranscriptional modification of
newly synthesized RNAs
• The newly synthesized RNAs (mRNA, tRNA and rRNA) are called
precursor molecules
• RNAs must be modified before they become mature and functional.
• These modifications mainly occur in eukaryotes
• Modification involves removal of some nucleotides and
transformation of other nucleotides.
Messenger RNA posttranscriptional
modification
• Precursor mRNA undergoes capping, polyadenylation and splicing
before it becomes mature mRNA.
• Capping is the addition of a methylated guanosine head on the 5´
end.
• The cap does the following;
i. protects mRNA from degradation
ii. helps in nuclear export
iii. assists in ribosome binding during translation
Messenger RNA posttranscriptional
modification
• Polyadenylation is the addition of poly (A) tail on the 3´ end:
The poly-A tail does the following;
i. protects mRNA from degradation
ii. aid in the export of mRNA from the nucleus to the
cytoplasm;
iii. aids in transcription termination
iv. helps in binding proteins involved in initiating translation.
Messenger RNA
posttranscriptional modification
• RNA transcripts in eukaryotes cannot leave the nucleus to
be translated until their introns are removed.
• Splicing is a process of removal of unnecessary parts of
RNA, the introns, and joining of necessary parts, the
exons, together.
• mRNA splicing is unique to eukaryotes (in prokaryotes
splicing though rare is found to occur in non-coding RNA
i.e usually in tRNA).
• In prokaryotes, the genetic material is small in quantity
and comprises entirely of coding genes.
Figure 6: Messenger RNA post-transcription modification
Ribosomal RNA posttranscriptional
modification
• Precursor rRNA undergoes methylation and shortening to form
mature rRNA.
• Mature rRNA combines with protein to produce ribosome.
• A complete ribosome has two subunits.
• The prokaryotic ribosome is 70s (30s + 50s) while the eukaryotic
ribosome is 80s (60s + 40s).
Figure 7: Formation of prokaryotic (a) and eukaryotic (b)
ribosomes.
Transfer RNA posttranscriptional
modification
• Precursor tRNA is converted into cloverleaf shaped mature tRNA
which has the following features:
i. D arm
ii. T arm
iii. Anticodon arm and
iv. Amino acid acceptor arm.
• Every tRNA has the CCA sequence at its 3′-terminus
• The CCA-3′ moiety is required for amino acid attachment onto the 3′-
end of the tRNA and for peptide-bond formation on the ribosome.
Figure 8: Structure of Transfer RNA
Functions of different RNA
molecules
• mRNA molecules carry the coding sequences for protein
synthesis and are called transcripts.
• rRNA molecules form the core of a cell's ribosomes (the structures in
which protein synthesis takes place)
• tRNA molecules carry amino acids to the ribosomes during protein
synthesis.
Transcription inhibiting drugs

• Transcription inhibitors can be used as antibiotics against, pathogenic

organisms.
• Examples of such drugs are;
i. rifampicin is an antibacterial drug which inhibits prokaryotic DNA
transcription
ii. 8-Hydroxyquinoline is an antifungal drug which is a transcription
inhibitor.
Summary
• The synthesis of RNA using information from DNA is called
transcription.
• The RNA molecule is intermediate between DNA and proteins.
• The transcription process involves initiation, elongation and
termination.
• The stretch of DNA that is transcribed into an RNA molecule is called
a transcription unit.
• The section of DNA that holds the information for one polypeptide is
called a gene.
Summary
• Only one of the DNA strands is used as a template for the
synthesis of RNA, the sense DNA strand.
• The other strand which is not transcribed is called the anti-
sense/coding s t ra n d
• T h e a n t i s e n s e s t ra n d has the same code as the RNA
transcript except for T in place of U.